Neuronal Apolipoprotein E4 Expression Results in Proteome-Wide Alterations and Compromises Bioenergetic Capacity by Disrupting Mitochondrial Function

In neurons, apoE4 is associated with decreased neurite outgrowth.

“decreased neurite outgrowth”

• cites [10] Mahley, 2006, Apolipoprotein E4: A causative factor and therapeutic target in neuropathology, including Alzheimer’s disease, Proc Natl Acad Sci U S A sourceSupported
• cites [17] Chang, 2005, Lipid- and receptor-binding regions of apolipoprotein E4 fragments act in concert to cause mitochondrial dysfunction and neurotoxicity, Proc Natl Acad Sci U S A sourceCould not verify

  The Results report greater cell death from an apoE4 fragment in Neuro-2a cells, but do not report decreased neurite outgrowth. Searches for “neurite outgrowth” and “neurite extension” found no passage in our copy addressing that outcome. (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourceCould not verify

  The section on mitochondrial respiratory complexes in neuronal cells reports lower levels of those complexes in apoE4-expressing neurons, but does not assess neurite outgrowth. Searches for “neurite outgrowth” and “neurite extension and neuronal process growth” found no passage addressing the claimed outcome. Page numbers were not available in our copy. (LLM, conf=0.73)

  Experimental — accuracy not yet measured
• cites [19] Chen, 2012, Small molecule structure correctors abolish detrimental effects of apolipoprotein E4 in cultured neurons, J Biol Chem sourceSupported
• cites [34] Brodbeck, 2008, Rosiglitazone increases dendritic spine density and rescues spine loss caused by apolipoprotein E4 in primary cortical neurons, Proc Natl Acad Sci U S A sourcePartly supported

  The Results report that apoE4 reduced dendritic spine density in primary cortical neurons compared with apoE3. The introduction mentions neurite outgrowth, but our copy does not clearly state that apoE4 decreases it; spine density and neurite outgrowth are distinct measures. No page number is available in the indexed passages. (LLM, conf=0.83)

  Experimental — accuracy not yet measured
• cites [35] Brodbeck, 2011, Structure-dependent impairment of intracellular apolipoprotein E4 trafficking and its detrimental effects are rescued by small-molecule structure correctors, J Biol Chem sourceSupported

ApoE fragments escape the secretory pathway into the cytosol.

“The resulting apoE fragments escape the secretory pathway into the cytosol”

• cites [17] Chang, 2005, Lipid- and receptor-binding regions of apolipoprotein E4 fragments act in concert to cause mitochondrial dysfunction and neurotoxicity, Proc Natl Acad Sci U S A sourceSupported
• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourcePartly supported

  The Discussion (page not provided) says neuronal apoE can be cleaved into fragments and that an apoE4 fragment mislocalized to mitochondria when expressed in N2A cells. It does not say the fragments escape the secretory pathway into the cytosol; it describes mitochondrial targeting of proteolytic fragments in this study as a possibility, not an established route. (LLM, conf=0.81)

  Experimental — accuracy not yet measured
• cites [19] Chen, 2012, Small molecule structure correctors abolish detrimental effects of apolipoprotein E4 in cultured neurons, J Biol Chem sourceCould not verify

  The Discussion (page not available) mentions neuron-specific proteolysis and neurotoxic apoE4 fragment formation. Neither that passage nor passages retrieved for “cytosol” and “secretory pathway” say that apoE fragments leave the secretory pathway or enter the cytosol. (LLM, conf=0.74)

  Experimental — accuracy not yet measured

In neurons, apoE4 is associated with impaired mitochondrial respiration.

“In neurons, apoE4 is associated with impaired mitochondrial respiration”

• cites [10] Mahley, 2006, Apolipoprotein E4: A causative factor and therapeutic target in neuropathology, including Alzheimer’s disease, Proc Natl Acad Sci U S A sourcePartly supported

  The section “ApoE4 Neuropathology Independent of Aβ” (page unavailable) associates apoE4 with reduced neuronal glucose utilization and describes mitochondrial dysfunction in cultured neurons. That supports impaired mitochondrial function, but the paper does not specifically establish impaired mitochondrial respiration. (LLM, conf=0.82)

  Experimental — accuracy not yet measured
• cites [17] Chang, 2005, Lipid- and receptor-binding regions of apolipoprotein E4 fragments act in concert to cause mitochondrial dysfunction and neurotoxicity, Proc Natl Acad Sci U S A sourcePartly supported

  The Results report impaired mitochondrial function in Neuro-2a cells expressing truncated apoE4 fragments, measured by MitoTracker fluorescence. They do not report a direct measure of mitochondrial respiration, and the effect was observed with fragments rather than full-length apoE4. (LLM, conf=0.87)

  Experimental — accuracy not yet measured
• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourceSupported
• cites [19] Chen, 2012, Small molecule structure correctors abolish detrimental effects of apolipoprotein E4 in cultured neurons, J Biol Chem sourceSupported
• cites [34] Brodbeck, 2008, Rosiglitazone increases dendritic spine density and rescues spine loss caused by apolipoprotein E4 in primary cortical neurons, Proc Natl Acad Sci U S A sourcePartly supported

  The introduction (page unavailable) says that apoE4 fragments in neurons impair mitochondrial membrane potential and function, and the Results likewise describe impaired mitochondrial function. The copy does not specifically identify impaired mitochondrial respiration, so it supports the broader association but not that particular measure. (LLM, conf=0.85)

  Experimental — accuracy not yet measured
• cites [35] Brodbeck, 2011, Structure-dependent impairment of intracellular apolipoprotein E4 trafficking and its detrimental effects are rescued by small-molecule structure correctors, J Biol Chem sourceCould not verify

  The Abstract and the section on apoE4 trafficking in mouse primary hippocampal neurons report impaired intracellular trafficking, not mitochondrial respiration. Searches for mitochondrial respiration and oxygen consumption found no passage in our copy addressing the claimed respiratory impairment. (LLM, conf=0.75)

  Experimental — accuracy not yet measured

In neurons, apoE4 is associated with neurotoxicity.

“and neurotoxicity [10, 17–19, 34, 35]”

• cites [10] Mahley, 2006, Apolipoprotein E4: A causative factor and therapeutic target in neuropathology, including Alzheimer’s disease, Proc Natl Acad Sci U S A sourceSupported
• cites [17] Chang, 2005, Lipid- and receptor-binding regions of apolipoprotein E4 fragments act in concert to cause mitochondrial dysfunction and neurotoxicity, Proc Natl Acad Sci U S A sourceSupported
• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourceSupported
• cites [19] Chen, 2012, Small molecule structure correctors abolish detrimental effects of apolipoprotein E4 in cultured neurons, J Biol Chem sourceSupported
• cites [34] Brodbeck, 2008, Rosiglitazone increases dendritic spine density and rescues spine loss caused by apolipoprotein E4 in primary cortical neurons, Proc Natl Acad Sci U S A sourceSupported
• cites [35] Brodbeck, 2011, Structure-dependent impairment of intracellular apolipoprotein E4 trafficking and its detrimental effects are rescued by small-molecule structure correctors, J Biol Chem sourceSupported

The largest apoE4 fragment, apoE4(1-272), causes mitochondrial fragmentation.

“and cause mitochondrial fragmentation [10, 17, 20]”

• cites [10] Mahley, 2006, Apolipoprotein E4: A causative factor and therapeutic target in neuropathology, including Alzheimer’s disease, Proc Natl Acad Sci U S A sourcePartly supported

  In “ApoE4 Neuropathology Independent of Aβ” (page unavailable), the paper says apoE4(1-272) accumulates in mitochondria and describes mitochondrial dysfunction associated with apoE fragments. It does not say that apoE4(1-272) causes mitochondrial fragmentation, a distinct change in mitochondrial structure. (LLM, conf=0.84)

  Experimental — accuracy not yet measured
• cites [17] Chang, 2005, Lipid- and receptor-binding regions of apolipoprotein E4 fragments act in concert to cause mitochondrial dysfunction and neurotoxicity, Proc Natl Acad Sci U S A sourcePartly supported

  The Results report that apoE4(1-272) interacted with mitochondria in Neuro-2a cells and reduced MitoTracker fluorescence, which the authors interpret as impaired mitochondrial function and integrity. They do not report mitochondrial fragmentation; the filamentous inclusions described in the Results concern the apoE4 fragment, not mitochondrial shape. (LLM, conf=0.85)

  Experimental — accuracy not yet measured
• cites [20] Nakamura, 2009, Apolipoprotein E4 (1–272) fragment is associated with mitochondrial proteins and affects mitochondrial function in neuronal cells, Mol Neurodegener sourcePartly supported

  The Conclusion (page unavailable) reports that apoE4(1–272) expression in Neuro2a cells reduced the activities of mitochondrial respiratory complexes III and IV. This supports an effect on mitochondrial function, but the copy does not report mitochondrial fragmentation; searches for fragmentation and mitochondrial morphology found no such result. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

Young, cognitively normal apoE4 carriers have glucose hypometabolism in brain regions affected by Alzheimer's disease.

“Young, cognitively normal apoE4 carriers have glucose hypometabolism in brain regions affected by AD [29–31]”

• cites [29] Reiman, 1996, Preclinical evidence of Alzheimer’s disease in persons homozygous for the ɛ4 allele for apolipoprotein E, N Engl J Med sourcePartly supported

  The Abstract reports reduced glucose metabolism in Alzheimer’s-affected brain regions among cognitively normal ε4 homozygotes. But its Methods specify volunteers aged 50–65, and Figure 2 gives a mean age of 55; this does not establish the claim for young carriers or for carriers generally. Page numbers are not supplied in our copy. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [30] Small, 2000, Cerebral metabolic and cognitive decline in persons at genetic risk for Alzheimer’s disease, Proc Natl Acad Sci U S A sourcePartly supported

  The Results report lower glucose metabolism in the inferior parietal, lateral temporal, and posterior cingulate regions of nondemented APOE-4 carriers. The Methods give an age range of 50–84 years, and the Results give carriers a mean age of 65.9 years; this does not support describing the studied group as young. Their memory scores were within age norms, although many had age-associated memory impairment or mild cognitive impairment. Page numbers are unavailable in our copy. (LLM, conf=0.90)

  Experimental — accuracy not yet measured
• cites [31] Reiman, 2004, Functional brain abnormalities in young adults at genetic risk for late-onset Alzheimer’s dementia, Proc Natl Acad Sci U S A sourceNot checked — no full text of the source

ApoE4 impairs respiratory function in Neuro-2a cells.

“and impairs respiratory function [18]”

• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourceSupported

In nondemented apoE4 carriers, mitochondrial complex IV activity is decreased in cortical regions susceptible to Alzheimer's disease.

“in nondemented apoE4 carriers, the activity of mitochondrial complex IV is decreased in cortical regions susceptible to AD [32]”

• cites [32] Valla, 2010, Reduced posterior cingulate mitochondrial activity in expired young adult carriers of the APOE ɛ4 allele, the major late-onset Alzheimer’s susceptibility gene, J Alzheimers Dis sourcePartly supported

  The Results report significantly lower cytochrome oxidase (mitochondrial complex IV) activity in young-adult APOE ε4 carriers than in noncarriers, but the measurements were from the posterior cingulate cortex. The Discussion says further studies are needed to determine whether the reduction extends beyond that region, so the claim's plural “cortical regions” is broader than this study establishes. Page numbers are unavailable in our copy. (LLM, conf=0.89)

  Experimental — accuracy not yet measured

The single amino acid difference increases apoE4 cleavage by a stress-induced neuron-specific protease.

“increases its cleavage by a stress-induced neuron-specific protease [7, 12, 16]”

• cites [7] Huang, 2012, Alzheimer mechanisms and therapeutic strategies, Cell sourcePartly supported

  In the ApoE4 section (page unavailable), the paper says apoE isoforms differ by single amino acid substitutions and neuronal apoE4 is more susceptible to cleavage than apoE3. It says stress or injury induces neuronal apoE expression, not that the protease is stress-induced; it reports cleavage in neurons but does not establish that the protease itself is neuron-specific. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [12] Mahley, 2012, Apolipoprotein E sets the stage: Response to injury triggers neuropathology, Neuron sourcePartly supported

  The ‘ApoE Hypothesis’ section says structural differences make apoE4 more susceptible to neuronal proteolytic cleavage than apoE3 or apoE2. The ‘Neuron-Specific Proteolysis’ section describes the responsible protease as unidentified. The paper says injury or stress induces neuronal apoE synthesis, not that it induces the protease. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [16] Huang, 2001, Apolipoprotein E fragments present in Alzheimer’s disease brains induce neurofibrillary tangle-like intracellular inclusions in neurons, Proc Natl Acad Sci U S A sourceNot checked — no full text of the source

ApoE4 increases risk for Alzheimer's disease.

“ApoE4 increases risk for AD”

• cites [4] Biundo, 2015, Interaction of apoE3 and apoE4 isoforms with an ITM2b/BRI2 mutation linked to the Alzheimer disease-like Danish dementia: Effects on learning and memory, Neurobiol Learn Mem sourceSupported
• cites [5] Bu, 2009, Apolipoprotein E and its receptors in Alzheimer’s disease: Pathways, pathogenesis and therapy, Nat Rev Neurosci sourceSupported
• cites [6] Holtzman, 2012, Apolipoprotein E and apolipoprotein E receptors: Normal biology and roles in Alzheimer disease, Cold Spring Harb Perspect Med sourceNot checked — no full text of the source
• cites [7] Huang, 2012, Alzheimer mechanisms and therapeutic strategies, Cell sourceSupported
• cites [8] Kim, 2009, The role of apolipoprotein E in Alzheimer’s disease, Neuron sourceSupported
• cites [9] Liu, 2013, Apolipoprotein E and Alzheimer disease: Risk, mechanisms and therapy, Nat Rev Neurol sourceSupported
• cites [10] Mahley, 2006, Apolipoprotein E4: A causative factor and therapeutic target in neuropathology, including Alzheimer’s disease, Proc Natl Acad Sci U S A sourceSupported
• cites [11] Mahley, 2009, Apolipoprotein E: Structure determines function, from atherosclerosis to Alzheimer’s disease to AIDS, J Lipid Res sourceSupported
• cites [12] Mahley, 2012, Apolipoprotein E sets the stage: Response to injury triggers neuropathology, Neuron sourceSupported
• cites [13] Mahley, 2012, Small-molecule structure correctors target abnormal protein structure and function: The structure corrector rescue of apolipoprotein E–associated neuropathology, J Med Chem sourceSupported

Mice expressing human apoE4 show deficient uptake and metabolism of glucose in the brain.

“Mice expressing human apoE4 show deficient uptake and metabolism of glucose in the brain [33].”

• cites [33] Wu, 2018, Human apoE isoforms differentially modulate brain glucose and ketone body metabolism: Implications for Alzheimer’s disease risk reduction and early intervention, J Neurosci sourceSupported

Expression of apoE4 fragments containing the lipid-binding domain in N2a cells results in neurotoxicity.

“expression of fragments containing this domain in N2a cells results in neurotoxicity [17]”

• cites [17] Chang, 2005, Lipid- and receptor-binding regions of apolipoprotein E4 fragments act in concert to cause mitochondrial dysfunction and neurotoxicity, Proc Natl Acad Sci U S A sourcePartly supported

  The Results report that expressing apoE4(1–272), which contains the lipid-binding region, caused greater cell death in Neuro-2a cells. But the Results also state that the lipid-binding region alone caused no neurotoxicity: the receptor-binding region was required too. The claim omits that qualification. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

The interaction of apoE4 fragments with mitochondria is dependent on the apoE lipid-binding domain.

“The interaction of apoE4 fragments with mitochondria is dependent on the apoE lipid-binding domain”

• cites [17] Chang, 2005, Lipid- and receptor-binding regions of apolipoprotein E4 fragments act in concert to cause mitochondrial dysfunction and neurotoxicity, Proc Natl Acad Sci U S A sourceSupported

ApoE4 reduces the level of respiratory chain subunits in Neuro-2a cells.

“ApoE4 also reduces the level of respiratory chain subunits in Neuro-2a (N2a) cells”

• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourceSupported

Expression of apoE4 in mouse neurons, but not astrocytes, selectively decreases the expression of mitochondrial respiratory subunits.

“Expression of apoE4 in mouse neurons, but not astrocytes, selectively decreases the expression of mitochondrial respiratory subunits [18].”

• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourceSupported

Impaired PDH activity might be partly responsible for the respiratory deficits in apoE4 cells.

“Since PDH deficiency limits respiratory capacity and causes lactic acidosis in humans [92], impaired PDH activity might be partly responsible for the respiratory deficits in apoE4 cells.”

• cites [92] Fluge, 2016, Metabolic profiling indicates impaired pyruvate dehydrogenase function in myalgic encephalopathy/chronic fatigue syndrome, JCI Insight sourceSupported

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

Using MitoMiner 4.0 and the Gene Ontology "Mitochondrion" category, 953 proteins associated with mitochondria were identified.

“Using the MitoMiner 4.0 database [73] and the Gene Ontology “Mitochondrion” category [74, 75], we identified 953 proteins associated with mitochondria; the levels of 42% of those proteins were significantly changed by apoE4 expression, again with equal numbers increased and decreased (Fig. 5A).”

• cites [73] Smith, 2016, MitoMiner v3.1, an update on the mitochondrial proteomics database, Nucleic Acids Res sourcePartly supported

  The Introduction identifies MitoMiner as a database for investigating mitochondrial proteins and says it incorporates Gene Ontology annotations. But this paper describes MitoMiner v3.1, not the cited version 4.0. The 953-protein count is presented as the citing authors’ own result, not a finding attributed to this source. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [74] Ashburner, 2000, Gene Ontology: Tool for the unification of biology, Nat Genet sourceSupported
• cites [75] The Gene Ontology Consortium, 2017, Expansion of the Gene Ontology knowledgebase and resources, Nucleic Acids Res sourceSupported

• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Pscan and PASTAA identified transcription factors that may coordinate the changes in protein levels in apoE4-expressing cells.

“To determine which regulatory pathways help compensate for apoE4-induced mitochondrial dysfunction, we used Pscan [64] and PASTAA [63] to identify transcription factors that may coordinate the changes in protein levels in apoE4-expressing cells (Table 1).”

• cites [64] Zambelli, 2009, Pscan: Finding over-represented transcription factor binding site motifs in sequences from co-regulated or co-expressed genes, Nucleic Acids Res sourceSupported
• cites [63] Roider, 2009, PASTAA: Identifying transcription factors associated with sets of co-regulated genes, Bioinformatics sourceSupported

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Previously, the authors showed that apoE4 decreases levels of cytochrome c oxidase 1 (COX1).

“Previously we showed that apoE4 decreases levels of cytochrome c oxidase 1 (COX1) [18], a mitochondrially encoded subunit that initiates complex IV assembly [82].”

• cites [18] Chen, 2011, Apolipoprotein E4 domain interaction mediates detrimental effects on mitochondria and is a potential therapeutic target for Alzheimer disease, J Biol Chem sourceSupported

Several of the transcription factors identified are implicated in neurodegeneration, including nuclear respiratory factor 1, nuclear factor erythroid-2-like 2, ETS domain-containing protein Elk-1, Kruppel-like factor 14, and Sp4.

“Additionally, several of the transcription factors identified are implicated in neurodegeneration, including nuclear respiratory factor 1 [117, 118], nuclear factor erythroid-2-like 2 [110, 111], ETS domain-containing protein Elk-1 [119], Kruppel-like factor 14 [120], and Sp4 [121].”

• cites [117] Taherzadeh-Fard, 2011, PGC-1alpha downstream transcription factors NRF-1 and TFAM are genetic modifiers of Huntington disease, Mol Neurodegener sourceSupported
• cites [118] Urfer-Buchwalder, 2017, Identification of a nuclear respiratory factor 1 recognition motif in the apolipoprotein E variant APOE4 linked to Alzheimer’s disease, Sci Rep sourceSupported
• cites [110] Scarpulla, 2005, Nuclear control of respiratory gene expression in mammalian cells, J Cell Biochem sourceNot checked — no full text of the source
• cites [111] Ohtsuji, 2008, Nrf1 and Nrf2 play distinct roles in activation of antioxidant response element-dependent genes, J Biol Chem sourceCould not verify

  The Abstract and Discussion describe Nrf1 and Nrf2 in antioxidant-gene regulation and mouse liver pathology, but the passages do not link either factor to neurodegeneration. A search for neurodegenerative disease and neuronal terms found no passage establishing that link. The paper’s CNC-family Nrf1 should also not be assumed to mean nuclear respiratory factor 1. (LLM, conf=0.75)

  Experimental — accuracy not yet measured
• cites [119] Besnard, 2011, Elk-1 a transcription factor with multiple facets in the brain, Front Neurosci sourceSupported

ApoE4 also affects respiratory complex formation.

“In concert with mitochondrial proteases, which respond to altered mitochondrial protein stoichiometry [107], apoE4 also affects respiratory complex formation [108].”

• cites [108] Friedman, 2015, MICOS coordinates with respiratory complexes and lipids to establish mitochondrial inner membrane architecture, eLife sourceCould not verify

  The Results and Discussion passages address MICOS, respiratory complexes, and mitochondrial architecture, but do not address apoE4. Searches for “apoE4,” “apolipoprotein E4,” and its effect on respiratory complex formation found no supporting passage in our copy. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Increased ER-mitochondrial interactions increase MAM activity in neurons treated with conditioned medium from apoE4- but not apoE3-expressing astrocytes.

“Increased ER-mitochondrial interactions increase MAM activity in neurons treated with conditioned medium from apoE4- but not apoE3-expressing astrocytes [104].”

• cites [104] Tambini, 2016, ApoE4 upregulates the activity of mitochondria-associated ER membranes, EMBO Rep sourcePartly supported

  The Results and Discussion report increased phosphatidylethanolamine synthesis in mouse hippocampal neurons treated with ApoE4 versus ApoE3 astrocyte-conditioned medium, supporting increased MAM-related activity. They do not show that increased physical ER–mitochondrial interactions caused that neuronal effect; the paper says the mechanism is unclear, and its apposition measurement showed only a trend in ApoE4-treated cells. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

Global proteomic profiling revealed widespread changes in mitochondrial processes in apoE4 cells.

“Global proteomic profiling revealed widespread changes in mitochondrial processes in apoE4 cells”

Glycolysis was enhanced in apoE4 cells under basal conditions.

“Glycolysis was enhanced in apoE4 cells under basal conditions; however, the glycolytic rate and ATP production were insufficient to maintain total cellular energy under conditions of high metabolic demand.”

Under maximal stress stimulation, apoE4-expressing neurons lacked spare capacity to generate significant ATP levels by OXPHOS and glycolysis.

“apoE4-expressing neurons, unlike those expressing apoE3, lacked spare capacity to generate significant ATP levels by OXPHOS and glycolysis (Fig. 1D)”

ApoE4 cells had altered levels of proteins related to proteases.

“proteases”

The rate of Ca2+ influx into mitochondria in response to extracellular Ca2+ was much greater in apoE4-expressing cells than in apoE3-expressing cells.

“In contrast, in response to extracellular Ca2+, both the basal mitochondrial Ca2+ level (Fig. 4E) and the rate of Ca2+ influx into mitochondria in response to extracellular Ca2+ were much greater in cells expressing apoE4 (Fig. 4F, H), culminating in twofold higher levels of mitochondrial Ca2+ (Fig. 4G).”

Maximal glycolysis rate was similar in apoE3- and apoE4-expressing cells when glycolysis was stimulated by monensin and respiration was inhibited by antimycin A.

“the maximal glycolysis rate was similar in apoE3- and apoE4-expressing cells when glycolysis was stimulated by monensin and respiration was inhibited by antimycin A”

ApoE4 expression increased mitochondrial calcium levels.

“and mitochondrial calcium”

The total NAD+ + NADH pool was smaller in apoE4-expressing cells than in apoE3-expressing cells.

“Interestingly, the total NAD+ + NADH pool was smaller in the apoE4 cells, suggesting that apoE4 impairs NAD+ synthetic capacity [68].”

ApoE4 altered a wide range of proteins involved in OXPHOS, redox homeostasis, and glucose and amino acid metabolism.

“ApoE4 altered a wide range of proteins involved in OXPHOS, redox homeostasis, and glucose and amino acid metabolism (Fig. 6).”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

ApoE4 cells had altered levels of proteins related to mitochondrial ribosomal proteins.

“and mitochondrial ribosomal proteins”

ApoE4 cells had altered levels of proteins related to mitochondrial protein translocation.

“mitochondrial protein translocation”

Combined total ATP production rates under basal conditions were similar in apoE3 and apoE4 cells.

“The combined total ATP production rates under basal conditions were similar.”

Volcano plots confirmed that there were both significantly higher and significantly lower levels of subsets of mitochondrial proteins in apoE4-expressing cells.

“Volcano plots of –log10 (adjusted p-value) significance levels versus fold change in protein levels confirmed that there were both significantly higher and significantly lower levels of subsets of mitochondrial proteins in the apoE4-expressing cells (Fig. 5B).”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

ApoE4 decreased the levels of 33 of 64 detected subunits.

“Our proteomic analysis showed that apoE4 decreased the levels of 33 of 64 detected subunits, undoubtedly altering complex assembly.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Cytosolic Ca2+ influx after thapsigargin treatment was not significantly lower in apoE4 than in apoE3 cells.

“Cytosolic Ca2+ influx after treatment with thapsigargin was not significantly lower in apoE4 than apoE3 cells (Fig. 4I, J), but mitochondrial Ca2+ influx was significantly greater in apoE4 cells (Fig. 4K), resulting in nearly eightfold higher levels (Fig. 4L).”

Under basal conditions, apoE4 increased glycolysis in N2a cells.

“Under basal conditions, apoE4 impaired respiration and increased glycolysis”

• Also stated in: § ApoE4 expression alters the bioenergetics profile in neuronal cells · plain

  “apoE4-expressing cells shift to glycolysis to meet energy demands under basal conditions (Fig. 1B)”

ApoE4 expression increased the levels of reactive oxygen species.

“increased the levels of reactive oxygen species”

MITRAC12, a central component of the MITRAC assembly complex, was reduced by 47% in apoE4-expressing cells (p < 1×10-9).

“For example, complex IV assembly requires the MITRAC assembly complex, a central component of which, MITRAC12, was reduced by 47% in apoE4-expressing cells (p < 1×10–9).”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.
• AssumptionRests on an assumption: Complex IV assembly requires the MITRAC assembly complex.

TIGAR was also decreased significantly in apoE4-expressing cells.

“Interestingly, TIGAR, an enzyme that negatively regulates glycolytic activity [88, 89], was also decreased significantly, perhaps to enhance glycolytic activity in the context of mitochondrial dysfunction and reduced levels of glycolytic enzymes.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

ATP production rates by OXPHOS in apoE4 cells were 33% lower than in apoE3 cells.

“ATP production rates by OXPHOS in apoE4 cells were 33% lower”

NAD+ was significantly lower in apoE4-expressing cells than in apoE3-expressing cells.

“We found that NADH was significantly higher, and NAD+ was significantly lower, in apoE4-expressing cells, resulting in a lower NAD+/NADH ratio (Fig. 3A).”

ApoE4 cells had major disruptions to all detected subunits in complex V (ATP synthase).

“and major disruptions to all detected subunits in complex V (ATP synthase)”

In response to extracellular Ca2+, the basal mitochondrial Ca2+ level was much greater in apoE4-expressing cells than in apoE3-expressing cells.

“In contrast, in response to extracellular Ca2+, both the basal mitochondrial Ca2+ level (Fig. 4E) and the rate of Ca2+ influx into mitochondria in response to extracellular Ca2+ were much greater in cells expressing apoE4 (Fig. 4F, H), culminating in twofold higher levels of mitochondrial Ca2+ (Fig. 4G).”

ATP production rates by glycolysis in apoE4 cells were increased to a greater extent than in apoE3 cells.

“were increased to a greater extent by glycolysis than in apoE3 cells (Fig. 1C)”

ApoE4-expressing neurons display a coordinated decrease in respiratory chain subunits.

“and a coordinated decrease in respiratory chain subunits”

ApoE4-expressing cells had significantly lower rates of basal and FCCP-stimulated respiration than apoE3-expressing cells.

“apoE4-expressing neuronal cells had significantly lower rates of basal and FCCP-stimulated respiration than apoE3-expressing cells (Fig. 1A)”

Mitochondrial Ca2+ levels 6 min after thapsigargin treatment were nearly eightfold higher in apoE4 cells than in apoE3 cells.

“Cytosolic Ca2+ influx after treatment with thapsigargin was not significantly lower in apoE4 than apoE3 cells (Fig. 4I, J), but mitochondrial Ca2+ influx was significantly greater in apoE4 cells (Fig. 4K), resulting in nearly eightfold higher levels (Fig. 4L).”

ApoE4 cells had altered levels of proteins related to mitochondrial fusion/fission.

“mitochondrial fusion/fission”

ApoE4-expressing neurons display impaired mitochondrial energy production.

“ApoE4-expressing neurons display impaired mitochondrial energy production”

There were alterations in a wide range of proteins involved in glycolysis, redox regulation, intermediary metabolism, mitochondrial morphology, and the import, synthesis, and degradation of mitochondrial proteins.

“There were also alterations in a wide range of proteins involved in glycolysis, redox regulation, and intermediary metabolism; in mitochondrial morphology; and in the import, synthesis, and degradation of mitochondrial proteins.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Basal cytosolic Ca2+ levels did not differ between apoE3- and apoE4-expressing cells.

“The two cell lines did not differ in basal cytosolic Ca2+ levels (Fig. 4A) or in overall Ca2+ influx induced by extracellular Ca2+ (Fig. 4B).”

Label-free quantitative proteomic profiling of whole-cell lysates from N2a-apoE3 and N2a-apoE4 cells by LC-MS/MS identified approximately 4500 distinct proteins.

“Label-free, quantitative proteomic profiling of whole-cell lysates from N2a-apoE3 and N2a-apoE4 cells by LC-MS/MS identified approximately 4500 distinct proteins (Fig. 5A).”

• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

COX7a is significantly decreased in apoE4-expressing neurons.

“For example, COX7a has a critical role in stabilizing complexes III and IV [39] and is significantly decreased in apoE4-expressing neurons.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

ApoE4-expressing cells shift to glycolysis to meet energy demands under basal conditions.

“apoE4-expressing cells shift to glycolysis to meet energy demands under basal conditions (Fig. 1B)”

Under conditions that prevent net ATP production by glycolysis (basal respiration only), apoE4-expressing cells maintained total ATP levels during the 1-h challenge.

“Surprisingly, under conditions that prevent net ATP production by glycolysis (basal respiration only) [65], apoE4-expressing cells maintained total ATP levels during the 1-h challenge, possibly by decreasing energy utilization or increasing reliance on glycogen stores.”

The NAD+/NADH ratio was lower in apoE4-expressing cells than in apoE3-expressing cells.

“We found that NADH was significantly higher, and NAD+ was significantly lower, in apoE4-expressing cells, resulting in a lower NAD+/NADH ratio (Fig. 3A).”

Many other mitochondrial matrix enzymes, including those involved in the tricarboxylic acid cycle and in amino acid and ketone body metabolism, were significantly increased in apoE4-expressing cells.

“In contrast, many other mitochondrial matrix enzymes, including those involved in the tricarboxylic acid cycle and in amino acid and ketone body metabolism, were significantly increased in apoE4-expressing cells (Fig. 6).”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

ApoE4-expressing cells were more reliant on glycolysis under basal conditions than apoE3 cells.

“apoE4-expressing cells were more reliant on glycolysis under basal conditions than apoE3 cells”

Both analyses highlighted transcription factors that regulate the expression of mitochondrial genes and genes related to metabolism and redox homeostasis.

“Both analyses highlighted transcription factors that regulate the expression of mitochondrial genes and genes related to metabolism and redox homeostasis.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Glycolytic ATP production by apoE3- and apoE4-expressing cells increased to comparable levels when mitochondrial ATP synthesis was blocked with oligomycin or when respiration was inhibited with antimycin A and monensin.

“glycolytic ATP production by apoE3- and apoE4-expressing cells increased to comparable levels when mitochondrial ATP synthesis was blocked with oligomycin or when respiration was inhibited with antimycin A and monensin”

Overall Ca2+ influx induced by extracellular Ca2+ did not differ between apoE3- and apoE4-expressing cells.

“The two cell lines did not differ in basal cytosolic Ca2+ levels (Fig. 4A) or in overall Ca2+ influx induced by extracellular Ca2+ (Fig. 4B).”

Gene Ontology and KEGG pathway-enrichment analyses revealed a diverse array of effects of apoE4 on cellular processes.

“These analyses revealed a diverse array of effects of apoE4 on cellular processes.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

When stressed with FCCP, ATP production by OXPHOS in apoE4-expressing cells was markedly lower than in apoE3-expressing cells.

“but ATP production by OXPHOS was markedly lower”

The changes in apoE4-expressing neurons are associated with altered calcium handling (increased mitochondrial calcium).

“altered calcium handling (increased mitochondrial calcium)”

ApoE4 cells showed increases in many other mitochondrial components, including proteins involved in mitochondrial morphology, protein import, protein maturation and degradation, and mitochondrial protein synthesis.

“We also detected increases in many other mitochondrial components in apoE4 cells, including proteins involved in mitochondrial morphology (fusion/fission; cristae formation), protein import (TOM/TIM complexes and assembly), protein maturation and degradation (proteases), and mitochondrial protein synthesis (ribosomal subunits) (Fig. 9).”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Mitochondrial processes and pathways were identified from the sets of proteins increased or decreased by apoE4.

“Interestingly, mitochondrial processes and pathways were identified from the sets of proteins increased or decreased by apoE4.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

ApoE4 cells had altered levels of proteins related to mitochondrial endoplasmic reticulum-associated membranes.

“Also altered in apoE4 cells were levels of proteins related to mitochondrial endoplasmic reticulum–associated membranes”

Total reserve capacity was about 50% lower in apoE4-expressing cells.

“the total reserve capacity was about 50% lower in apoE4-expressing cells”

Mitochondrial Ca2+ influx after thapsigargin treatment was significantly greater in apoE4 cells than in apoE3 cells.

“Cytosolic Ca2+ influx after treatment with thapsigargin was not significantly lower in apoE4 than apoE3 cells (Fig. 4I, J), but mitochondrial Ca2+ influx was significantly greater in apoE4 cells (Fig. 4K), resulting in nearly eightfold higher levels (Fig. 4L).”

Under basal conditions, apoE4 impaired respiration in N2a cells.

“Under basal conditions, apoE4 impaired respiration and increased glycolysis”

• Also stated in: § ApoE4 expression alters the bioenergetics profile in neuronal cells · plain

  “apoE4-expressing neuronal cells had significantly lower rates of basal and FCCP-stimulated respiration than apoE3-expressing cells (Fig. 1A)”

ApoE4 expression decreased the NAD+/NADH ratio.

“ApoE4 expression also decreased the NAD+/NADH ratio”

Under conditions that increase ATP demand by glycolysis and impair mitochondrial ATP production (stimulated glycolysis only), apoE3- and apoE4-expressing cells showed a similar partial loss of cellular ATP production.

“Under conditions that increase ATP demand by glycolysis and impair mitochondrial ATP production (stimulated glycolysis only), apoE3- and apoE4-expressing cells showed a similar partial loss of cellular ATP production.”

ApoE4 cells had reduced levels of numerous respiratory complex subunits.

“including reduced levels of numerous respiratory complex subunits”

NADH was significantly higher in apoE4-expressing cells than in apoE3-expressing cells.

“We found that NADH was significantly higher, and NAD+ was significantly lower, in apoE4-expressing cells, resulting in a lower NAD+/NADH ratio (Fig. 3A).”

The changes in apoE4-expressing neurons are associated with increased ROS production.

“increased ROS production”

The changes in apoE4-expressing neurons are countered by increased levels of subsets of other mitochondrial proteins involved in mitochondrial morphology, protein transport, and protein degradation.

“are countered by increased levels of subsets of other mitochondrial proteins involved in mitochondrial morphology, protein transport, and protein degradation”

The levels of all glycolytic enzymes in apoE4-expressing cells that differed from the levels in apoE3-expressing cells were lower in the apoE4 cells.

“For example, the levels of all glycolytic enzymes in apoE4-expressing cells that differed from the levels in apoE3-expressing cells were lower in the apoE4 cells.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Intracellular ROS levels determined by CellROX assay were approximately twofold higher in apoE4-expressing cells than in apoE3-expressing cells.

“Intracellular ROS levels determined by CellROX assay were approximately twofold higher in apoE4-expressing cells (Fig. 3B).”

When stressed with FCCP, ATP production by glycolysis in apoE4-expressing cells was similar to that in apoE3-expressing cells.

“when cells were stressed with FCCP, ATP production by glycolysis in apoE4-expressing cells was similar to that in apoE3-expressing cells”

The levels of both E1 subunits of PDH in apoE4 cells were reduced by 20%.

“Indeed, levels of both E1 subunits of PDH in apoE4 cells were reduced by 20%.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

When challenged or stressed, apoE4-expressing neurons had 50% less reserve capacity to generate ATP to meet energy requirements than apoE3-expressing neurons.

“when challenged or stressed, apoE4-expressing neurons had 50% less reserve capacity to generate ATP to meet energy requirements than apoE3-expressing neurons”

Mitochondrial respiration and OXPHOS-dependent ATP production are impaired in apoE4-expressing cells, being 40-50% lower than in apoE3-expressing cells.

“We found that mitochondrial respiration and OXPHOS-dependent ATP production are impaired in apoE4-expressing cells, being 40–50% lower than in apoE3-expressing cells.”

The changes in apoE4-expressing neurons are associated with a lower NAD+/NADH ratio.

“These changes are associated with a lower NAD+/NADH ratio”

ApoE4 expression altered the mitochondrial cristae-organizing system.

“ApoE4 expression also altered the mitochondrial cristae–organizing system, which stabilizes the cristae junction between the inner and outer membranes.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

The levels of four major proteins directly involved in mitochondrial dynamics were significantly increased by apoE4 expression.

“Levels of four major proteins directly involved in mitochondrial dynamics were significantly increased by apoE4 expression, as shown by proteomics analyses.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

Unbiased analyses to predict which transcription factors might control the proteomic changes in apoE4 cells identified a surprising number of factors known to influence mitochondrial biogenesis and regulate the expression of additional metabolism- and redox-related genes.

“Our unbiased analyses to predict which transcription factors might control the proteomic changes in apoE4 cells identified a surprising number of factors known to influence mitochondrial biogenesis and regulate the expression of additional metabolism- and redox-related genes.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

The levels of all of these MAM proteins except GRP75 were significantly increased in apoE4 cells.

“Remarkably, levels of all of these MAM proteins except GRP75 were significantly increased in the apoE4 cells (Fig. 8).”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

The levels of all mitochondrial ribosomal proteins detected by proteomics were increased in apoE4 cells.

“Interestingly, the levels of all mitochondrial ribosomal proteins [109] detected by proteomics were increased.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: The N2a neuronal cell model recapitulates the cell-autonomous effects of apoE4 in human neurons relevant to neurodegeneration.

The single amino acid difference alters the intramolecular interactions in apoE4.

“which alters the intramolecular interactions in apoE4”

• cites [15] Mahley, 1988, Apolipoprotein E: Cholesterol transport protein with expanding role in cell biology, Science sourceNot checked — no full text of the source

ApoE4 impairs NAD+ synthetic capacity.

“Interestingly, the total NAD+ + NADH pool was smaller in the apoE4 cells, suggesting that apoE4 impairs NAD+ synthetic capacity [68].”

• cites [68] Verdin, 2015, NAD+ in aging, metabolism, and neurodegeneration, Science sourceNot checked — no full text of the source

The decrease in 33 of 64 detected subunits alters complex assembly.

“Our proteomic analysis showed that apoE4 decreased the levels of 33 of 64 detected subunits, undoubtedly altering complex assembly.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

Despite numerous cellular adaptations, apoE4-expressing neurons remained vulnerable to metabolic stress.

“despite numerous cellular adaptations, apoE4-expressing neurons remained vulnerable to metabolic stress”

The altered stoichiometry of respiratory complex subunits could adversely affect various aspects of mitochondrial activity.

“Thus, the altered stoichiometry of respiratory complex subunits could adversely affect various aspects of mitochondrial activity.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

As a result, apoE4-expressing neuronal cells were more severely affected by stress.

“As a result, apoE4-expressing neuronal cells were more severely affected by stress.”

ApoE4 disrupts the metabolism of glycolysis-derived pyruvate by pyruvate dehydrogenase (PDH).

“Our results suggest that apoE4 disrupts the metabolism of glycolysis-derived pyruvate by pyruvate dehydrogenase (PDH).”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

COX7a has a critical role in stabilizing complexes III and IV.

“For example, COX7a has a critical role in stabilizing complexes III and IV [39] and is significantly decreased in apoE4-expressing neurons.”

• cites [39] Letts, 2016, The architecture of respiratory supercomplexes, Nature sourceNot checked — no full text of the source

ApoE4 expression is associated with metabolic inflexibility under conditions of high-energy demand.

“Evidently, apoE4 expression is associated with metabolic inflexibility under conditions of high-energy demand.”

Total ATP content under the standard condition (Basal Glycolysis + Respiration) was the same in both cell lines.

“Total ATP content under the standard condition (“Basal Glycolysis + Respiration”) was the same in both cell lines”

The induction of specific enzymes likely accommodates altered metabolite fluxes in response to slowed respiration and NADH build-up.

“This induction of specific enzymes likely accommodates altered metabolite fluxes in response to slowed respiration and NADH build-up.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

The glycolytic rate and ATP production were insufficient to maintain total cellular energy under conditions of high metabolic demand in apoE4 cells.

“Glycolysis was enhanced in apoE4 cells under basal conditions; however, the glycolytic rate and ATP production were insufficient to maintain total cellular energy under conditions of high metabolic demand.”

ApoE4-induced bioenergetic deficits led to extensive metabolic rewiring.

“ApoE4-induced bioenergetic deficits led to extensive metabolic rewiring”

Even if the interaction of apoE4 with mitochondria disrupts just a few critical assembly factors or subunits, it might cause the widespread depletion of functional complexes, impaired respiration, and the reduced ATP levels observed in the cells.

“Therefore, even if the interaction of apoE4 with mitochondria disrupts just a few critical assembly factors or subunits, it might cause the widespread depletion of functional complexes, impaired respiration, and the reduced ATP levels we observed in our cells.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.
• AssumptionRests on an assumption: Complex IV assembly requires the MITRAC assembly complex.

Despite enhanced glycolysis under basal conditions, apoE4-expressing cells cannot call upon greater glycolytic reserves during periods of high energy demand to compensate for their lower respiration rate.

“despite enhanced glycolysis under basal conditions, apoE4-expressing cells cannot call upon greater glycolytic reserves during periods of high energy demand to compensate for their lower respiration rate”

The profound effect on all detected subunit levels in complex V (ATP synthase) would disrupt mitochondrial ATP production.

“Likewise, the profound effect on all detected subunit levels in complex V (ATP synthase) would disrupt mitochondrial ATP production.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

ApoE4 cells regulate mitochondrial Ca2+ differently from apoE3 cells.

“Thus, apoE4 cells regulate mitochondrial Ca2+ differently from apoE3 cells.”

Glycolysis in apoE4 cells was insufficient to compensate for the impaired mitochondrial ATP production during metabolic stress.

“Regardless, glycolysis in apoE4 cells was insufficient to compensate for the impaired mitochondrial ATP production during metabolic stress.”

Ca2+ released from the ER of apoE4 cells flowed more quickly into mitochondria, bypassing the cytoplasm.

“The initial lower cytosolic Ca2+ influx coincided with the initial higher mitochondrial Ca2+ influx in apoE4 than in apoE3 cells, suggesting that Ca2+ released from the ER of apoE4 cells flowed more quickly into mitochondria, bypassing the cytoplasm.”

Despite various adaptations, apoE4-expressing neuronal cells remained vulnerable to adverse metabolic events.

“Despite various adaptations, apoE4-expressing neuronal cells remained vulnerable to adverse metabolic events.”

These changes in factors involved in mitochondrial morphology, biogenesis, and quality control may compensate for the large-scale apoE4-induced decreases in OXPHOS complexes.

“These changes in factors involved in mitochondrial morphology, biogenesis, and quality control may compensate for the large-scale apoE4-induced decreases in OXPHOS complexes.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

Mitochondrial-associated apoE4 disrupts the balance of core subunits of the respiratory complexes to prevent complete maturation of assembled complexes in the inner mitochondrial membrane.

“Judging from the broad effects of apoE4 on the respiratory complexes, we propose that mitochondrial-associated apoE4 disrupts the balance of core subunits of the respiratory complexes to prevent complete maturation of assembled complexes in the inner mitochondrial membrane.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.

The increased levels of MAM proteins in apoE4-expressing cells might reflect increased tethering that is associated with the increased Ca2+ flux and increased ROS production observed.

“Thus, the increased levels of MAM proteins in apoE4-expressing cells might reflect increased tethering that is associated with the increased Ca2+ flux and increased ROS production we observed.”

• AssumptionRests on an assumption: Changes in protein levels detected by proteomics produce corresponding functional changes in the affected pathways.