Guidelines for the use and interpretation of assays for monitoring autophagy

In the paper

“This results in the preferential degradation of mitochondrial outer membrane proteins by the proteasome, while inner membrane proteins and mitochondrial DNA 558 remain intact.”

What the sources say

• Disputed

  In Results, under “p62/SQSTM1 is not essential for mitochondrial degradation” (page unavailable), the paper reports that mitochondrial DNA decreased after CCCP treatment, contrary to the claim that it remained intact. It identifies autophagic and lysosomal degradation of depolarized mitochondria; it does not report preferential proteasomal degradation of outer-membrane proteins while inner-membrane proteins remain intact. (LLM, conf=0.88)

  Checked by a reasoning model — may be wrong.

  cites [558] sourceExperimental — accuracy not yet measured

In the paper

“This post-absorptive release of amino acids is strongly inhibited by infusion of insulin or by exogenous supply of amino acids, suggesting that it is mainly mediated by a lysosomal/autophagic mechanism of protein catabolism.[643][644][645][646][647][648][649][650]”

What the sources say

• ?Not checked — no full text of the sourcecites [643] source
• ?Not checked — no full text of the sourcecites [644] source
• Disputed

  The Results (p. 5) report that amino-acid infusion did not significantly change outward amino-acid transport or intracellular amino-acid appearance, an index of proteolysis. The Discussion (pp. 6–7) says protein breakdown remained unchanged; net balance shifted from release to uptake because synthesis increased. The paper does not attribute that shift to inhibited lysosomal/autophagic catabolism or establish an effect of insulin infusion. (LLM, conf=0.88)

  Checked by a reasoning model — may be wrong.

  cites [645] sourceExperimental — accuracy not yet measured

In the paper

“tubule-specific ATG5-deficient mice 656 display a degeneration phenotype earlier than podocyte-specific ATG5-deficient mice, 657 suggesting that autophagy, and hence LC3 levels, might actually be more prominent in the former.”

What the sources say

• Partly supported

  The Results report early cellular changes in proximal tubule-specific Atg5-deficient mice, and the paper describes eventual tubular degeneration. They also report suppressed LC3-I-to-LC3-II conversion after Atg5 deletion. Our copy does not compare the timing of degeneration or LC3 levels with podocyte-specific Atg5-deficient mice, so it does not support those comparative claims. (LLM, conf=0.86)

  cites [656] sourceExperimental — accuracy not yet measured
• Disputed

  The section “Glomerular podocytes exert high levels of autophagy under basal conditions” (page not available) reports more detectable autophagosomes in podocytes than in other kidney cells and faster LC3-II accumulation in podocytes than in the tubular cells compared. This is contrary to the suggestion that autophagy and LC3 are more prominent in tubules. The paper reports late degeneration in podocyte-specific Atg5-deficient mice, but does not establish an earlier phenotype in proximal tubule-specific Atg5-deficient mice. (LLM, conf=0.86)

  Checked by a reasoning model — may be wrong.

  cites [657] sourceExperimental — accuracy not yet measured

In the paper

“In a limited number of cases, autophagy has been established as the cause of cell death; 97,199,[658][659][660][661][662][663][664] although opposite results have been reported using analogous experimental settings.665”

What the sources say

• Supported

  The citation is attached to the clause about opposite results, not to the preceding claim that autophagy causes cell death. The Discussion reports that inducing autophagy increased survival in cells expressing mutant huntingtin, while the Results report that interfering with p62 increased huntingtin-induced cell death. These findings support the cited contrast in a cell-death setting. (LLM, conf=0.86)

  cites [97] source
• Disputed

  The citation is attached to the claim that autophagy causes cell death, not to the later clause about opposite results. The Discussion (page unavailable) instead says this work supports the hypothesis that autophagy is required to maintain cellular viability; it reports accelerated senescence in autophagy-gene mutants. (LLM, conf=0.86)

  Checked by a reasoning model — may be wrong.

  cites [199] sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [658] source

In the paper

“Finally, plant root tips cultured in nutrient-sufficient medium display constitutive autophagic flux (i.e., a basal level), which is enhanced in nutrient-deprived medium.696,778,779”

What the sources say

• Disputed

  In the Results section on barley root tips (page unavailable), pilot experiments found no difference in autolysosome accumulation with versus without sucrose, contrary to the claimed enhancement in nutrient-deprived medium. The Discussion speculates about normal low-level autophagic activity but does not establish constitutive flux in nutrient-sufficient root tips. (LLM, conf=0.87)

  Checked by a reasoning model — may be wrong.

  cites [696] sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [778] source
• Supported

  The Addendum (pp. 360–362) states that autophagy occurs constitutively in plant root cells under nutrient-sufficient conditions and is activated under nutrient-limiting conditions. It also reports increased autophagosome numbers in Arabidopsis root tips cultured in sucrose-free medium. (LLM, conf=0.93)

  cites [779] source

In the paper

“Autophagy was first detected by TEM in the 1950s (reviewed in ref. 8).”

What the sources say

• Supported

  The Abstract says autophagy was first discovered by transmission electron microscopy and dates that discovery to the mid-1950s. The section on its discovery also describes TEM observations from 1954 and 1957. This supports the citing sentence’s use of the review. (LLM, conf=0.94)

  cites [8] source

In the paper

“The impact of autophagic flux on cell death and human pathologies therefore demands accurate tools to measure not only the current flux of the system, but also its capacity, 20 and its response time, when exposed to a defined insult.21”

What the sources say

• ?Not checked — no full text of the sourcecites [20] source
• Supported

  In “Autophagy and the Dynamics between Modes of Cell Death” (page not supplied), the paper recommends measuring basal autophagic flux and the time needed to reach maximal attainable activity. It also discusses autophagic capacity and the time needed to respond to a stress stimulus, supporting the attributed need to assess flux, capacity, and response time. (LLM, conf=0.90)

  cites [21] source

In the paper

“a putative advantage of mCherry over mRFP is its enhanced photostability and intensity, which are an order of magnitude higher (and comparable to GFP), enabling acquisition of images at similar exposure settings as are used for GFP, thus minimizing potential bias in interpretation.182”

What the sources say

• ?Could not verify

  The Introduction discusses red fluorescent proteins generally and attributes brightness and photostability to DsRed-Express2, but does not compare mCherry with mRFP. Searches for mCherry, the claimed order-of-magnitude difference, and GFP-like imaging exposure settings found no passage addressing those claims. Our copy therefore cannot verify the attributed comparison or its proposed effect on interpretation. (LLM, conf=0.75)

  cites [182] sourceExperimental — accuracy not yet measured

In the paper

“Dendra2 is an improved version of the green-to-red photoswitchable fluorescent protein Dendra, which is derived from the octocoral Dendronephthya sp.184”

What the sources say

• ?Could not verify

  The Methods sections on GFP–Hsp70 construction and confocal microscopy describe an EGFP fusion protein and LysoTracker staining, not Dendra2. Searches for Dendra2, Dendronephthya and photoconvertible coral proteins found no passage in our copy addressing Dendra2’s relationship to Dendra or its origin. (LLM, conf=0.75)

  cites [184] sourceExperimental — accuracy not yet measured

In the paper

“It is also possible to use anti-LC3/Atg8 antibodies for immunocytochemistry or immunohistochemistry, 140,[204][205][206][207][208][209] procedures that have the advantages of detecting the endogenous protein, obviating the need for transfection and or the generation of a transgenic organism, as well as avoiding potential artifacts resulting from overexpression”

What the sources say

• ?Not checked — no full text of the sourcecites [140] source
• ?Not checked — no full text of the sourcecites [204] source
• ?Not checked — no full text of the sourcecites [205] source
• Supported

  Split into 2 atoms — 2 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [206] source
• Supported

  Split into 2 atoms — 2 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [207] source
• ?Not checked — no full text of the sourcecites [208] source
• Partly supported

  Split into 2 atoms — 1 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [209] sourceExperimental — accuracy not yet measured

In the paper

“Another alternative to mRFP or mCherry is to use the Venus variant of YFP, which is brighter than mRFP and less sensitive to pH than GFP.186”

What the sources say

• ?Could not verify

  The DNA constructions and LC3-II localization sections describe a GFP–LC3 construct and its use in microscopy. Searches for Venus/YFP, mRFP, brightness, and pH sensitivity found no passage comparing these fluorescent proteins, so our copy does not address the attributed claim. The indexed passages provide no page numbers. (LLM, conf=0.78)

  cites [186] sourceExperimental — accuracy not yet measured

In the paper

“a caution concerning the use of the EGFP fluorescent protein for microscopy is that this fluorophore has a relatively neutral pH optimum for fluorescence, 180 so that its signal may diminish quickly during live cell imaging due to the acidic environment of the lysosome.”

What the sources say

• Supported

  The Results state that GFP has a pKa of 6.0 and that its fluorescence is attenuated in acidic lysosomes. The Discussion says GFP-LC3 signals disappear rapidly in the lysosomal environment and explains that protonation can attenuate fluorescence in less than 1 ms. The paper uses pEGFP-LC3 constructs; the cited passages do not provide page numbers. (LLM, conf=0.91)

  cites [180] source

In the paper

“The pH optimum of EGFP is important to consider when using GFP-LC3 constructs, as the original GFP-LC3 marker 187 uses the EGFP variant, which may result in a reduced signal upon the formation of amphisomes or autolysosomes.”

What the sources say

• Partly supported

  The DNA and siRNA Transfection section (page unavailable) identifies the construct used in this study as EGFP-LC3, supporting the claim that a GFP-LC3 construct uses EGFP. The paper does not describe it as the original GFP-LC3 marker or discuss EGFP’s pH optimum or reduced signal in amphisomes or autolysosomes. (LLM, conf=0.83)

  cites [187] sourceExperimental — accuracy not yet measured

In the paper

“Late/degradative autophagic vacuoles and autolysosomes (AVd) usually have only one limiting membrane, and contain cytoplasmic material and/or organelles at various stages of degradation (Fig. 2A and B).43,47”

What the sources say

• Partly supported

  The Autophagic Pathways section defines AVd as containing partially degraded cytoplasmic material and says AVd can be amphisomes or autolysosomes. The copy does not establish that AVd usually have only one limiting membrane; the Figure 2 description in Fusion Events with Endosomes and Lysosomes describes an AVd with both an inner and an outer membrane. Page numbers are not available in the indexed passages. (LLM, conf=0.85)

  cites [43] sourceExperimental — accuracy not yet measured
• Partly supported

  The section on leupeptin-induced alterations (page unavailable) reports that hepatocytic lysosomes contained autophagocytosed cytoplasm at various stages of degradation. The Discussion (page unavailable) identifies secondary lysosomes with autophagic contents as autolysosomes. Our copy does not state that these vacuoles usually have only one limiting membrane; it specifies double or multiple membranes for autophagosomes instead. (LLM, conf=0.85)

  cites [47] sourceExperimental — accuracy not yet measured

In the paper

“TORC1 is downstream of AKT1; however, oxidative stress inhibits MTOR, thus allowing autophagy, despite the concomitant activation of AKT1.115”

What the sources say

• Partly supported

  The Abstract (page unavailable) reports that increased oxidative stress after stable PINK1 silencing activates mitophagy, an autophagy-based process. Neither that passage nor the Discussion passages establish that oxidative stress inhibits MTOR, that TORC1 is downstream of AKT1, or that AKT1 is activated concurrently. (LLM, conf=0.78)

  cites [115] sourceExperimental — accuracy not yet measured

In the paper

“ATG16L1 has been used to monitor the movement of plasma membrane as a donor for autophagy, and thus an early step in the process; ATG16L1 is located on phagophores, but not on completed autophagosomes.269,411”

What the sources say

• ?Could not verify

  The Introduction discusses GFP-Atg8p and LC3 as autophagy markers, and the paper reports vacuolar turnover in yeast. Searches for ATG16L1, phagophores, and plasma membrane as an autophagy membrane donor found no passage addressing ATG16L1’s use or its location on developing versus completed autophagosomes. (LLM, conf=0.76)

  cites [269] sourceExperimental — accuracy not yet measured
• Supported

  In Results, “Plasma membrane contributes to autophagosome precursors” (page not specified), the paper uses Atg16L1-positive vesicles to trace plasma-membrane-derived vesicles entering early autophagosome precursors. In Results, “Influence of clathrin-mediated endocytosis on Atg16L1-positive autophagosome precursors” (page not specified), it describes Atg16L1-positive maturing structures and says the Atg16L1 complex is removed when autophagosomes form. (LLM, conf=0.93)

  cites [411] source

In the paper

“ATG5, ATG12, and ATG16L1, associate with the phagophore and have been detected by fluorescence or immunofluorescence (Fig. 16).400,401”

What the sources say

• Partly supported

  In “Apg16L localizes to autophagic isolation membrane” (page not provided), the paper reports that the Apg12–Apg5 conjugate associates with isolation membranes and shows fluorescently tagged Apg5 and Apg16L co-labeling those membranes. Fig. 8 also documents immunofluorescence detection of Apg16L. The paper does not show Apg12 individually detected there by fluorescence or immunofluorescence. (LLM, conf=0.86)

  cites [400] sourceExperimental — accuracy not yet measured
• Partly supported

  The Results sections “Apg5 Colocalizes with Punctate LC3 Structures” and “Apg5 Localizes to Isolation Membranes” report GFP-Apg5 on isolation membranes and anti-Apg12 staining overlapping its punctate signals. These observations support the ATG5 and ATG12 portion, but our copy does not report fluorescence or immunofluorescence detection of ATG16L1 at the phagophore. (LLM, conf=0.88)

  cites [401] sourceExperimental — accuracy not yet measured

In the paper

“Furthermore, in some cells ULK1 has functions in addition to autophagy, such as in axonal transport and outgrowth, and its activity state may thus reflect its role in these processes.[390][391][392][393][394][395]”

What the sources say

• Supported

  The Discussion section identifies mouse Unc51.1 as ULK1 and links it to neurite outgrowth. It also argues that autophagy does not underlie the axonal and dendritic abnormalities studied. In the Results section on axonal transport defects (page unavailable), wild-type Unc-51 rescued vesicle accumulations whereas a kinase-deficient form did not, supporting the qualified point about activity. (LLM, conf=0.90)

  cites [390] source
• Partly supported

  The Results section reports that Unc-51 has roles beyond autophagy, including a likely role in axonal transport; the Discussion also notes its requirement for axon outgrowth in worms and mice. The paper does not establish that ULK1’s activity state reflects those roles, so that part remains an inference rather than a finding of this source. No page numbers are available in our copy. (LLM, conf=0.82)

  cites [391] sourceExperimental — accuracy not yet measured
• Partly supported

  The paper identifies ULK1 among kinases implicated in axonal events and in its screen of neurite outgrowth and retraction (opening summary; Discussion, page not available). It does not establish a role for ULK1 in axonal transport or test whether ULK1's activity state reflects a role in these processes. (LLM, conf=0.85)

  cites [392] sourceExperimental — accuracy not yet measured

In the paper

“ATG14-GFP or BATS-GFP detected by fluorescence microscopy or TEM can be used as a phagophore marker; however, ATG14 is not localized exclusively to phagophores, as it can also be detected on mature autophagosomes as well as the ER.409,410”

What the sources say

• Partly supported

  The Results (page unavailable) use GFP-tagged BATS in fluorescence microscopy and show that BATS puncta colocalize with Atg16, a phagophore marker. The Results also describe ATG14L association with the ER. They do not show ATG14 on mature autophagosomes or demonstrate GFP-tagged ATG14/BATS as a marker detected by TEM; TEM is used to examine autophagic vacuoles. (LLM, conf=0.85)

  cites [409] sourceExperimental — accuracy not yet measured
• Partly supported

  The Results and discussion (page unavailable) show GFP-Atg14L puncta partially colocalizing with Atg16L, a phagophore marker, and show Atg14L at the ER. They do not establish that Atg14L is present on mature autophagosomes: LC3 marks both phagophores and completed autophagosomes. The passages do not support BATS-GFP or TEM as a phagophore-marker method. (LLM, conf=0.86)

  cites [410] sourceExperimental — accuracy not yet measured

In the paper

“Finally, in Arabidopsis and possibly other eukaryotes, the ATG1 and ATG13 proteins are targets of autophagy, which means that their levels may drop substantially under conditions that induce autophagic turnover.174”

What the sources say

• ?Could not verify

  The Introduction (page unavailable) describes the Atg1/Atg13 complex as a regulator of autophagy in budding yeast, not as a target of autophagic degradation. Searches for Arabidopsis and for declining ATG1/ATG13 protein levels found no passage addressing the claimed turnover. (LLM, conf=0.75)

  cites [174] sourceExperimental — accuracy not yet measured

In the paper

“In Arabidopsis and some mammalian cells it appears that essentially all of the ATG5 and ATG12 proteins exist in the conjugated form and the expression levels do not change, at least during short-term starvation.150,[400][401][402]”

What the sources say

• ?Could not verify

  The paper’s description of the conjugation cascades says ATG12 attaches to ATG5, but gives no fraction of either protein present in that form. Its nutritional-stress results concern maize ATG transcripts and ATG8, not ATG5–ATG12 protein levels in Arabidopsis or mammalian cells during short-term starvation. Searches for the conjugated fraction and starvation-related changes did not locate a passage addressing the claim. (LLM, conf=0.72)

  cites [150] sourceExperimental — accuracy not yet measured
• Partly supported

  The section “Apg12-Apg5 and Apg16L form a ~800 kDa protein complex” (page not provided) says that almost all Apg5 in wild-type mouse ES cells was conjugated to Apg12. It does not establish that almost all Apg12 was conjugated, address that proportion in Arabidopsis, or report unchanged ATG5 and ATG12 expression during short-term starvation. Its statement that complex formation was unaffected by nutrient conditions concerns the complex, not protein expression levels. (LLM, conf=0.87)

  cites [400] sourceExperimental — accuracy not yet measured
• Partly supported

  The Results sections “Production of Apg5-deficient Cells” and “Apg5 Colocalizes with Punctate LC3 Structures” report that nearly all Apg5 in the mouse embryonic stem-cell transformants was conjugated with Apg12 and that the total amount of the conjugate did not change significantly after amino-acid starvation. The paper does not establish the same findings for Arabidopsis, nor does these passages show that essentially all Apg12 was conjugated or that the expression levels of both individual proteins were unchanged. (LLM, conf=0.87)

  cites [401] sourceExperimental — accuracy not yet measured

In the paper

“Yeast Atg14 is the autophagy-specific subunit of the Vps34 complex I, 404 and a human homolog, named ATG14 (ATG14L/BARKOR), has been identified.[405][406][407][408]”

What the sources say

• Supported

  The Results section (‘Apg14p and Vps38p Constitute Distinct Complexes’) identifies the Apg14p-containing Vps34 complex as complex I and says it functions in autophagy, while the distinct Vps38p-containing complex functions in CPY sorting. Citation 404 follows the yeast clause; the human-homolog clause carries separate citations [405]–[408]. (LLM, conf=0.92)

  cites [404] source
• ?Not checked — no full text of the sourcecites [405] source
• Partly supported

  The passage describing protein identification (section labeled Abstract; page unavailable) reports a newly identified mammalian protein with modest similarity to yeast Atg14 and names it Atg14L. However, the protein was isolated from mouse tissues, and our copy does not identify it as a human homolog or use the name BARKOR. The yeast-complex point is also described in the paper, but the citation in question follows the human-homolog point. (LLM, conf=0.84)

  cites [406] sourceExperimental — accuracy not yet measured

In the paper

“It should be noted that there are two isoforms of WIPI2 (2B and 2D), 421 and in C. elegans WIPI4 (EPG-6) has been identified as the WIPI homolog required for autophagy.422”

What the sources say

• ?Could not verify

  Citation 422 is attached to the C. elegans clause. The paper’s Discussion addresses human WIPI-1 and WIPI-2, but the passages found do not identify WIPI4/EPG-6 or establish its requirement for autophagy in C. elegans (searched: C. elegans WIPI4 EPG-6; EPG-6 WIPI-4 Caenorhabditis elegans). The WIPI2 isoform clause has a separate citation, 421. (LLM, conf=0.76)

  cites [421] sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [422] source

In the paper

“For this purpose, the WatershedCounting3D plug-in for ImageJ may be useful.211,212”

What the sources say

• ?Could not verify

  The passages available from our copy are lists of links with no section or page labels. Searches for WatershedCounting3D, ImageJ, and objective puncta counting found no passage addressing whether the plug-in is useful for obtaining a puncta count per cell. (LLM, conf=0.69)

  cites [211] sourceExperimental — accuracy not yet measured
• ?Could not verify

  The Fluorescence microscopy section describes scoring macrophages by their Atg7-labelled structures, but does not mention ImageJ or the WatershedCounting3D plug-in. Searches for the plug-in, ImageJ, and automated puncta counting found no passage addressing whether it is useful for obtaining an objective count per cell. Page numbers were not available in our copy. (LLM, conf=0.76)

  cites [212] sourceExperimental — accuracy not yet measured

In the paper

“ZFYVE1 partially colocalizes with WIPI1 upon nutrient starvation, 421 and also with WIPI2.418”

What the sources say

• Supported

  The results passage accompanying Fig. 4C reports that DFCP1 (ZFYVE1) and WIPI-1 partially colocalized upon nutrient starvation. This source is cited for the WIPI1 clause; the WIPI2 clause has a separate citation. (LLM, conf=0.90)

  cites [421] source
• ?Not checked — no full text of the sourcecites [418] source

In the paper

“Although a population of BECN1 may localize in proximity to the trans-Golgi network, 433 it is also present at the ER and mitochondria.425”

What the sources say

• Supported

  The Localization of Beclin and PtdIns 3-kinase section reports that Beclin colocalized with trans-Golgi network markers and concludes that it localizes to the TGN. Citation 433 is attached only to the TGN clause; the ER and mitochondria clause carries a different citation. (LLM, conf=0.94)

  cites [433] source
• ?Not checked — no full text of the sourcecites [425] source

In the paper

“The elimination of DRAM1 by siRNA blocks autophagy, 439,440 as effectively as elimination of BECN1, indicating it is an essential component for this process, although its mechanism of action is not known.”

What the sources say

• Partly supported

  The Results section reports that siRNA knockdown of DRAM and BECN1 each prevented UV-induced degradation of VRK1 in WS1 fibroblasts. The Discussion says DRAM’s role is not yet known. The paper does not show that DRAM knockdown blocks autophagy generally, or establish that it does so as effectively as BECN1 knockdown across the process. (LLM, conf=0.88)

  cites [439] sourceExperimental — accuracy not yet measured
• Partly supported

  In the section “Evidence for the interplay between JNK and p53-DRAM signalling pathways in the regulation of autophagy” (page unavailable), the paper says DRAM silencing or depletion prevents 2-methoxyestradiol-induced autophagy in Ewing sarcoma cells. It does not specify siRNA in the passages available, and that context-specific result does not establish that DRAM1 elimination blocks autophagy generally. The paper also leaves DRAM’s mechanism of action open. (LLM, conf=0.85)

  cites [440] sourceExperimental — accuracy not yet measured

In the paper

“Transcriptional upregulation of Drosophila Atg8a and Atg8b is also observed in the fat body following induction of autophagy at the end of larval development, 453 Atg5, Atg6, Atg8a and Atg18 are upregulated in the ovary of starved flies, 454 and an increase in Drosophila Atg8b is observed in cultured Drosophila l(2)mbn cells following starvation (Gorski S, personal communication).”

What the sources say

• Partly supported

  Split into 3 atoms — 1 confirmed, 2 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [453] sourceExperimental — accuracy not yet measured
• Partly supported

  Split into 3 atoms — 1 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 1 unverifiable. Expand the claim to see per-atom verdicts.

  cites [454] sourceExperimental — accuracy not yet measured
• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites Gorski S, personal communication

In the paper

“and in C. elegans, the FOXA transcription factor PHA-4 regulates the expression of several autophagy-related genes.458”

What the sources say

• Supported

  The Discussion reports that increased expression of several autophagy genes in germline-less C. elegans depends on the FOXA transcription factor PHA-4. The Results likewise identify PHA-4 as a transcriptional regulator of autophagy genes in glp-1 animals. (LLM, conf=0.93)

  cites [458] source

In the paper

“Increased expression of Atg5 in vivo after optic nerve axotomy in mice 446 and increased expression of Atg7, Becn1 and Lc3a during neurogenesis at different embryonic stages in the mouse olfactory bulb are also seen.447”

What the sources say

• Supported

  Citation 446 attaches to the first clause, about Atg5 after optic nerve axotomy in mice. The Results report that Atg5 mRNA expression was induced in axotomized retinas; the separate olfactory-bulb claim is attached to citation 447. (LLM, conf=0.94)

  cites [446] source
• Supported

  Citation 447 follows the olfactory-bulb clause, not the separate optic-nerve-axotomy clause marked 446. The Results describe measuring autophagy-gene expression in mouse olfactory bulbs at different embryonic stages, and the Discussion reports progressive upregulation of Atg7, Becn1 and LC3 during the E13.5–E15.5 neurogenic period. (LLM, conf=0.91)

  cites [447] source

In the paper

“and an autophagy-dedicated microarray was developed as a high-throughput tool to simultaneously monitor the transcriptional regulation of all genes involved in, and related to, autophagy.444”

What the sources say

• Supported

  The Results and Discussion section (page not supplied) says the authors designed and produced a human autophagy-dedicated microarray to simultaneously analyze expression of the whole repertoire of genes involved in or related to autophagy. This supports the claim's description of the tool and its purpose. (LLM, conf=0.94)

  cites [444] source

In the paper

“It is detected as a subpopulation in the Golgi and cis-Golgi, colocalizing with GOLGB1/giantin and GOLGA2/ GM130, and also in early and late endosomes and lysosomes, colocalizing with EEA1 and LAMP2.439”

What the sources say

• Partly supported

  The Results section, “A subpopulation of DRAM colocalizes in Golgi apparatus and endosome vesicles with VRK1” (page unavailable), reports partial colocalization with giantin and colocalization with GM130, EEA1, and LAMP2. It identifies EEA1 as an early-endosome marker and LAMP2 as a lysosomal marker, but does not establish localization in late endosomes. Figure 3 describes these observations in cells transfected to express DRAM. (LLM, conf=0.86)

  cites [439] sourceExperimental — accuracy not yet measured

In the paper

“Autophagy is critical for Drosophila midgut cell death, which is accompanied by transcriptional upregulation of all of the Atg genes tested, including Atg8a (Fig. 18).199,451”

What the sources say

• ?Could not verify

  The Results passages concern ATG8 and ATG4 expression and autophagy in Arabidopsis, including induction under nitrogen starvation, not Drosophila midgut cell death. Searches for Drosophila midgut cell death and Atg8a transcription found no passage addressing the claim in our copy. (LLM, conf=0.76)

  cites [199] sourceExperimental — accuracy not yet measured
• Partly supported

  The Abstract says that disrupting autophagy severely delays Drosophila larval midgut destruction and that several Atg genes are transcriptionally upregulated during midgut cell death. It does not say that all Atg genes tested were upregulated or identify Atg8a as one of them; it describes Atg8a as a marker of autophagic puncta. (LLM, conf=0.86)

  cites [451] sourceExperimental — accuracy not yet measured

In the paper

“It is important to be aware, however, that certain forms of macroautophagy are induced in a BECN1-independent manner, and are not blocked by PtdIns3K inhibitors.97,431”

What the sources say

• ?Could not verify

  The Results section (‘p62 bodies are degraded by autophagy’; page unavailable) reports that 3-methyladenine inhibits a step of autophagy and causes p62 accumulation. Our copy does not address whether any forms of macroautophagy are BECN1-independent or resist PtdIns3K inhibitors (searched: BECN1-independent, PtdIns3K inhibitors, 3-methyladenine). The p62 finding does not rule out the existence of other forms. (LLM, conf=0.74)

  cites [97] sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [431] source

In the paper

“Interestingly, caspase-mediated cleavage of BECN1 inactivates BECN1-induced autophagy and enhances apoptosis in several cell types, 432”

What the sources say

• Partly supported

  The ‘Crosstalk between apoptosis and autophagy by Beclin 1’ section says caspase cleavage destroys Beclin 1’s pro-autophagic activity and produces fragments that sensitize cells to apoptotic signals. It gives HeLa cells as one example, but does not establish the claimed effect across several cell types. Our copy provides no page number for this section. (LLM, conf=0.82)

  cites [432] sourceExperimental — accuracy not yet measured

In the paper

“Autophagy is induced by the release of BECN1 from BCL2 by pro-apoptotic BH3 proteins, phosphorylation of BECN1 by DAPK (at Thr119, located in the BH3 domain), 426 or phosphorylation of BCL2 by MAPK8/JNK1 (at Thr69, Ser70 and Ser87).427,428”

What the sources say

• Supported

  The Introduction states that DAPK phosphorylates beclin 1 at Thr119 in its BH3 domain. The Discussion links that phosphorylation to dissociation from Bcl-XL and induction of autophagy. Citation 426 is attached to this DAPK mechanism, not the separately cited JNK1 mechanism. (LLM, conf=0.94)

  cites [426] source
• Supported

  The passage labeled Abstract reports that JNK1 phosphorylates Bcl-2 at T69, S70 and S87, causing Bcl-2 to dissociate from Beclin 1 and stimulating autophagy. This supports the BCL2 phosphorylation mechanism to which this source's citation is attached; the DAPK mechanism has a separate citation. (LLM, conf=0.94)

  cites [427] source
• ?Not checked — no full text of the sourcecites [428] source

In the paper

“DRAM1 (damage-regulated autophagic modulator 1) is a gene induced by activated TP53/p53 in response to different types of cellular stress, including DNA damage.438,439”

What the sources say

• Supported

  The Results report that several genotoxic agents induced DRAM in control cells but not when p53 was silenced, demonstrating p53-dependent induction in response to cellular stress, including DNA-damaging stress. The paper calls the gene DRAM rather than DRAM1; this does not change the substance of the claim. Page numbers are not provided in our copy. (LLM, conf=0.93)

  cites [438] source
• Partly supported

  The Results section, “DRAM gene expression is induced in response to DNA damage” (page not provided), reports that several DNA-damaging agents activated DRAM expression alongside p53 accumulation, and that wild-type p53 induced DRAM expression. The copy does not establish DRAM induction in response to other types of cellular stress; its reference to several types of stress concerns autophagy more generally. (LLM, conf=0.85)

  cites [439] sourceExperimental — accuracy not yet measured

In the paper

“Detection of DRAM1 RNA is very easy by qRT-PCR during autophagy; 438,439 however, detection of the DRAM1 protein is very difficult because of its small size and hydrophobicity, which makes generation of specific antibodies a complicated process, and in general these have very low sensitivity.”

What the sources say

• Partly supported

  The citation is attached to the RNA-detection clause. The Results report measuring DRAM mRNA by real-time quantitative RT-PCR after p53 induction, but do not establish that detection is “very easy” or specifically describe qRT-PCR detection during autophagy. The later protein-detection clause follows the citation and is not assessed here. (LLM, conf=0.78)

  cites [438] sourceExperimental — accuracy not yet measured
• Partly supported

  The citation follows the RNA-detection clause. The qRT-PCR Methods describe detecting human DRAM RNA, and the Results report measuring DRAM gene expression by qRT-PCR. The paper does not characterize that detection as “very easy”; the protein-detection clause comes after the citation. (LLM, conf=0.82)

  cites [439] sourceExperimental — accuracy not yet measured

In the paper

“Although less work has been done on post-transcriptional regulation, several studies implicate microRNAs in controlling the expression of proteins associated with autophagy.164,[468][469][470]”

What the sources say

• ?Not checked — no full text of the sourcecites [164] source
• ?Not checked — no full text of the sourcecites [468] source
• Supported

  The Discussion reports that miR-93 strongly decreased SQSTM1 protein expression and states that SQSTM1 is involved in transporting proteins destined for autophagy. It concludes that SQSTM1 expression is post-transcriptionally regulated by miRNAs, making this paper an example of the studies described in the citing sentence. (LLM, conf=0.93)

  cites [469] source

In the paper

“Even increases in LC3 mRNA can be quite modest and are cell type-and organism-dependent.471”

What the sources say

• Partly supported

  The ‘LC3 as a Potential Biomarker for Autophagy’ passage in Detection of Starvation-induced Autophagy reports weak LC3 mRNA upregulation—up to threefold—in starved HepG2 cells. It does not establish that LC3 mRNA increases vary by cell type or organism; the other cell and mouse observations concern LC3 protein rather than mRNA. (LLM, conf=0.86)

  cites [471] sourceExperimental — accuracy not yet measured

In the paper

“For example, yeast pexophagy utilizes the processing of Pex14-GFP and Pot1 (thiolase)-GFP, 494,495”

What the sources say

• ?Not checked — no full text of the sourcecites [494] source
• Partly supported

  The Results section (‘The MAPK Slt2p is required for pexophagy’; page unavailable) describes monitoring yeast pexophagy through cleavage of Pot1p-GFP, identifying Pot1p as thiolase. Our copy does not describe processing Pex14-GFP, so it supports only the Pot1-GFP part of the claim. (LLM, conf=0.88)

  cites [495] sourceExperimental — accuracy not yet measured

In the paper

“In addition, yeast mitophagy requires both the Slt2 and Hog1 signaling pathways; the activation and phosphorylation of Slt2 and Hog1 can be monitored with commercially available phospho-specific antibodies (Fig. 19).381”

What the sources say

• Supported

  The Introduction states that both Slt2 and Hog1 are required for yeast mitophagy. The Results (Fig. 5) show their activation and phosphorylation monitored with phospho-specific antibodies, and Materials and methods identifies Cell Signaling Technology as the supplier of both antibodies. (LLM, conf=0.94)

  cites [381] source

In the paper

“Finally, CEBPB/C/EBPβ is a transcription factor that regulates autophagy in response to the circadian cycle.467”

What the sources say

• Supported

  The Introduction identifies C/EBPβ as a transcription factor linking autophagy to the circadian pacemaker. The section on physiological regulation of autophagy in the liver reports that C/EBPβ knockdown disrupts circadian regulation of autophagy genes and alters diurnal autophagy markers. (LLM, conf=0.94)

  cites [467] source

In the paper

“VMP1 is an essential autophagy gene that is conserved from Dictyostelium to mammals, 232,462 and the VMP1 protein regulates early steps of the autophagic pathway.420”

What the sources say

• ?Could not verify

  The passages labeled Abstract discuss p62/SQSTM1 and its possible role in autophagy, not VMP1. Searches for VMP1, its conservation from Dictyostelium to mammals, and its role in early autophagy steps found no passage addressing the claim; the available passages provide no page numbers. (LLM, conf=0.75)

  cites [232] sourceExperimental — accuracy not yet measured
• Supported

  The citation to this paper follows the claim about VMP1 regulating early steps; the conservation and essentiality claim has separate citations. In “Mammalian VMP1, EI24, and mEPG5 Are Required for Starvation-Induced Autophagy,” the authors explicitly conclude that VMP1 regulates an early step of the autophagy pathway. (LLM, conf=0.95)

  cites [462] source
• ?Not checked — no full text of the sourcecites [420] source

In the paper

“A gene regulatory network, named CLEAR (coordinated lysosomal enhancement and regulation) that controls both lysosome and autophagosome biogenesis was identified using a systemsbiology approach.[463][464][465]”

What the sources say

• Partly supported

  The passage discussing CLEAR (section labeled Abstract; page unavailable) calls it the “Coordinated Lysosomal Expression and Regulation” network, not “enhancement and regulation.” The paper reports that TFEB co-regulates lysosome and autophagosome biogenesis, but it treats CLEAR as an existing network and does not say it identified the network using a systems-biology approach. (LLM, conf=0.88)

  cites [463] sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [464] source
• ?Not checked — no full text of the sourcecites [465] source

In the paper

“TFEB activity is regulated by starvation, 463 and can thus serve as a new tool for monitoring transcriptional regulation connected with autophagy.”

What the sources say

• Supported

  The citation follows the clause that TFEB activity is regulated by starvation. The paper reports that nutrient starvation rapidly induces TFEB nuclear translocation and links TFEB activity to nutrient levels (passages 1–2). The subsequent suggestion that TFEB could serve as a monitoring tool is the citing authors’ inference, not the cited claim. (LLM, conf=0.93)

  cites [463] source

In the paper

“In this case the relative amount of the free GFP and the complete fusion protein is the relevant parameter for quantification; although it may not be possible to detect clear changes in the amount of the full-length chimera, especially under conditions of limited flux.29,36”

What the sources say

• Partly supported

  Split into 2 atoms — 1 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 1 unverifiable. Expand the claim to see per-atom verdicts.

  cites [29] sourceExperimental — accuracy not yet measured
• Partly supported

  Split into 2 atoms — 1 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [36] sourceExperimental — accuracy not yet measured

In the paper

“Along these lines, the erythroid transcription factor GATA1 and its coregulator Friend of GATA1 induce the transcription of multiple genes encoding autophagy components; this developmentally regulated transcriptional response is coupled to increases in autophagosome number as well as the percent of cells that contain autophagosomes.466”

What the sources say

• Supported

  Split into 2 atoms — 2 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [466] source

In the paper

“In cells, it is also possible to measure the release of an unlabeled amino acid by chromatographic methods, thereby obviating the need for prelabeling; 490 however, it is important to keep in mind that amino acid release is also regulated by protein synthesis, which in turn is modulated by many different factors.”

What the sources say

• ?Could not verify

  The available indexed passages are an IRIS catalog entry (labeled Abstract, no page number), not the paper’s substantive text. Searches for chromatographic measurement of unlabeled amino-acid release and for regulation of release by protein synthesis found no passage addressing either point, so the claim cannot be verified from our copy. (LLM, conf=0.75)

  cites [490] sourceExperimental — accuracy not yet measured

In the paper

“GFP-Vid24, Vid30-GFP and Sec28-GFP colocalize with actin during prolonged glucose starvation and for up to 30 min following glucose replenishment in wild-type cells; however, colocalization is less obvious by the 60 min time point.540,543”

What the sources say

• Supported

  The Vid30-GFP results (Fig. 3B; page unavailable) report colocalization with actin patches during glucose starvation and through 30 minutes after glucose re-feeding, with less at 60 minutes. The GFP-Vid24 results (Fig. 7A) report the same pattern in wild-type cells. The paper also reports that Sec28 associates with actin patches during starvation and through 30 minutes after re-feeding, with less colocalization at 60 minutes; it attributes those Sec28 findings to an earlier study. (LLM, conf=0.88)

  cites [540] source
• ?Not checked — no full text of the sourcecites [543] source

In the paper

“In addition, mutants such as vac8D and tlg2D accumulate prApe1 under rich conditions, but not during autophagy.380,532”

What the sources say

• Partly supported

  The section “Vac8p Is Not Essential for Autophagy” reports negligible prAPI delivery in vac8Δ cells in nitrogen-containing medium and near-wild-type delivery during starvation. The paper names tlg2 among mutants primarily defective in one pathway or the other, but the passages do not establish the claimed rich-medium versus autophagy pattern for tlg2Δ. Page numbers are unavailable in our copy. (LLM, conf=0.87)

  cites [380] sourceExperimental — accuracy not yet measured
• Partly supported

  The sections “API maturation is the major biosynthetic trafficking defect in tlg2∆ cells” and “Rapamycin-induced autophagy bypasses the API transport block in tlg2∆ cells” report blocked API maturation in tlg2∆ cells under ordinary conditions and rapid, near-complete proAPI maturation when autophagy is induced. The cited paper does not report the corresponding result for vac8∆ mutants, so it supports only the tlg2∆ example. (LLM, conf=0.88)

  cites [532] sourceExperimental — accuracy not yet measured

In the paper

“Vid vesicles can be purified by fractionation and gradient centrifugation; western blotting analysis using antibodies against organelle markers and Fbp1, and the subsequent verification of fractions by EM facilitate their identification.539”

What the sources say

• Supported

  The Results section, “Purification of the FBPase-associated Vesicles,” describes fractionation followed by two sucrose gradients, immunoblotting with antibodies against FBPase and organelle markers to distinguish the FBPase-containing fractions, and EM examination of the isolated fractions. EM showed a near-homogeneous population of vesicles, supporting the claimed identification procedure. (LLM, conf=0.94)

  cites [539] source

In the paper

“Immunodetection of LC3 as definite puncta is possible in paraffin-embedded tissue sections and fresh frozen tissue, by either immunohistochemistry or immunofluorescence; 140,[634][635][636][637] however, this methodology has not received extensive evaluation, and does not lend itself well to dynamic assays.”

What the sources say

• ?Not checked — no full text of the sourcecites [140] source
• Partly supported

  Split into 2 atoms — 0 confirmed, 2 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [634] sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [635] source

In the paper

“In addition, multiple disease states have been associated with an altered activity of the ubiquitin-proteasome system, further complicating the use of the amino acid exchange rate as a marker of autophagy.654”

What the sources say

• Partly supported

  The Main section links UPS malfunction to several disease categories, including cancer, cardiovascular disease and neurodegenerative disorders. It does not discuss amino acid exchange rate as a marker of autophagy or say that altered UPS activity complicates its use. (LLM, conf=0.86)

  cites [654] sourceExperimental — accuracy not yet measured

In the paper

“Compared with classical biochemical approaches, PCP allows the proteome-wide recording of protein gradient profiles.”

What the sources say

• Partly supported

  The Results section (page unavailable) describes density-gradient fractionation and mass-spectrometric measurement of protein distributions across fractions. It does not identify the method as PCP or establish the claimed proteome-wide scope or comparison with classical biochemical approaches. (LLM, conf=0.82)

  cites [278] sourceExperimental — accuracy not yet measured

In the paper

“Twelve h of fasting in mice may be sufficient to trigger autophagy in muscle, 639,640 but the appropriate time should be determined empirically.”

What the sources say

• Supported

  The Results section (‘Mitochondria elongate upon induction of autophagy’) states that autophagy was induced in vivo by fasting mice for 12 hours and reports changes in their muscle mitochondria. This supports the hedged claim that a 12-hour fast may be sufficient; the recommendation to determine the appropriate time empirically is the citing authors’ advice. (LLM, conf=0.92)

  cites [639] source
• ?Not checked — no full text of the sourcecites [640] source

In the paper

“Bone marrow transfer has been used to document in vivo the role of autophagy in the reverse cholesterol transport pathway from peripheral tissues or cells (e.g., macrophages) to the liver for secretion in bile and for excretion, 638”

What the sources say

• Partly supported

  The Results section, “Lipophagy Contributes to RCT In Vivo” (page unavailable), reports that autophagy-deficient macrophages showed reduced cholesterol transport to plasma, liver, gallbladder, and feces. But the Experimental Procedures, “In Vivo RCT Studies” (page unavailable), describe injecting labeled macrophages into mice, not bone marrow transfer. The in vivo role is supported; the named method is not. (LLM, conf=0.88)

  cites [638] sourceExperimental — accuracy not yet measured

In the paper

“Chloroquine suppresses the immunological response in a manner that is not due to its pHdependent lysosomotropic accumulation (chloroquine interferes with lipopolysaccharide-induced TNF/TNFa gene expression by a nonlysosmotropic mechanism), 651 as well as through its pHdependent inhibition of antigen presentation.652”

What the sources say

• Supported

  The citation to Weber and Levitz is attached to the nonlysosomotropic mechanism, not to the antigen-presentation claim, which has a separate citation. In “Pretranslational effect of CQ occurs by a nonlysosomotropic mechanism,” the paper reports that reducing pH-dependent chloroquine accumulation did not prevent its suppression of LPS-stimulated TNF-α mRNA in human PBMC. (LLM, conf=0.92)

  cites [651] source
• ?Not checked — no full text of the sourcecites [652] source

In the paper

“This quantitative technique is capable of monitoring the fusion of autophagosomes with lysosomes, that is, the formation of an autolysosome, and the assay does not depend on the analysis of LC3.”

What the sources say

• Supported

  The SIGNIFICANCE section says the quantitative Keima technique was designed to detect autophagosome–lysosome fusion. The section on EGFP-LC3 and Keima probes says Keima records autolysosome formation; the paper also describes its use where conventional LC3-based probes are ineffective. (LLM, conf=0.94)

  cites [549] source

In the paper

“For example, cellular viability is related to high CTSB activity and low CTSD/ cathepsin D activities.642 Therefore, the appearance of the opposite levels of activities may be one indication of the initiation of autophagy (lysosome)-dependent cell death.”

What the sources say

• Partly supported

  In serum-deprived PC12 cells, the paper reports that cathepsin B activity falls while cathepsin D activity rises during cell death (III-1, p. 6), and describes a cathepsin-regulated death pathway downstream of autophagy (III-2, p. 10). But it does not establish high B and low D as a general condition for cellular viability: PC12 cells overexpressing B had improved survival even though their cathepsin D protein amount increased (Fig. 7, p. 10). (LLM, conf=0.86)

  cites [642] sourceExperimental — accuracy not yet measured

In the paper

“In addition, cleavage of GFP-LC3 to generate free GFP can be evaluated.This has been successfully performed in mouse liver, 172,631 suggesting the GFP-LC3 cleavage assay may also be applied to in vivo studies.”

What the sources say

• ?Not checked — no full text of the sourcecites [172] source
• Supported

  The Results section (‘Acute ethanol exposure induced autophagy,’ Fig. 1C) reports a single GFP moiety produced by breakdown of GFP-LC3 in ethanol-treated mouse livers. This directly supports the claim that GFP-LC3 cleavage can be evaluated in vivo. (LLM, conf=0.93)

  cites [631] source

In the paper

“Similarly, adenoviral-mediated expression of the transcription factor CEBPB/C/EBPβ induces autophagy in hepatocytes.467”

What the sources say

• Supported

  In “C/EBPβ induces autophagy gene expression and protein degradation,” adenoviral expression of C/EBPβ in primary hepatocytes increased LC3-II and enhanced protein degradation that was sensitive to the autophagy inhibitor 3-MA. The authors conclude that C/EBPβ is sufficient to enhance autophagic protein degradation. (LLM, conf=0.94)

  cites [467] source

In the paper

“There are fewer compounds that act as inducers of autophagy, but the initial characterization of this process was due in large part to the inducing effects of glucagon, which appears to act through indirect inhibition of MTOR via the activation of STK11/LKB1-AMPK.615,616,761”

What the sources say

• Partly supported

  The Introduction (p. C11) calls glucagon a powerful inducer of autophagy in liver, and the results (p. C12) report numerous autophagic vacuoles in glucagon-injected rats compared with controls. This supports glucagon’s role in the early study of autophagy, but the paper does not address MTOR inhibition or STK11/LKB1–AMPK activation. (LLM, conf=0.86)

  cites [615] sourceExperimental — accuracy not yet measured
• Partly supported

  The Introduction (p. 437) says the discovery that glucagon induces autophagic vacuoles opened the way to studying autophagy under reproducible, timed conditions. The paper does not address the proposed STK11/LKB1–AMPK or MTOR mechanism, so it supports the historical and inducing-effects portion but not that mechanistic explanation. (LLM, conf=0.88)

  cites [616] sourceExperimental — accuracy not yet measured
• Partly supported

  The Results and Discussion report that glucagon represses mTOR signaling in perfused rat liver and is associated with phosphorylation of LKB1 and AMPK, supporting the proposed indirect pathway. Our copy does not address glucagon-induced autophagy or the claim that glucagon was important to autophagy’s initial characterization. (LLM, conf=0.88)

  cites [761] sourceExperimental — accuracy not yet measured

In the paper

“For example, this can be achieved either through overexpression or post-translational activation of the gene encoding TFEB (see Transcriptional and translational regulation), a transcriptional regulator of the biogenesis of both lysosomes and autophagosomes.463,464”

What the sources say

• Supported

  The Abstract-labeled passages report that TFEB overexpression increases autophagosome formation and autophagic flux, while TFEB drives expression of lysosomal and autophagy genes. They also report that phosphorylation regulates TFEB activity and that a phosphorylation-site mutant more strongly induces the autophagic–lysosomal system. Page numbers are not provided in our copy. (LLM, conf=0.92)

  cites [463] source
• ?Not checked — no full text of the sourcecites [464] source

In the paper

“Along these lines ICP34.5, viral BCL2 homologs and viral CFLAR/FLIP of herpesviruses block autophagosome formation, 425,580,759”

What the sources say

• ?Not checked — no full text of the sourcecites [425] source
• ?Not checked — no full text of the sourcecites [580] source
• Supported

  In the passages labeled Abstract, the paper reports that herpesvirus vFLIP suppresses autophagy and describes FLIP as inhibiting autophagosome elongation. This supports the viral FLIP portion of the multi-citation sentence; the citation need not establish its separate ICP34.5 claim. (LLM, conf=0.90)

  cites [759] source

In the paper

“MTOR is a major regulatory protein that is part of the insulin signaling pathway, and it controls processes other than autophagy, so that rapamycin will ultimately affect many metabolic pathways.379,[762][763][764]”

What the sources say

• Partly supported

  The Introduction (page not provided) describes Tor as a nutrient-responsive regulator and rapamycin as its inhibitor; it also reports effects beyond autophagy, including cell-cycle arrest and glycogen accumulation. The Discussion identifies mammalian Tor (mTOR), but our copy does not place it in the insulin signaling pathway or establish that rapamycin affects many metabolic pathways in mammals. (LLM, conf=0.86)

  cites [379] sourceExperimental — accuracy not yet measured
• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites [762][763][764]

In the paper

“Clearly claim (b) is stronger than claim (a), and needs to be justified by proof that inhibiting autophagy, through either genetic or chemical means, prevents cell death.669”

What the sources say

• Supported

  The ‘Definition of Autophagic Cell Death’ section says that cell death mediated by autophagy must be suppressible by chemical or genetic inhibition of the autophagic pathway. It also says that cell death showing autophagy markers but not blocked by such inhibition should not be classified as autophagic cell death. The comparison between claims (a) and (b) is the citing authors’ framing. (LLM, conf=0.94)

  cites [669] source

In the paper

“however, a substantial amount of neuronal cell death occurs during neurogenesis, making it problematic to carry out a correct analysis in vivo or ex vivo.677,678”

What the sources say

• ?Not checked — no full text of the sourcecites [677] source
• ?Could not verify

  The indexed passages are repository file listings, not passages from the paper; they have no section or page numbers. Searches for neuronal death during neurogenesis and for difficulties analyzing it in vivo or ex vivo returned no substantive text, so the claim cannot be checked against our copy. (LLM, conf=0.76)

  cites [678] sourceExperimental — accuracy not yet measured

In the paper

“The activation of the lysosomal two pore channel (TPC), by nicotinic acid adenine dinucleotide phosphate (NAADP) induces autophagy, which can selectively be inhibited by the TPC blocker NED-19, or by pre-incubation with BAPTA, showing that lysosomal calcium also modulates autophagy.244”

What the sources say

• ?Could not verify

  Split into 2 atoms — 0 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 2 unverifiable. Expand the claim to see per-atom verdicts.

  cites [244] sourceExperimental — accuracy not yet measured

In the paper

“Autophagy may also be regulated by the release of calcium from the endoplasmic reticulum under stress conditions; 212,726,766”

What the sources say

• ?Could not verify

  The Introduction links autophagy to cellular stress and discusses several regulators, but does not mention calcium release. The section on secretory traffic discusses vesicles from the endoplasmic reticulum, not calcium signaling. Searches for calcium release and ER-stress regulation found no passage addressing the claimed mechanism. (LLM, conf=0.76)

  cites [212] sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [726] source
• ?Not checked — no full text of the sourcecites [766] source

In the paper

“In vivo treatment of embryos with cadmium results in an increase in autophagy, probably to counter the stress, allowing cell survival through the elimination/ recycling of damaged structures.632”

What the sources say

• Supported

  The paper reports higher autophagy in cadmium-treated sea urchin embryos than during normal development. In its discussion of cadmium treatment, it suggests autophagy could support cell survival; the Introduction describes autophagy as removing damaged proteins and organelles under stress. The claim’s tentative wording reflects that uncertainty. (LLM, conf=0.90)

  cites [632] source

In the paper

“Some of these, like TLR-mediated RNA recognition, 756 can influence autophagy by either masking any inhibitory effect or compromising autophagy independent of the knockdown probe.”

What the sources say

• Partly supported

  The Introduction and the section ‘ssRNA induces autophagy through TLR7’ show that RNA recognition through TLR7 can induce autophagy; the latter also notes that experimental manipulations sometimes increased background LC3 puncta. The paper does not establish that this response masks a knockdown probe’s inhibitory effect or compromises autophagy independently of the probe. (LLM, conf=0.84)

  cites [756] sourceExperimental — accuracy not yet measured

In the paper

“For example, upon induction by starvation of multicellular development in the protist Dictyostelium, autophagy (or at least Atg1) is required to protect against starvation-induced cell death, allowing vacuolar developmental cell death to take place instead.672,673”

What the sources say

• ?Not checked — no full text of the sourcecites [672] source
• Supported

  The Introduction states that Atg1 protects Dictyostelium cells from starvation-induced cell death and allows subsequent autophagic cell death. The Discussion describes Atg1 as allowing starvation-induced cell death to be replaced by autophagic cell death, whose vacuolization contributes to stalk formation. In the paper’s model, that later death requires a second signal after starvation/cAMP. (LLM, conf=0.93)

  cites [673] source

In the paper

“In this respect it is noteworthy that some cell-types of the immune system such as dendritic cells 260 seem to tolerate loss of autophagy fairly well, whereas others such as T and B cells are compromised in their development and function after autophagy inhibition.757,758”

What the sources say

• ?Not checked — no full text of the sourcecites [260] source
• Partly supported

  The Abstract and Results and Discussion (pages not provided) report reduced peripheral T- and B-cell numbers in Atg5-deficient chimeras and impaired T-cell proliferation, supporting developmental and functional compromise in lymphocytes. The paper does not show impaired B-cell function; it also reports that Atg5-deficient T cells underwent full maturation. The dendritic-cell comparison has a separate citation in the citing sentence and is not attributed to this paper. (LLM, conf=0.86)

  cites [757] sourceExperimental — accuracy not yet measured
• Partly supported

  The Introduction and the section on peripheral B-1 cells report impaired B-cell development and reduced B-1a cell numbers after ATG5 loss. The Introduction also reports a cited finding on T-cell survival and proliferation. This paper does not establish impaired immune-cell function generally or address whether dendritic cells tolerate autophagy loss; the citing sentence assigns those points to other citations. (LLM, conf=0.84)

  cites [758] sourceExperimental — accuracy not yet measured

In the paper

“M2 of influenza virus and HIV Nef block autophagosome degradation.278,760”

What the sources say

• Supported

  The Discussion identifies influenza A virus M2 as the protein that blocks autophagosome degradation. In this sentence with two citations, this source supports the M2 part; the HIV Nef part is not attributed to it. (LLM, conf=0.94)

  cites [278] source
• Supported

  The cited paper supports the HIV Nef part of the sentence. In Results, under “Nef inhibits autophagic maturation,” it concludes that Nef blocks maturation of early autophagic organelles into degradative autolysosomes. The sentence has two citations; this HIV paper is evidence for the Nef claim, not the influenza M2 claim. (LLM, conf=0.92)

  cites [760] source

In the paper

“This results in the induction of BNIP3 and BNIP3L/NIX and”

What the sources say

• Supported

  The Results report autophagy when HIF was stabilized in pVHL-deficient cells, and show that coexpression of BNIP3 and BNIP3L induced autophagic flux in normoxia, mimicking the hypoxic response. The Discussion identifies HIF-dependent coinduction of both proteins as critical to hypoxia-induced autophagy. The paper presents these as related findings rather than directly testing every step of the proposed sequence under normoxia. (LLM, conf=0.87)

  cites [775] source

In the paper

“One example is the chicken lymphoma DT40 cells (see Chicken B-lymphoid DT40 cells below) and their knockout variant for all three inositol 1,4,5-trisphosphate receptor isoforms.241,772,773 In these cells, no differences in basal levels of LC3-II can be observed up to 4 h after addition of fresh medium, but differences can be observed after longer times (Vicencio JM, Szabadkai G, personal comm”

What the sources say

• ?Could not verify

  Split into 2 atoms — 0 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 2 unverifiable. Expand the claim to see per-atom verdicts.

  cites [241] sourceExperimental — accuracy not yet measured
• Supported

  Split into 2 atoms — 2 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [772] source
• ?Not checked — no full text of the sourcecites [773] source
• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites Vicencio JM, Szabadkai G, personal communication

In the paper

“Note that the band running just below LC3-I at approximately 17.5 kDa may be a processing intermediate of LC3-I”

In the paper

“note that an equivalent assay has not been developed for higher eukaryotic cells”

In the paper

“This modified version of the fluorophore has certain properties including a monomeric state, low phototoxic activation and efficient chromophore maturation, which make it suitable for real-time tracking of LC3 and SQSTM1 (Fig. 8; Kaarniranta K, personal communication).”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites Kaarniranta K, personal communication

In the paper

“it is detectable in freshly prepared homogenates, but is less visible after the sample is subjected to a freeze-thaw cycle.”

In the paper

“but we note that it is not always possible to detect endogenous Atg8/LC3.”

In the paper

“for example, high levels of overexpressed GFP-LC3 can result in its nuclear localization, although the protein can still relocate to the cytosol upon starvation”

In the paper

“thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes”

In the paper

“In contrast, during nonselective autophagy, the content of the autophagosome is morphologically identical to the cytoplasm, containing similar densities of ribosomes, and intact sequestered organelles are clearly identifiable.”

In the paper

“In the case of selective autophagy, the cargo should correspond to the specific substrate being targeted for sequestration-bulk cytoplasm is essentially excluded.”

In the paper

“That is, autophagic activity includes not just the increased synthesis or lipidation of Atg8/LC3 (LC3 is a mammalian homolog of yeast Atg8), or an increase in the formation of autophagosomes, but, most importantly, flux through the entire 460”

In the paper

“One critical point is that autophagy is a highly dynamic, multi-step process.”

In the paper

“The focal degradation of cytoplasmic areas sequestered by the phagophore, which matures into the autophagosome, is the morphological hallmark of autophagy.”

In the paper

“Thus, increases in the level of phosphatidylethanolamine (PE)-modified Atg8/LC3 (Atg8-PE/LC3-II), or even the appearance of autophagosomes are not measures of autophagic flux per se, but can reflect the induction of autophagic sequestration and/or inhibition of autophagosome or amphisome clearance.”

In the paper

“For example, inefficient fusion with endosomes and/or lysosomes, respectively, or perturbation of the transport machinery, 18 would inhibit autophagosome maturation to amphisomes or autolysosomes”

What the sources say

• ?Not checked — no full text of the sourcecites [18] source

In the paper

“It should not be assumed that an autophagic system is at steady-state in the strict biochemical meaning of this term, as this implies that the level of autophagosomes does not change with time, and the flux through the system is constant.”

In the paper

“Accordingly, the use of autophagy markers such as LC3-II needs to be complemented by assays to estimate overall autophagic flux, or flow, to permit a correct interpretation of the results.”

In the paper

“we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy.”

In the paper

“An accumulation of autophagosomes [measured by transmission electron microscopy (TEM) image analysis, as fluorescent GFP-MAP1LC3 (GFP-LC3) dots, or as LC3 lipidation on a western blot], could, for example, reflect induction of autophagy, reduction in autophagosome turnover, [14][15][16] or the inability of turnover to keep pace with increased autophagosome formation (Fig. 1).17”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites [14][15][16]
• ?Not checked — no full text of the sourcecites [17] source

In the paper

“Like other cellular pathways, it can be modulated at several steps, both positively and negatively.”

In the paper

“By responding to perturbations in the extracellular environment, cells tune autophagic flux to meet intracellular metabolic demands.”

In the paper

“Therefore, autophagic substrates need to be monitored dynamically over time to verify that they have reached the lysosome/vacuole, and, when appropriate, are degraded.”

In the paper

“this assay is not always performed in a quantitative manner (for example, western blot exposures need to be in the linear range).”

In the paper

“appropriate caution must be used as with any enzyme assay regarding, for example, substrate concentrations and linearity”

In the paper

“all tagged proteins are prone to show enrichment in lysosomes during nonspecific autophagy of the cytoplasm, especially at higher expression levels.”

In the paper

“Alternatively, the lower pH of lysosomes relative to that of the vacuole may contribute to differences in detecting free GFP.”

In the paper

“Also, GFP is relatively resistant to denaturation, and boiling for 5 min may be needed to prevent the folded protein from being trapped in the stacking gel during SDS-PAGE.”

In the paper

“Dendra2 is capable of irreversible photoconversion from a green to a red fluorescent form, but can be used also as a normal GFP or RFP vector.”

In the paper

“The nature of these structures is not fully understood.”

In the paper

“it would be better to use a saturating concentration of lysosomal inhibitors when performing an autophagic flux assay.172”

What the sources say

• ?Not checked — no full text of the sourcecites [172] source

In the paper

“although lysosomotropic compounds should help stabilize GFP by neutralizing the pH in the lysosome”

In the paper

“The cytoplasmic origin of the contents must still be morphologically identifiable, if morphology is the only criterion that is being used for the identification of autolysosomes.”

In the paper

“In quickly frozen and fractured cells the fracture runs preferentially along the hydrophobic plane of the membranes, allowing characterization of the limiting membranes of the different types of autophagic vacuoles and visualization of their limited protein intramembrane particles (IMPs, or integral membrane proteins).”

In the paper

“they do not have an immediate effect on lysosomal hydrolase activity”

In the paper

“Apparently, GFP is more sensitive to mammalian lysosomal hydrolases than to the degradative milieu of the yeast vacuole.”

In the paper

“The GFP-Atg8/LC3 processing assay (the generation of free GFP within the vacuole/lysosome) is a convenient way to monitor autophagy, but it does not work in all cell types, and is not as easy to quantify as enzyme-based assays.”

In the paper

“Accordingly, it is critical that any chimeras be constructed with the fluorescent tag at the N terminus of Atg8/LC3.”

In the paper

“Conclusion: Assays for Atg1/ULK1 can provide detailed insight into the induction of autophagy, but are not a direct measurement of the process.”

In the paper

“Monitoring the endogenous protein, however, obviously depends on the ability to detect it in the system of interest.”

In the paper

“With regard to detection of the WIPI proteins, endogenous WIPI1 puncta cannot be detected in many cell types, 418 and the level of transiently expressed GFP-WIPI1 puncta is cell context-dependent; 418,419”

What the sources say

• ?Not checked — no full text of the sourcecites [418] source
• ?Not checked — no full text of the sourcecites [419] source

In the paper

“Mutants such as atg1D accumulate Atg9 exclusively at the PAS, and this phenotype forms the basis of the transport of Atg9 after knocking out ATG1 (TAKA) assay.63”

What the sources say

• ?Not checked — no full text of the sourcecites [63] source

In the paper

“Nonetheless, it must be kept in mind that monitoring changes in the activity of Atg1/ ULK1 is not a direct assay for autophagy, although such changes may correlate with autophagy activity.”

In the paper

“GFP-WIPI1 and GFP-WIPI2 have been detected on the completed (mature) autophagosome by freeze-fracture analysis, 60 but endogenous WIPI2 has not been detected on mRFP-LC3-or LAMP2-positive autophagosomes or autolysosomes using immunolabeling.418”

What the sources say

• ?Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  cites [60]
• ?Not checked — no full text of the sourcecites [418] source