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Cited by 35Cholesterol and Lipid MetabolismLipoproteins and Cardiovascular HealthLipid metabolism and disorders

The DOI matches its Crossref record; 48 of the 51 references it cites were found in a registry or an index; 16 of the 65 sources its claims cite were checked for support.

Asialoglycoprotein receptor 1 is a novel PCSK9-independent ligand of liver LDLR cleaved by furin

Open access
Susan-Resiga, Delia, Girard, Emmanuelle, Essalmani, Rachid, Roubtsova, Anna, Marcinkiewicz, Jadwiga, Derbali, Rabeb M., Evagelidis, Alexandra, Byun, Jae H., Lebeau, Paul F., Austin, Richard C., Seidah, Nabil G.·2021·Journal of Biological Chemistry
DOI 10.1016/j.jbc.2021.101177PMC PMC8479480

Published version of a preprint

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Abstract

ABSTRACT The hepatic carbohydrate-recognizing asialoglycoprotein receptor (ASGR1) mediates the endocytosis/lysosomal degradation of desialylated glycoproteins following binding to terminal galactose/N-acetylgalactosamine. Human heterozygote-carriers of ASGR1- deletions exhibited ∼34% lower risk of coronary artery disease and ∼10-14% non-HDL-cholesterol reduction. Since PCSK9 is a major degrader of LDLR, the regulation of LDLR and/or PCSK9 by ASGR1 was studied. Thus, we investigated the role of endogenous/overexpressed ASGR1 on LDLR degradation and functionality by Western-blot and immunofluorescence in HepG2 naïve and HepG2-PCSK9-knockout cells. ASGR1, like PCSK9, targets LDLR and both interact with/enhance the degradation of the receptor independently. Such lack of cooperativity between PCSK9 and ASGR1 on LDLR expression was confirmed in livers of wild-type (WT) versus Pcsk9 -/- mice. ASGR1-knockdown in HepG2 naïve cells significantly increased total (∼1.2-fold) and cell-surface (∼4-fold) LDLR protein. In HepG2-PCSK9-knockout cells ASGR1-silencing led to ∼2-fold higher levels of LDLR protein and DiI-LDL uptake, associated with ∼9-fold increased cell-surface LDLR. Overexpression of WT-ASGR1/2 reduced primarily the immature non-O-glycosylated LDLR (∼110 kDa), whereas the triple Gln 240 /Trp 244 /Glu 253 Ala-mutant (loss of carbohydrate-binding) reduced the mature form of the LDLR (∼150 kDa), suggesting that ASGR1 binds the LDLR in sugar-dependent and -independent fashion. Furin sheds ASGR1 at R KM K 103 ↓ into a secreted form, likely resulting in a loss-of-function on LDLR. LDLR is the first example of a liver-receptor ligand of ASGR1. Additionally, we demonstrate that lack of ASGR1 enhances LDLR levels and DiI-LDL incorporation, independently of PCSK9. Overall, silencing of ASGR1 and PCSK9 may lead to higher LDL-uptake by hepatocytes, thereby providing a novel approach to further reduce LDL-cholesterol.

Overview

AI snapshot

Written from the paper’s full text.

TL;DR

ASGR1 is a novel PCSK9-independent ligand of liver LDLR that promotes LDLR degradation and reduces LDL uptake; furin cleaves ASGR1 at RKMK103↓SL into a secreted ~28-kDa form. Silencing ASGR1 increases total and cell-surface LDLR and DiI-LDL uptake independently of PCSK9, suggesting combined ASGR1/PCSK9 inhibition may further lower LDL cholesterol.

Key findings

  • PCSK9 and ASGR1 independently reduce LDLR: PCSK9 lowered cell-surface LDLR by ~90% (IF) and total LDLR by ~40% (WB) but did not alter ASGR1; ASGR1 knockdown increased total LDLR ~1.2-fold and cell-surface LDLR ~4-fold in HepG2-naïve cells, and ~2-fold total and ~9-fold cell-surface LDLR in HepG2-PCSK9-KO cells, with ~2-fold higher DiI-LDL uptake.
  • ASGR1 binds LDLR: endogenous ASGR1 co-immunoprecipitated with LDLR in HepG2-PCSK9-KO cells and mouse liver; WT ASGR1 bound both immature ~110-kDa and mature ~150-kDa LDLR, whereas the Q240A/W244A/E253A CRD mutant primarily bound immature LDLR and did not efficiently reduce cell-surface LDLR or DiI-LDL uptake.
  • Furin cleaves ASGR1 at RKMK103↓SL into a secreted ~28-kDa form; mutation of this site abrogated cleavage, while an RRRR103↓EL substitution caused complete cleavage of mature ~48-kDa ASGR1.
  • ASGR1 specificity: ASGR1 knockdown increased LDLR ~2-fold but did not affect CD36 or insulin receptor β levels in HepG2-PCSK9-KO cells.

Methods

  • Cell models: HepG2-naïve, HepG2-PCSK9-KO, IHH, and HEK293 cells; ASGR1/PCSK9 siRNA knockdown, ASGR1/ASGR2 and LDLR overexpression, purified PCSK9 treatment, DiI-LDL uptake, immunofluorescence, Western blotting, qRT-PCR, co-immunoprecipitation, and cell-surface biotinylation.
  • Mouse models: WT, Pcsk9−/−, and Ldlr−/− mice; liver immunohistochemistry and Western blotting for LDLR/ASGR1, plus in situ hybridization for ASGR1/ASGR2 mRNA during development and adulthood.
  • Statistics: averages ± SD, Student’s t test, p < 0.05 considered significant.

Limitations

  • Stated in the paper: the repertoire of liver glycoproteins targeted by ASGR1 is unknown; other ASGR1 functions related to inflammation, atherosclerosis, and cardiovascular health are yet to be identified; possible extrahepatic roles of shed ASGR1 are speculative.
  • Evident from the text: mechanistic conclusions rely heavily on immortalized cell lines and overexpression systems, with in vivo work limited to mouse liver protein levels and co-IP rather than ASGR1 loss-of-function lipid or cardiovascular endpoints.
  • Evident from the text: ASGR1/ASGR2 homo- and hetero-oligomer context and overexpression may not reflect endogenous stoichiometry, and human evidence is cited from prior ASGR1 variant studies rather than directly tested here.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1016/j.jbc.2021.101177

Cites

48 of 48 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Verification

Claims the paper makes

16 of 65 cited sources checked · 8 supported · 1 flagged · accuracy not yet measured

  • Premisecitation

    The reduction in cardiovascular disease risk observed in ASGR1 del12 carriers (approximately 34%) is approximately 2-fold greater than would have been predicted (approximately 18%) from the associated modest reduction of LDL cholesterol.

    “Comparison of plasma LDLc decrease in LOF PCSK9 R46L (−17 mg/dl; ∼30% reduction in CVD risk) with that in LOF ASGR1 del12 mutation (−11 mg/dl) illustrates that reduction in CVD risk observed in this ASGR1 mutant (∼34%) was ∼2-fold greater than would have been predicted (i.e., ∼18%) from the associated modest reduction of LDLc (12).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [12] Nioi, 2016, Variant ASGR1 associated with a reduced risk of coronary artery disease, N. Engl. J. Med. source

      Figure 2 reports a combined odds ratio of 0.66 for coronary artery disease in ASGR1 del12 carriers, corresponding to the cited 34% reduction. The Coronary Risk and del12 section says the reduction is greater than predicted by the variant’s effect on non-HDL cholesterol. Our copy does not give the claimed 18% prediction or twofold comparison; its lipid table reports a combined LDL-cholesterol decrease of 12.5 mg/dl, rather than 11 mg/dl. (LLM, conf=0.87)

      Experimental — accuracy not yet measured
  • Findingcitation

    ASGR1 transcripts are exclusively expressed in liver, specifically in hepatocytes.

    “A Tabula Muris compendium of single-cell RNA-Seq transcriptome data from 20 mouse organs and tissues (czbiohub.org) (14) revealed that ASGR1 transcripts are exclusively expressed in liver, specifically in hepatocytes (Fig. 1C).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [14] Tabula Muris Consortium, 2018, Single-cell transcriptomics of 20 mouse organs creates a Tabula Muris, Nature source
  • Premisecitation

    Two human ASGR1-deletion variants found in Iceland, resulting in truncated proteins likely inactive, are associated with a modest ∼10% to 14% reduction in LDLc.

    “two human ASGR1-deletion variants found in Iceland, resulting in truncated proteins, likely inactive, are associated with a modest ∼10% to 14% reduction in LDLc”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [12] Nioi, 2016, Variant ASGR1 associated with a reduced risk of coronary artery disease, N. Engl. J. Med. source

      The Abstract and Second Loss-of-Function Variant section identify two Icelandic ASGR1 loss-of-function variants, but only del12 is a deletion; p.W158X is a 4-bp insertion. The lipid-results table (p. 2134) reports lower LDL cholesterol for del12, while the p.W158X results report lower non-HDL cholesterol, not an LDL reduction of approximately 10% to 14% for both variants. (LLM, conf=0.90)

      Experimental — accuracy not yet measured
  • Premisecitation

    The ASGR1 del12 mutation is associated with a significant 34% lower risk of cardiovascular disease.

    “In addition, the del12 mutation was associated with a significant 34% lower risk of CVD.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [12] Nioi, 2016, Variant ASGR1 associated with a reduced risk of coronary artery disease, N. Engl. J. Med. source

      The Coronary Risk and del12 section reports a combined odds ratio of 0.66 (P = 4.0×10⁻⁶), corresponding to 34% lower odds among del12 carriers. That result is specifically for coronary artery disease, not cardiovascular disease broadly. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
  • Findingown data

    Insertion of an optimal Furin-like site RRRR103 down arrow EL resulted in complete cleavage of mature approximately 48-kDa ASGR1 by endogenous Furin into a soluble secreted approximately 28-kDa sASGR1.

    “Indeed, insertion of an optimal Furin-like site RRRR103↓EL (17, 29) resulted in complete cleavage of mature ∼48-kDa ASGR1 (upper band, likely endoH resistant (17)) by endogenous Furin into a soluble secreted ∼28-kDa sASGR1 (Fig. 11D).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [17] Essalmani, 2021, Implications of spike-glycoprotein processing at S1/S2 by furin, at S2’ by furin and/or TMPRSS2 and shedding of ACE2: Cell-to-cell fusion, cell entry and infectivity of SARS-CoV-2, bioRxiv

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

    • cites [29] Benjannet, 2006, The proprotein convertase PCSK9 is inactivated by furin and/or PC5/6A: Functional consequences of natural mutations and post-translational modifications, J. Biol. Chem. source
  • Probably not a substantial claim. 33
  • It may state two separate claims as one. 40

  • Probably not a substantial claim.

    Premisecitation

    Shedding of the LDLR by metalloproteases is thought to mostly target iLDLR.

    “Finally, shedding of the LDLR by metalloproteases is thought to mostly target iLDLR (23).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [23] Nassoury, 2007, The cellular trafficking of the secretory proprotein convertase PCSK9 and its dependence on the LDLR, Traffic source

      In “Interaction of PCSK9 and the LDLR” (page unavailable), the paper distinguishes immature LDLR from shed LDLR and reports that an ER-retained LDLR mutant, which does not mature, is not shed into the medium. That runs contrary to the claim that shedding mostly targets immature LDLR; the paper does not quantify which form metalloproteases target most often. (LLM, conf=0.78)

      Checked by a reasoning model — may be wrong.

      Experimental — accuracy not yet measured

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    Loss-of-function PCSK9 variants enhance LDLR levels and promote LDL cholesterol clearance.

    “Unlike gain-of-function PCSK9 variants (7), loss-of-function (LOF) variants enhance LDLR levels and promote LDLc clearance (4, 8).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [4] Cohen, 2005, Low LDL cholesterol in individuals of African descent resulting from frequent nonsense mutations in PCSK9, Nat. Genet. source
    • cites [8] Cohen, 2006, Sequence variations in PCSK9, low LDL, and protection against coronary heart disease, N. Engl. J. Med. source

      The Conclusions report that mice lacking Pcsk9 have more hepatic LDL receptors and clear LDL from plasma faster. But the same section says the molecular mechanisms by which the human PCSK9 sequence variants lower LDL cholesterol are not known, so it does not establish that those variants act through increased LDLR levels and clearance. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • Rests on an assumption: Removal of plasma LDL cholesterol is primarily mediated by the LDLR located on the surface of hepatocytes.

  • It may state two separate claims as one.

    Premisecitation

    Human heterozygous carriers of ASGR1 deletions exhibit approximately 34% lower risk of coronary artery disease and approximately 10% to 14% reduction of non-HDL cholesterol.

    “Human heterozygote carriers of ASGR1 deletions exhibit ∼34% lower risk of coronary artery disease and ∼10% to 14% reduction of non-HDL cholesterol.”
    • Also stated in: § Abstract · plain
      “Recent studies showed that heterozygous carriers of the early termination LOF del12 mutation in the ASGR1 gene (1 in 120 persons in the Icelandic study) and another early termination heterozygote LOF ASGR1 mutant (W158X) had lower plasma levels of non-HDL cholesterol than noncarriers (12).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [12] Nioi, 2016, Variant ASGR1 associated with a reduced risk of coronary artery disease, N. Engl. J. Med. source

  • Probably not a substantial claim. It may state two separate claims as one.

    Premisecitation

    PCSK9 is highly expressed in the liver and, upon secretion into plasma, binds to and enhances the degradation of LDLR in a nonenzymatic fashion, thereby increasing circulating LDL cholesterol.

    “The proprotein convertase subtilisin/kexin type 9 (PCSK9) is highly expressed in the liver (2) and, upon its secretion into plasma, binds to and enhances the degradation of the LDLR in a nonenzymatic fashion (3, 4, 5, 6), thereby increasing circulating LDLc.”
    Quote in the paperSubstantial claim1 of 5 sources checked
    • cites [2] Seidah, 2003, The secretory proprotein convertase neural apoptosis-regulated convertase 1 (NARC-1): Liver regeneration and neuronal differentiation, Proc. Natl. Acad. Sci. U. S. A. source
    • cites [3] Maxwell, 2004, Adenoviral-mediated expression of Pcsk9 in mice results in a low-density lipoprotein receptor knockout phenotype, Proc. Natl. Acad. Sci. U. S. A. source
    • cites [4] Cohen, 2005, Low LDL cholesterol in individuals of African descent resulting from frequent nonsense mutations in PCSK9, Nat. Genet. source
    • cites [5] McNutt, 2007, Catalytic activity is not required for secreted PCSK9 to reduce low density lipoprotein receptors in HepG2 cells, J. Biol. Chem. source

      Checked before — its earlier verdict is held back until a new check runs.

    • cites [6] Seidah, 2017, The proprotein convertases in hypercholesterolemia and cardiovascular diseases: Emphasis on proprotein convertase subtilisin/kexin 9, Pharmacol. Rev. source
    • Rests on an assumption: Removal of plasma LDL cholesterol is primarily mediated by the LDLR located on the surface of hepatocytes.

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    Furin can cleave and inactivate PCSK9 in liver hepatocytes.

    “It can also cleave and inactivate PCSK9 in liver hepatocytes (29, 30).”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [29] Benjannet, 2006, The proprotein convertase PCSK9 is inactivated by furin and/or PC5/6A: Functional consequences of natural mutations and post-translational modifications, J. Biol. Chem. source

      The paper states that furin cleaves and inactivates PCSK9 and reports evidence from hepatic HepG2 cells (passage [1], labeled Abstract in our copy). It does not directly demonstrate that cleavage and inactivation occur in liver hepatocytes; its discussion says liver-furin knockout evidence was not available. (LLM, conf=0.84)

      Experimental — accuracy not yet measured
    • cites [30] Essalmani, 2011, In vivo evidence that furin from hepatocytes inactivates PCSK9, J. Biol. Chem. source

  • It may state two separate claims as one.

    Findingcitation

    In Pcsk9-knockout mice, LDLR protein levels at the cell surface of hepatocytes and in whole tissue are increased, while ASGR1 levels are not affected.

    “The data showed that, although LDLR protein levels at the cell surface of hepatocytes (IHC) and in whole tissue (WB) were increased in Pcsk9-KO livers, in accordance with (21), those of ASGR1 were not affected.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [21] Roubtsova, 2015, PCSK9 deficiency unmasks a sex/tissue-specific subcellular distribution of the LDL and VLDL receptors in mice, J. Lipid Res. source

      In “LDLR subcellular distribution is affected by sex in PCSK9 KO livers” (page unavailable), the paper reports increased LDLR in liver extracts from both sexes, but increased hepatocyte-surface LDLR only in knockout males, not females. The citation is attached to the LDLR comparison; the sentence presents the ASGR1 result as the citing authors’ own data, so ASGR1 is not assessed as a claim attributed to this source. (LLM, conf=0.90)

      Experimental — accuracy not yet measured

  • It may state two separate claims as one.

    Findingown data

    Lack of PCSK9 does not affect the mRNA levels of LDLR, ASGR1, ASGR2, or SREBP2 in HepG2 cells.

    “As expected (18, 19), lack of PCSK9 did not affect the mRNA levels of LDLR. In addition, no effect was observed on the mRNA levels of ASGR1, ASGR2, and the transcription factor SREBP2 implicated in PCSK9 mRNA regulation (20) (Fig. 2C).”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [18] Zaid, 2008, Proprotein convertase subtilisin/kexin type 9 (PCSK9): Hepatocyte-specific low-density lipoprotein receptor degradation and critical role in mouse liver regeneration, Hepatology source

      The citation is attached to the LDLR mRNA statement. In “Analysis of Liver LDLR and Plasma PCSK9” (page unavailable), the source reports higher LDLR protein levels in PCSK9-knockout mouse livers, but does not report whether LDLR mRNA changed. Searches for LDLR mRNA and transcripts also found no HepG2 result; the ASGR1, ASGR2, and SREBP2 findings are presented with the citing paper’s Fig. 2C, not attributed to this source. (LLM, conf=0.75)

      Experimental — accuracy not yet measured
    • cites [19] Gupta, 2010, A locked nucleic acid antisense oligonucleotide (LNA) silences PCSK9 and enhances LDLR expression in vitro and in vivo, PLoS One source
    • cites [20] Dubuc, 2004, Statins upregulate PCSK9, the gene encoding the proprotein convertase neural apoptosis-regulated convertase-1 implicated in familial hypercholesterolemia, Arterioscler. Thromb. Vasc. Biol. source

  • It may state two separate claims as one.

    Premisecitation

    O-glycosylated chains of mLDLR are predominantly present close to the C-terminal transmembrane domain but are also found in linker regions separating the N-terminal repeat domains of the LDLR.

    “The latter are predominantly present close to the C-terminal transmembrane domain but are also found in the linker regions separating the N-terminal repeat domains of the LDLR (22).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [22] Pedersen, 2014, Low density lipoprotein receptor class A repeats are O-glycosylated in linker regions, J. Biol. Chem. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Findingown data

    In most cases, mRNA levels of ASGR1 were higher than those of ASGR2, especially in mouse liver.

    “In most cases the mRNA levels of ASGR1 were higher than those of ASGR2, especially in mouse liver.”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    ASGR1 knockdown in HepG2-naive cells significantly increases total LDLR protein by approximately 1.2- to 1.3-fold and cell-surface LDLR by approximately 4-fold.

    “ASGR1 knockdown in HepG2-naïve cells significantly increased total (∼1.2-fold) and cell-surface (∼4-fold) LDLR protein.”
    • Also stated in: § ASGR1 regulates LDLR levels and functionality independently of PCSK9 · hedged
      “Specifically, we estimated a total LDLR increase of ∼1.3-fold (Fig. 5A) and ∼2-fold (Fig. 5B) in each cell line, respectively. The corresponding cell-surface increase in LDLR was at least ∼4-fold in naïve HepG2 cells (Fig. 5, C and D) and ∼9-fold in HepG2-PCSK9-KO cells (Fig. 5E).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Endogenous ASGR1 is present at the plasma membrane and colocalizes with LDLR in HepG2-PCSK9-KO cells.

    “Immunofluorescence (IF) staining of endogenous ASGR1 in HepG2-PCSK9-KO cells confirmed its presence at the plasma membrane and revealed its colocalization with the LDLR (−PCSK9; Fig. 2A).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Cell-surface levels of the triple-Ala mutant of ASGR1 are approximately 60% higher than wildtype, mirroring its approximately 70% increased total cellular levels.

    “Furthermore, we noticed that the cell-surface levels of the triple-Ala mutant of ASGR1 are ∼60% (∼1.6 fold) higher than the WT (Fig. 9B; top left panel), mirroring its ∼70% (∼1.7 fold) increased total cellular levels (Fig. 9B; right panel) and suggesting that even at higher levels, this mutant that binds iLDLR (Fig. 8C; left panel) does not efficiently reduce LDLR levels (Fig. 9, A and B).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Wildtype ASGR1 reduces levels of iLDLR, which is not seen with CRD mutants that primarily reduce mLDLR.

    “As observed at the plasma membrane (Fig. 9B; left panel), in total cell lysate (intracellular and plasma membrane) WT ASGR1 also reduced the levels of iLDLR, which is not seen with the CRD mutants that primarily reduce the levels of mLDLR (Fig. 9C; lower left panel).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    PCSK9 knockdown results in approximately 1.5-fold increase in LDLR protein with no change in ASGR1 protein in HepG2 cells.

    “At the protein level, PCSK9 KD resulted in ∼1.5-fold increase in LDLR, with no change in ASGR1 (Fig. 2D).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    In mouse, ASGR1 and ASGR2 mRNA are mainly expressed in liver starting at embryonic days 17 and 15, respectively, and their expression increases until adulthood.

    “In mouse, whole-body in situ hybridization histochemistry of ASGR1 and ASGR2 mRNA expression during development and in the adult revealed that both transcripts are mainly expressed in liver starting at embryonic days 17 and 15, respectively, and showed that their expression increases until adulthood (Fig. 1B).”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    Both cell-surface LDLR and DiI-LDL uptake are significantly reduced in the presence of wildtype ASGR1 but not its QA/WA/EA triple Ala-mutant in HEK293 cells.

    “In nonpermeabilized cells, IF showed that both cell-surface LDLR and DiI-LDL uptake are significantly reduced in the presence of WT ASGR1 but not of its QA/WA/EA triple Ala-mutant (Fig. 9A).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Loss of endogenous ASGR1 in hepatic cell lines enhances LDLR levels and the ability to uptake DiI-LDL.

    “demonstrated that loss of endogenous ASGR1 in hepatic cells lines enhanced the levels of the LDLR and its ability to uptake DiI-LDL (Fig. 5)”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Overexpression of ASGR1 decreases LDLR levels and the ability to uptake DiI-LDL.

    “whereas the reverse is true upon overexpression of ASGR1 (Fig. 6)”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    The ratio of cell-surface iLDLR to total LDLR is significantly reduced by approximately 40% by wildtype ASGR1 but not by the triple mutant.

    “The data showed that the ratio of cell-surface [iLDLR]/[total LDLR] is significantly reduced (∼40%) by WT ASGR1 but not by the triple mutant (Fig. 9B; lower left panel).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Lack of endogenous ASGR1 in HepG2 cells does not affect PCSK9 mRNA or its protein levels in cells and media.

    “Finally, lack of endogenous ASGR1 in HepG2 cells does not affect PCSK9 mRNA or its protein levels in cells and media (Fig. 4).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Coexpression of ASGR1 and ASGR2 at a 1:1 ratio significantly decreases total levels of endogenous LDLR by approximately 30% and overexpressed LDLR by approximately 55% in HepG2-PCSK9-KO cells.

    “Coexpression of ASGR1 and ASGR2 at a 1:1 ratio resulted in a significant decrease in the total levels of both endogenous (−30%) and overexpressed (−55%) LDLR (WB; Fig. 6A).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    ASGR1 knockdown increases LDLR by approximately 2-fold, while CD36 and IR-beta levels are not affected in HepG2-PCSK9-KO cells.

    “The data show that, although KD of ASGR1 increased the levels of LDLR by ∼2-fold, those of CD36 and IR-β were not affected (Fig. 10).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    In Ldlr-knockout mice, ASGR1 cell-surface immunoreactivity and total ASGR1 protein are significantly increased by approximately 1.6-fold and approximately 1.3-fold, respectively.

    “To analyze the impact of the lack of LDLR on ASGR1 protein, we now show that, in the liver of Ldlr-KO mice, both ASGR1 cell-surface immunoreactivity (Fig. 3A) and total ASGR1 protein by WB (Fig. 3B) were significantly increased by ∼1.6- and ∼1.3-fold, respectively.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    In HepG2-PCSK9-knockout cells, ASGR1 silencing leads to approximately 2-fold higher total LDLR protein and approximately 9-fold increased cell-surface LDLR.

    “In HepG2-PCSK9-knockout cells, ASGR1 silencing led to ∼2-fold higher levels of LDLR protein and DiI (1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate)-LDL uptake associated with ∼9-fold increased cell-surface LDLR.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Exogenous PCSK9 reduces cell-surface LDLR by approximately 90% by immunofluorescence and total LDLR by approximately 40% by Western blot in HepG2 cells.

    “As expected, PCSK9 reduced the endogenous levels of cell-surface LDLR (−90% by IF) and total LDLR (−40% by WB).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Silencing ASGR1 in HepG2-PCSK9-KO cells significantly increases total cellular LDLR levels, associated with approximately 3-fold enhanced shedding of LDLR (approximately 10% of total LDLR), especially iLDLR.

    “Indeed, silencing ASGR1 in HepG2-PCSK9-KO cells (that still endogenously express ASGR2, Fig. 1D) significantly increased total cellular LDLR levels, associated with an ∼3-fold enhanced shedding of the LDLR (∼10% of total LDLR), especially iLDLR (Fig. 9D).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Only Furin can cleave ASGR1 at RKMK103↓SL and shed it into the media.

    “Our data showed that only Furin can cleave ASGR1 at RKMK103↓SL and shed it into the media (Fig. 11, A, C and E)”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingreasoning

    Overexpression of WT ASGR1 decreases LDLR shedding, likely due to degradation of mainly iLDLR.

    “Overexpression of WT ASGR1 decreased LDLR shedding, likely due to degradation of mainly iLDLR (Fig. 9C)”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    Inhibitory human monoclonal antibodies against PCSK9 reduce the risk of cardiovascular events in clinical populations.

    “The development of inhibitory human monoclonal antibodies against PCSK9 represents a powerful treatment strategy for the management of CVD, which has been shown to reduce the risk of cardiovascular events in clinical populations (9).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [9] Sabatine, 2017, Cardiovascular safety and efficacy of the PCSK9 inhibitor evolocumab in patients with and without diabetes and the effect of evolocumab on glycaemia and risk of new-onset diabetes: A prespecified analysis of the FOURIER randomised controlled trial, Lancet Diabetes Endocrinol. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    ASGRs are highly expressed in hepatocytes.

    “ASGRs are highly expressed in hepatocytes (10).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [10] Ashwell, 1982, Carbohydrate-specific receptors of the liver, Annu. Rev. Biochem. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    Hepatic ASGR is pivotal for the efficient clearance of desialylated glycoproteins from circulation by receptor-mediated endocytosis and lysosomal degradation, whereupon ASGR is recycled back to the cell surface.

    “The hepatic ASGR is pivotal for the efficient clearance of desialylated glycoproteins from circulation by receptor-mediated endocytosis and lysosomal degradation, whereupon ASGR is recycled back to the cell surface (13, 31).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [13] Bischoff, 1987, Two asialoglycoprotein receptor polypeptides in human hepatoma cells, J. Biol. Chem. source
    • cites [31] Weigel, 2002, Glycans as endocytosis signals: The cases of the asialoglycoprotein and hyaluronan/chondroitin sulfate receptors, Biochim. Biophys. Acta source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    Furin, the third member of the proprotein convertase family, enhances the shedding of various cell-surface proteins at the general PC-like motif (R/K)Xn(R/K) down arrow.

    “Furin, the third member of the proprotein convertase (PC) family (26), has been shown to enhance the shedding of various cell-surface proteins at the general PC-like motif (R/K)Xn(R/K)↓, where Xn represents 0, 2, 4, or 6 spacer aa (27).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [26] Seidah, 2012, The biology and therapeutic targeting of the proprotein convertases, Nat. Rev. Drug Discov. source
    • cites [27] Lichtenthaler, 2018, Proteolytic ectodomain shedding of membrane proteins in mammals-hardware, concepts, and recent developments, EMBO J. source

      Checked before — its earlier verdict is held back until a new check runs.

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    Both ASGR subunits are type II, single-pass transmembrane proteins.

    “Both subunits are type II, single-pass transmembrane proteins.”
    • Also stated in: § Discussion · plain
      “Both subunits are type II transmembrane proteins comprising a cytosolic, transmembrane domain; a luminal stalk region; and a calcium-dependent CRD (Fig. 1A).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [13] Bischoff, 1987, Two asialoglycoprotein receptor polypeptides in human hepatoma cells, J. Biol. Chem. source
    • cites [31] Weigel, 2002, Glycans as endocytosis signals: The cases of the asialoglycoprotein and hyaluronan/chondroitin sulfate receptors, Biochim. Biophys. Acta source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    In ASGR1, residues Gln240, Trp244, and Glu253 in the carbohydrate recognition domain are critical for binding exposed terminal galactose or N-acetylgalactosamine residues.

    “In ASGR1, residues Gln240, Trp244, and Glu253 in the carbohydrate recognition domain (CRD) are critical for binding exposed terminal galactose or N-acetylgalactosamine residues (https://www.uniprot.org/uniprot/P07306) (Fig. 1A).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites https://www.uniprot.org/uniprot/P07306

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

  • It may state two separate claims as one.

    Interpretationown data

    Endogenous ASGR1 and LDLR interact specifically and form a complex independent of PCSK9.

    “This suggests that endogenous ASGR1 and LDLR interact in a specific manner and form a complex independent of PCSK9.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    Strategies aimed at inhibiting ASGR1 expression or activity are being explored as another approach to lowering plasma LDL cholesterol and cardiovascular disease risk.

    “Strategies aimed at inhibiting ASGR1 expression/activity are being explored as another approach to lowering plasma LDLc and CVD risk (11).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [11] Igdoura, 2017, Asialoglycoprotein receptors as important mediators of plasma lipids and atherosclerosis, Curr. Opin. Lipidol. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    ASGR is a hepatocyte type II transmembrane heterodimeric glycoprotein highly conserved among mammals.

    “The asialoglycoprotein receptor (ASGR) (10) is a hepatocyte type II transmembrane heterodimeric glycoprotein highly conserved among mammals.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [10] Ashwell, 1982, Carbohydrate-specific receptors of the liver, Annu. Rev. Biochem. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    ASGRs are internalized from the cell surface via clathrin-coated vesicles into endosomes and are rapidly recycled constitutively even in the absence of a ligand.

    “ASGRs are internalized from the cell surface via clathrin-coated vesicles into endosomes and are rapidly recycled constitutively even in the absence of a ligand (11).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [11] Igdoura, 2017, Asialoglycoprotein receptors as important mediators of plasma lipids and atherosclerosis, Curr. Opin. Lipidol. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    PCSK9-deficient mice exhibit higher levels of LDLR.

    “mice lacking PCSK9, which as expected exhibit higher levels of LDLR (18)”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [18] Zaid, 2008, Proprotein convertase subtilisin/kexin type 9 (PCSK9): Hepatocyte-specific low-density lipoprotein receptor degradation and critical role in mouse liver regeneration, Hepatology source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Interpretationcitation

    Endogenous ASGR1 has two bands likely representing a major mature form of approximately 48 kDa and a minor ER-localized immature form of approximately 42 kDa.

    “Note the presence of two bands for endogenous ASGR1 (Fig. 2B), likely representing a major mature form (∼48 kDa) that excited the endoplasmic reticulum (ER) and a minor ER-localized immature form (∼42 kDa), as observed in other proteins (16, 17).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [16] Susan-Resiga, 2017, The proprotein convertase subtilisin/kexin type 9-resistant R410S low density lipoprotein receptor mutation: A novel mechanism causing familial hypercholesterolemia, J. Biol. Chem. source

      Checked before — its earlier verdict is held back until a new check runs.

    • cites [17] Essalmani, 2021, Implications of spike-glycoprotein processing at S1/S2 by furin, at S2’ by furin and/or TMPRSS2 and shedding of ACE2: Cell-to-cell fusion, cell entry and infectivity of SARS-CoV-2, bioRxiv

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

  • It may state two separate claims as one.

    Interpretationown data

    Functional cell-surface LDLR levels are reduced only by wildtype ASGR1 and require its critical carbohydrate-binding residues.

    “Thus, functional cell-surface LDLR levels are reduced only by WT ASGR1 and require its critical carbohydrate-binding residues.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    Circulating PCSK9 originating from hepatocytes binds to cell-surface LDLR and targets it to degradation in endosomes/lysosomes.

    “Circulating PCSK9 originating from hepatocytes (2) binds to cell-surface LDLR and targets it to degradation in endosomes/lysosomes (6).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [2] Seidah, 2003, The secretory proprotein convertase neural apoptosis-regulated convertase 1 (NARC-1): Liver regeneration and neuronal differentiation, Proc. Natl. Acad. Sci. U. S. A. source
    • cites [6] Seidah, 2017, The proprotein convertases in hypercholesterolemia and cardiovascular diseases: Emphasis on proprotein convertase subtilisin/kexin 9, Pharmacol. Rev. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    At steady state, approximately 40% to 60% of total cellular ASGRs are on the cell surface.

    “Thus, at steady state ∼40% to 60% of total cellular ASGRs are on the cell surface (31).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [31] Weigel, 2002, Glycans as endocytosis signals: The cases of the asialoglycoprotein and hyaluronan/chondroitin sulfate receptors, Biochim. Biophys. Acta source

  • It may state two separate claims as one.

    Interpretationown data

    Under overexpression conditions, both ASGR1 and ASGR2 can degrade LDLR, likely as homo- or hetero-oligomers.

    “These results suggest that, under overexpression conditions, both ASGR1 and ASGR2 can degrade the LDLR, likely as homo- or hetero-oligomers (10).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [10] Ashwell, 1982, Carbohydrate-specific receptors of the liver, Annu. Rev. Biochem. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    So far, the known ligands of ASGRs are soluble desialylated proteins, including von Willebrand factor and exogenous proteins such as asialofuetin and asialo-orosomucoid.

    “So far, the known ligands of ASGRs are soluble desialylated proteins, including von Willebrand factor (11) and exogenous proteins (e.g., asialofuetin, asialo-orosomucoid) (32).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [11] Igdoura, 2017, Asialoglycoprotein receptors as important mediators of plasma lipids and atherosclerosis, Curr. Opin. Lipidol. source
    • cites [32] Meier, 2000, Crystal structure of the carbohydrate recognition domain of the H1 subunit of the asialoglycoprotein receptor, J. Mol. Biol. source

  • Probably not a substantial claim.

    Premisecitation

    ASGR1 binds galactose/N-acetyl-galactosamine.

    “Since ASGR1 binds galactose/N-acetyl-galactosamine”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [32] Meier, 2000, Crystal structure of the carbohydrate recognition domain of the H1 subunit of the asialoglycoprotein receptor, J. Mol. Biol. source

  • It may state two separate claims as one.

    Interpretationown data

    Endogenous ASGR1 negatively regulates protein levels of LDLR and its functionality independently of PCSK9.

    “Thus, endogenous ASGR1 negatively regulates protein levels of LDLR and its functionality independently of PCSK9.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Interpretationown data

    Independently, PCSK9 and ASGR1 can reduce the levels of LDLR protein.

    “Thus, we can conclude that independently PCSK9 and ASGR1 can reduce the levels of LDLR protein.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    ASGR plays a critical role in serum glycoprotein homeostasis by mediating the endocytosis and lysosomal degradation of desialylated glycoproteins with exposed terminal galactose or N-acetylgalactosamine residues.

    “It consists of two subunits, ASGR1 (major) and ASGR2 (minor), and plays a critical role in serum glycoprotein homeostasis by mediating the endocytosis and lysosomal degradation of desialylated glycoproteins with exposed terminal galactose or N-acetylgalactosamine residues (11).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [11] Igdoura, 2017, Asialoglycoprotein receptors as important mediators of plasma lipids and atherosclerosis, Curr. Opin. Lipidol. source

  • It may state two separate claims as one.

    Interpretationown data

    Furin sheds ASGR1, possibly resulting in a loss of function toward LDLR.

    “In conclusion, Furin sheds ASGR1, possibly resulting in a LOF toward LDLR.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    ASGR consists of two subunits, ASGR1 (major) and ASGR2 (minor).

    “It consists of two subunits, ASGR1 (major) and ASGR2 (minor), and plays a critical role in serum glycoprotein homeostasis by mediating the endocytosis and lysosomal degradation of desialylated glycoproteins with exposed terminal galactose or N-acetylgalactosamine residues (11).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [11] Igdoura, 2017, Asialoglycoprotein receptors as important mediators of plasma lipids and atherosclerosis, Curr. Opin. Lipidol. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    Glycoprotein ASGR ligands contain terminal galactose or N-acetylgalactosamine, with the latter being the preferred residue.

    “Glycoprotein ASGR ligands contain terminal galactose or N-acetylgalactosamine, with the latter being the preferred residue (31).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [31] Weigel, 2002, Glycans as endocytosis signals: The cases of the asialoglycoprotein and hyaluronan/chondroitin sulfate receptors, Biochim. Biophys. Acta source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    In humans, the functional ASGR is composed of two subunits, ASGR1 (major, approximately 46 kDa) and ASGR2 (minor, approximately 50 kDa), that exist as homo-oligomers or hetero-oligomers in a molar ratio of approximately 3:1.

    “In humans, the functional receptor is composed of two subunits, ASGR1 (major, ∼46 kDa) and ASGR2 (minor, ∼50 kDa), that exist as homo-oligomers or hetero-oligomers in a molar ratio of ∼3:1, respectively (13, 31).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [13] Bischoff, 1987, Two asialoglycoprotein receptor polypeptides in human hepatoma cells, J. Biol. Chem. source
    • cites [31] Weigel, 2002, Glycans as endocytosis signals: The cases of the asialoglycoprotein and hyaluronan/chondroitin sulfate receptors, Biochim. Biophys. Acta source

  • Probably not a substantial claim.

    Premisecitation

    Circulating PCSK9 is responsible for most of its activity to reduce the levels of cell-surface LDLR in hepatocytes.

    “Circulating PCSK9 is responsible for most of its activity to reduce the levels of cell-surface LDLR in hepatocytes (6).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [6] Seidah, 2017, The proprotein convertases in hypercholesterolemia and cardiovascular diseases: Emphasis on proprotein convertase subtilisin/kexin 9, Pharmacol. Rev. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    Oligomerization of the ASGR subunits via their respective stalk regions occurs in the endoplasmic reticulum.

    “Oligomerization of the ASGR subunits via their respective stalk regions occurs in the ER (31).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [31] Weigel, 2002, Glycans as endocytosis signals: The cases of the asialoglycoprotein and hyaluronan/chondroitin sulfate receptors, Biochim. Biophys. Acta source

  • Probably not a substantial claim.

    Premisecitation

    The first seven members of the proprotein convertase family cleave substrates at the canonical motif R/K-Xn-R/K↓, where Xn = 0, 2, 4, or 6 spacer amino acids.

    “The first seven members of the family cleave substrates at the canonical motif R/K-Xn-R/K↓, where Xn = 0, 2, 4, or 6 spacer aa (26).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [26] Seidah, 2012, The biology and therapeutic targeting of the proprotein convertases, Nat. Rev. Drug Discov. source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Study design and conductown data

    The identity of each ASGR1 mutant was confirmed by DNA sequencing.

    “the identity of each mutant was confirmed by DNA sequencing.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Study design and conductnone given

    Immunoblots from cell lysates and liver lysates were quantified and normalized to membranes probed for α-Tubulin or β-Actin, respectively.

    “Immunoblots from cell lysates and liver lysates were quantified and normalized to membranes probed for α-Tubulin or β-Actin, respectively.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Study design and conductnone given

    For immunohistochemistry quantification, the minimum threshold was set to 20 and negative control values were subtracted.

    “For all images the minimum threshold was set to a value of 20, negative control values were subtracted, and final values were analyzed using Microsoft Excel.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Study design and conductnone given

    Statistical significance was evaluated by Student's t test, and p < 0.05 was considered significant.

    “The statistical significance was evaluated by Student’s t test, and probability values (p) <0.05 were considered significant.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim.

    Premisecitation

    ASGRs are primarily known to interact with circulating factors.

    “since ASGRs are primarily known to interact with circulating factors (11, 31).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [11] Igdoura, 2017, Asialoglycoprotein receptors as important mediators of plasma lipids and atherosclerosis, Curr. Opin. Lipidol. source
    • cites [31] Weigel, 2002, Glycans as endocytosis signals: The cases of the asialoglycoprotein and hyaluronan/chondroitin sulfate receptors, Biochim. Biophys. Acta source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Study design and conductown data

    A negative control (-Biotin) was included in cell-surface biotinylation experiments.

    “A negative control (-Biotin) was included, to which PBS was added instead of Biotin at the biotinylation step.”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Interpretationown data

    Both ASGR1-induced degradation of the LDLR and cell-surface shedding of the latter mostly target iLDLR, whereas PCSK9 targets both LDLR forms.

    “Thus, both ASGR1-induced degradation of the LDLR and the cell-surface shedding of the latter mostly target iLDLR, whereas PCSK9 targets both LDLR forms.”
    Quote in the paperSubstantial claim
63 of 63

  • The paper relies on this without support.

    Removal of plasma LDL cholesterol is primarily mediated by the LDLR located on the surface of hepatocytes.

    “The removal of plasma LDLc is primarily mediated by the low-density lipoprotein receptor (LDLR) located on the surface of hepatocytes.”

  • The paper relies on this without support.

    There are currently no studies to suggest that ASGR1 and PCSK9 act in concert to impact lipid metabolism and/or atherosclerosis by modulating the binding or uptake of LDL cholesterol via the LDLR.

    “Despite the importance of PCSK9 and ASGR1 in lipid lowering and reduced CVD risk, there are currently no studies to suggest that ASGR1 and PSCK9 act in concert to impact lipid metabolism and/or atherosclerosis by modulating the binding/uptake of LDLc via the LDLR.”

  • The paper relies on this without support.

    The ASGR1-LDLR interaction and its effect on LDLR levels observed in cell lines and mice operate similarly in humans.

    “Our data provide a mechanism for the modest reduction of LDLc in humans lacking functional ASGR1 (12).”
    “We conclude that the LDLR is a ligand of hepatic ASGR1”

  • The paper relies on this without support.

    The co-immunoprecipitation of ASGR1 and LDLR reflects a direct physical interaction between the two proteins.

    “endogenous ASGR1 and LDLR form a complex (Fig. 8, A and B)”
    “We conclude that the LDLR is a ligand of hepatic ASGR1”
4 of 4

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • ASGR1 interacts with the LDLR and directs it towards the lysosome for degradation.

    “Proprotein convertase subtilisin/kexin type 9 and the asialoglycoprotein receptor 1 both independently interact with the LDLR and direct it towards the lysosome for degradation.”

    From Post-translational regulation of the low-density lipoprotein receptor provides new targets for cholesterol regulation (2024) · cites this paper as [42]

    The Results report that ASGR1 forms a complex with LDLR independently of PCSK9, and the Discussion describes ASGR1-induced LDLR degradation (pages not available). The Discussion specifies endosomal/lysosomal degradation for PCSK9, but does not establish that ASGR1 directs LDLR to lysosomes. (LLM, conf=0.87)

    Experimental — accuracy not yet measured

  • ASGR1 binds the glycosylated ectodomain of LDLR, resulting in internalization and ultimately degradation in the lysosomes.

    “ASGR1 binds the glycosylated ectodomain of LDLR resulting in internalization and ultimately degradation in the lysosomes.”

    From The low-density lipoprotein receptor: Emerging post-transcriptional regulatory mechanisms (2025) · cites this paper as [84]

    The Results section (Fig. 8C) reports that ASGR1 binds both mature N- and O-glycosylated LDLR and immature LDLR, and the Discussion (Fig. 9) says ASGR1 promotes LDLR degradation. Our copy does not establish binding specifically to the LDLR ectodomain or show that ASGR1 causes LDLR internalization followed by lysosomal degradation. (LLM, conf=0.87)

    Experimental — accuracy not yet measured

  • ASGR1 promotes LDLR internalization and degradation, decreasing LDLR.

    “ASGR1 (Asialoglycoprotein receptor 1) Promotes LDLR internalization and degradation; Regulates SREBP2dependent signaling and expression of PCSK9 ↓ [84,85]”

    From The low-density lipoprotein receptor: Emerging post-transcriptional regulatory mechanisms (2025) · cites this paper as [84]

    The Results section on ASGR1 binding and LDLR degradation (Fig. 9; page unavailable) reports that ASGR1 reduces cell-surface and total LDLR levels, while silencing ASGR1 increases total LDLR. The paper measures internalization of LDL, but the passages do not establish that ASGR1 promotes internalization of LDLR itself. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

  • ASGR1 regulates SREBP2-dependent signaling and expression of PCSK9, decreasing LDLR.

    “ASGR1 (Asialoglycoprotein receptor 1) Promotes LDLR internalization and degradation; Regulates SREBP2dependent signaling and expression of PCSK9 ↓ [84,85]”

    From The low-density lipoprotein receptor: Emerging post-transcriptional regulatory mechanisms (2025) · cites this paper as [84]

    The Results section reports that ASGR1 reduces LDLR protein levels and enhances its degradation, supporting that part of the claim. But the Results and Discussion say ASGR1 knockdown did not change SREBP2 mRNA or PCSK9 expression, and that its effect on LDLR is independent of PCSK9. The claimed regulation of SREBP2-dependent signaling and PCSK9 expression is therefore not supported. (LLM, conf=0.90)

    Experimental — accuracy not yet measured

  • ASGR1 promotes the lysosomal degradation of LDLR.

    “Asialoglycoprotein receptor 1 (ASGR1) promotes the lysosomal degradation of LDLR (Fig. 8D).”

    From Pathways and Molecular Mechanisms Governing LDL Receptor Regulation (2025) · cites this paper as [146]

    The Results (Figure 7 and the section on ASGR1–LDLR binding; page unavailable) report that ASGR1 enhances LDLR degradation. The Discussion describes lysosomal degradation of ASGR ligands generally, but our copy does not establish that ASGR1-mediated LDLR degradation specifically occurs in lysosomes. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Figure legends use a different p-value threshold

    Minor

    The Figure 6 and 7 legends assign an asterisk to p < 0.1, while the statistical methods say only p < 0.05 was considered significant. A result marked only by that asterisk could therefore fall outside the paper's stated significance criterion.

1 of 1

Cited works that were retracted

Works this paper cites that were later retracted, from the retraction notices publishers deposit. Major when the notice came out before this paper was published.

  • Cites a retracted work: 10.1101/2021.07.02.450896

    Major

    The cited work was retracted on 2021-07-15 (notice 10.1101/2021.07.02.450896), before this paper was published (2021-10). Its authors could have known.

    Cited work 10.1101/2021.07.02.450896 · retracted 2021-07-15 · notice 10.1101/2021.07.02.450896 · this paper published 2021-10

1 of 1

Citations

Cites

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3 of the 51 references couldn’t be matched to a known paper.

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Metadata

Authors
Susan-Resiga, D., Girard, E., Essalmani, R., Roubtsova, A., Marcinkiewicz, J., Derbali, R. M., Evagelidis, A., Byun, J. H., Lebeau, P. F., Austin, R. C., & Seidah, N. G.
Affiliations
  • Montreal Clinical Research Institute (CA)
  • St. Joseph’s Healthcare Hamilton (CA)
Corresponding author
Seidah, Nabil G.
ORCID
Venue
Journal of Biological Chemistry
Publisher
Elsevier BV
Published
2021-10
Volume
297
Issue
4
Pages
101177
ISSN
0021-9258 (ISSN-L)
DOI
10.1016/j.jbc.2021.101177
PMC
PMC8479480
OpenAlex
W3198540846
Cited by
34
References
51
Citations by year
2026: 7 · 2025: 7 · 2024: 9 · 2023: 5 · 2022: 2 · 2021: 4
FWCI
2.12
Type
journal article
Language
en
Open access
Licence
Locations
  • Journal of Biological Chemistry (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • Europe PMC (PubMed Central) (publishedVersion, other-oa)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
  • Canada Research Chairs (950-231335)
  • Canada Excellence Research Chairs
  • Heart and Stroke Foundation of Canada (G-13-0003064, G-15-0009389)
  • Canadian Institutes of Health Research (148363, 173520, 74477)
  • Leducq Foundation (13 CVD 03)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biochemistry › Cholesterol and Lipid Metabolism
  • Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Lipoproteins and Cardiovascular Health
  • Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Lipid metabolism and disorders
Keywords
LDL receptor, PCSK9, furin, receptor degradation, LDL uptake, hepatic cholesterol metabolism, carbohydrate recognition, receptor glycosylation, PCSK9 knockout mice, coronary artery disease risk, non-HDL cholesterol
MeSH terms
Proprotein Convertase 9, Animals, Humans, Liver, Receptors, LDL, Mice, Knockout, Asialoglycoprotein Receptor, Furin, Mice, Hep G2 Cells, HEK293 Cells

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Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

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