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Cited by 13Protease and Inhibitor MechanismsBlood Coagulation and Thrombosis MechanismsCholesterol and Lipid Metabolism

The DOI matches its Crossref record; 68 of the 68 references it cites were found in a registry or an index; 27 of the 56 sources its claims cite were checked for support.

Identification of amino acid residues in the MT-loop of MT1-MMP critical for its ability to cleave low-density lipoprotein receptor

Open access
Wang, Maggie, Alabi, Adekunle, Gu, Hong-mei, Gill, Govind, Zhang, Ziyang, Jarad, Suha, Xia, Xiao-dan, Shen, Yishi, Wang, Gui-qing, Zhang, Da-wei·2022·Frontiers in Cardiovascular Medicine
DOI 10.3389/fcvm.2022.917238PMC PMC9452735
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Abstract

Low-density lipoprotein receptor (LDLR) mediates clearance of plasma LDL cholesterol, preventing the development of atherosclerosis. We previously demonstrated that membrane type 1-matrix metalloproteinase (MT1-MMP) cleaves LDLR and exacerbates the development of atherosclerosis. Here, we investigated determinants in LDLR and MT1-MMP that were critical for MT1-MMP-induced LDLR cleavage. We observed that deletion of various functional domains in LDLR or removal of each of the five predicted cleavage sites of MT1-MMP on LDLR did not affect MT1-MMP-induced cleavage of the receptor. Removal of the hemopexin domain or the C-terminal cytoplasmic tail of MT1-MMP also did not impair its ability to cleave LDLR. On the other hand, mutant MT1-MMP, in which the catalytic domain or the MT-loop was deleted, could not cleave LDLR. Further Ala-scanning analysis revealed an important role for Ile at position 167 of the MT-loop in MT1-MMP’s action on LDLR. Replacement of Ile167 with Ala, Thr, Glu, or Lys resulted in a marked loss of the ability to cleave LDLR, whereas mutation of Ile167 to a non-polar amino acid residue, including Leu, Val, Met, and Phe, had no effect. Therefore, our studies indicate that MT1-MMP does not require a specific cleavage site on LDLR. In contrast, an amino acid residue with a hydrophobic side chain at position 167 in the MT-loop is critical for MT1-MMP-induced LDLR cleavage.

Overview

AI snapshot

Written from the paper’s full text.

TL;DR

MT1-MMP cleaves LDLR without requiring a specific LDLR cleavage site, but it requires its catalytic domain and the MT-loop. Within the MT-loop, a hydrophobic residue at position 167—especially Ile—is critical for MT1-MMP-induced LDLR cleavage.

Key findings

  • Deletion of LDLR functional domains (ΔR1-R7, ΔEGF, ΔO-Link, Δ812) or the J.D. mutation (Y807C) did not prevent MT1-MMP-induced reduction of mature LDLR; removal of five predicted MT1-MMP cleavage sites individually also did not abolish cleavage.
  • MT1-MMP mutants lacking the catalytic domain (ΔCAT) or the MT-loop (Δ163-170) could not cleave LDLR, whereas removal of the C-terminal cytoplasmic tail (ΔC-Term) or hemopexin domain (ΔHPX) did not impair cleavage; ΔHPX appeared to reduce mature LDLR more strongly than wild-type MT1-MMP (p < 0.001).
  • Alanine scanning of the MT-loop showed that I167A significantly impaired MT1-MMP’s ability to promote LDLR degradation, while mutation of other MT-loop residues had no significant impact.
  • Replacing Ile167 with Ala, Thr, Glu, or Lys caused marked loss of LDLR cleavage, whereas replacement with non-polar residues Leu, Val, Met, or Phe retained cleavage ability, indicating a hydrophobic side chain at position 167 is required.

Methods

  • HEK293 cells were co-transfected with wild-type or mutant human LDLR and wild-type or mutant human MT1-MMP constructs generated by site-directed mutagenesis.
  • LDLR cleavage was assessed by immunoblotting of whole-cell lysates and by cell-surface biotinylation, with densitometry normalized to actin or transferrin receptor; confocal microscopy and transwell migration assays assessed MT1-MMP trafficking and cell migration.
  • Statistical analysis used GraphPad Prism 9.0 and Student’s t-test, with significance defined as p < 0.05; experiments were repeated at least three times unless indicated.

Limitations

  • Stated: the authors cannot exclude that I167A may cause a subtle structural change in the MT-loop; the mechanism by which Ile167 contributes to LDLR cleavage is unclear; and for some combined LDLR cleavage-site mutations, impaired trafficking to the plasma membrane meant the impact on MT1-MMP-induced cleavage could not be determined.
  • Evident from the text: all experiments were performed in HEK293 cells with overexpressed proteins, so findings may not reflect endogenous MT1-MMP/LDLR biology in hepatocytes or in vivo; no direct cleavage-site mapping or structural validation beyond AlphaFold modeling is reported.

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

Integrity signals

Registry

Crossref · 10.3389/fcvm.2022.917238

Cites

68 of 68 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Verification

Claims the paper makes

27 of 56 cited sources checked · 15 supported · 0 flagged · accuracy not yet measured

  • Interpretationown data

    The catalytic activity of MT1-MMP is essential for MT1-MMP-promoted LDLR shedding.

    “These findings indicate that the catalytic activity and the MT-loop are essential for MT1-MMP-promoted LDLR shedding.”
    • Also stated in: § The requirement of domains in MT1-MMP for low-density lipoprotein receptor shedding · boosted
      “reinforcing the importance of the catalytic activity of the proteinase for LDLR cleavage”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [18] Alabi, 2021, Membrane type 1 matrix metalloproteinase promotes LDL receptor shedding and accelerates the development of atherosclerosis., Nat Commun. source
  • Premisecitation

    The catalytic activity of MT1-MMP is required for LDLR shedding.

    “Our previous study demonstrates that the catalytic activity of MT1-MMP is required for LDLR shedding (18).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [18] Alabi, 2021, Membrane type 1 matrix metalloproteinase promotes LDL receptor shedding and accelerates the development of atherosclerosis., Nat Commun. source
  • Findingown data

    The sextuple, quadruple, and double mutations of predicted cleavage sites impaired LDLR trafficking to the plasma membrane.

    “These observations indicate that these mutations appeared to impair LDLR trafficking to the plasma membrane.”
    • Also stated in: § Specific sites on low-density lipoprotein receptor for MT1-MMP-induced cleavage · plain
      “We observed the premature but not the mature form of all these mutations”
    Quote in the paperSubstantial claim
  • Findingreasoning

    The levels of the catalytic dead mutation E240A were much higher than that of the wild-type and other mutant MT1-MMP since it loses the self-cleavage activity.

    “The levels of the catalytic dead mutation E240A were much higher than that of the wild-type and other mutant MT1-MMP since it loses the self-cleavage activity.”
    Quote in the paperNot a substantial claim
  • Findingown data

    Deletion of the catalytic domain of MT1-MMP virtually abolished its ability to cleave LDLR.

    “Consistently, we found that deletion of the catalytic domain of MT1-MMP virtually abolished its ability to cleave LDLR.”
    Quote in the paperSubstantial claim
  • Probably not a substantial claim. 21
  • It may state two separate claims as one. 20

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

    Premisecitation

    The MT-loop is flexible and only present in the four membrane type MMPs, but the conformation of the MT-loop is different in each membrane type MMP and in MT1-MMP complexed with or without TIMPs.

    “The MT-loop is flexible and only present in the four membrane type MMPs, but the conformation of the MT-loop is different in each membrane type MMP (61) and in MT1-MMP complexed with or without TIMPs (62–64).”
    Quote in the paperNot a substantial claim3 of 4 sources checked
    • cites [61] Lang, 2004, Crystal structure of the catalytic domain of MMP-16/MT3-MMP: Characterization of MT-MMP specific features., J Mol Biol. source

      The retrieved passages are reference-list entries and repository metadata, not the paper’s discussion of MT-loop structure. Searches for MT-loop flexibility, its presence in four membrane-type MMPs, and differing conformations found no passage in our copy that establishes the part attributed to citation (61). The separate claim about MT1-MMP with or without TIMPs is attached to citations (62–64), not this source. (LLM, conf=0.70)

      Experimental — accuracy not yet measured
    • cites [62] Grossman, 2010, The intrinsic protein flexibility of endogenous protease inhibitor TIMP-1 controls its binding interface and affects its function., Biochemistry. source
    • cites [63] Decaneto, 2017, Solvent water interactions within the active site of the membrane type I matrix metalloproteinase., Phys Chem Chem Phys. source
    • cites [64] Fernandez-Catalan, 1998, Crystal structure of the complex formed by the membrane type 1-matrix metalloproteinase with the tissue inhibitor of metalloproteinases-2, the soluble progelatinase a receptor., EMBO J. source

      The MT1-MMP catalytic domain section (p. 5240) describes the MT-loop as characteristic of MT-MMPs and flexible, and the paper reports its structure in a TIMP-2 complex. It does not establish that the loop adopts a different conformation with and without TIMPs; the Discussion says structural comparisons with the free components must await their structures. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

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

    Premisecitation

    The crystallographic structure of the catalytic domain of MT1-MMP and the predicted structure of the full-length protein by AlphaFold reveal that the MT-loop protrudes from the main structure.

    “The crystallographic structure of the catalytic domain of MT1-MMP and the predicted structure of the full-length protein by AlphaFold reveal that the MT-loop protrudes from the main structure.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [68] Jumper, 2021, Highly accurate protein structure prediction with AlphaFold., Nature. source

      The Main section (Fig. 1; page unavailable) identifies AlphaFold as a protein-structure prediction method. The paper does not report a prediction of full-length MT1-MMP, its catalytic-domain crystal structure, or an MT-loop protruding from the protein; searches for MT1-MMP and MT-loop found no such passage. (LLM, conf=0.83)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim.

    Premisecitation

    CleavePredict is a validated free access web server for predicting the substrate cleavage pattern by matrix metalloproteinases (MMP).

    “CleavePredict is a validated free access web server for predicting the substrate cleavage pattern by matrix metalloproteinases (MMP).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [45] Kumar, 2015, CleavPredict: A platform for reasoning about matrix metalloproteinases proteolytic events., PLoS One. source

  • It may state two separate claims as one.

    Premisecitation

    Specific targeting of hepatocytes can be achieved using GalNAc-conjugation, such as GalNAc-siRNA or antisense oligonucleotides, which has been successfully used to silence hepatic PCSK9 expression in a clinical trial.

    “Specific targeting hepatocytes can be archived using N-acetylgalactosamine (GalNAc)-conjugation, such as GalNAc-siRNA or antisense oligonucleotides (ASOs) (50–52), which has been successfully used to silence hepatic PCSK9 expression in a clinical trial (51).”
    Quote in the paperSubstantial claim3 of 3 sources checked
    • cites [50] Springer, 2018, GalNAc-siRNA conjugates: Leading the way for delivery of RNAi therapeutics., Nucleic Acid Ther. source
    • cites [51] Cupido, 2020, Inclisiran for the treatment of hypercholesterolaemia: Implications and unanswered questions., Cardiovasc Res. source
    • cites [52] Debacker, 2020, Delivery of oligonucleotides to the liver with GalNAc: From research to registered therapeutic drug., Mol Ther. source

  • Probably not a substantial claim.

    Premisecitation

    The HPX domain contributes to the interaction of MT1-MMP with its substrates, such as CD44 and collagen.

    “The HPX domain contributes to the interaction of MT1-MMP with its substrates, such as CD44 and collagen (54–57).”
    Quote in the paperSubstantial claim4 of 4 sources checked
    • cites [54] Tam, 2004, Characterization of the distinct collagen binding, helicase and cleavage mechanisms of matrix metalloproteinase 2 and 14 (gelatinase A and MT1-MMP): The differential roles of the MMP hemopexin c domains and the MMP-2 fibronectin type II modules in collagen triple helicase activities., J Biol Chem. source

      The Results (page not provided) state that the MT1-MMP hemopexin C domain binds collagen and is required for native collagen cleavage, supporting the collagen example. No passage in our copy addresses an interaction with CD44, so that example is not supported by this source. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [55] Marcink, 2019, MT1-MMP binds membranes by opposite tips of its beta propeller to position it for pericellular proteolysis., Structure. source
    • cites [56] Van Doren, 2017, Peripheral membrane associations of matrix metalloproteinases., Biochim Biophys Acta Mol Cell Res. source
    • cites [57] Zarrabi, 2011, Inhibition of matrix metalloproteinase 14 (MMP-14)-mediated cancer cell migration., J Biol Chem. source

      The section on PEX-domain motifs reports that a blade I mutant failed to coprecipitate with CD44, supporting a role for the domain in the MMP-14–CD44 interaction. The Discussion also describes CD44 cleavage by MMP-14. Our copy does not establish that the PEX domain contributes to interaction with collagen; searches for collagen binding and substrate interaction did not find such a passage. No page number was available in the indexed copy. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • Rests on an assumption: Exosites are a promising target for the development of specific MMP inhibitors.

  • It may state two separate claims as one.

    Premisecitation

    Hepatic MT1-MMP stimulates LDLR shedding, thereby increasing plasma LDL-C levels and the development of atherosclerosis.

    “Our previous study demonstrates that hepatic MT1-MMP stimulates LDLR shedding, thereby increasing plasma LDL-C levels and the development of atherosclerosis (18).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [18] Alabi, 2021, Membrane type 1 matrix metalloproteinase promotes LDL receptor shedding and accelerates the development of atherosclerosis., Nat Commun. source

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

    Premisecitation

    The cytoplasmic tail of MT1-MMP plays an important role in endocytosis of MT1-MMP and the localization of MT1-MMP on specific microdomains in the plasma membrane.

    “The cytoplasmic tail of MT1-MMP plays an important role in endocytosis of MT1-MMP and the localization of MT1-MMP on the specific microdomains in the plasma membrane (19, 23–25).”
    Quote in the paperNot a substantial claim3 of 4 sources checked
    • cites [19] Xia, 2021, Membrane-type I matrix metalloproteinase (MT1-MMP), lipid metabolism and therapeutic implications., J Mol Cell Biol. source
    • cites [23] Houghton, 2015, Matrix metalloproteinases in destructive lung disease., Matrix Biol. source

      The Introduction discusses MMP functions in emphysema, and the MMP-12 section discusses a different enzyme’s activity, but neither addresses MT1-MMP trafficking. Searches for MT1-MMP, its cytoplasmic tail, endocytosis, and plasma-membrane microdomains found no passage supporting the attributed claim. (LLM, conf=0.74)

      Experimental — accuracy not yet measured
    • cites [24] Castro-Castro, 2016, Cellular and molecular mechanisms of MT1-MMP-dependent cancer cell invasion., Annu Rev Cell Dev Biol. source
    • cites [25] Feinberg, 2018, Divergent matrix-remodeling strategies distinguish developmental from neoplastic mammary epithelial cell invasion programs., Dev Cell. source

      The Results and Discussion passages address MT1-MMP’s role in collagen remodeling and mammary invasion, not the function of its cytoplasmic tail. Searches for “cytoplasmic tail,” “endocytosis,” and “microdomains” found no passage in our copy connecting the tail to endocytosis or localization at plasma-membrane microdomains; page numbers were unavailable. (LLM, conf=0.76)

      Experimental — accuracy not yet measured

  • It may state two separate claims as one.

    Premisecitation

    Exosites outside the catalytic core domain of MMPs are less conserved and contribute to substrate selection and binding.

    “exosites that are regions outside the catalytic core domain of MMPs are less conserved and contribute to substrate selection and binding (27–29).”
    Quote in the paperSubstantial claim1 of 3 sources checked
    • cites [27] Mori, 2013, Transmembrane/cytoplasmic, rather than catalytic, domains of Mmp14 signal to MAPK activation and mammary branching morphogenesis via binding to integrin beta1., Development. source
    • cites [28] Yanez-Mo, 2008, MT1-MMP collagenolytic activity is regulated through association with tetraspanin CD151 in primary endothelial cells., Blood. source
    • cites [29] Mori, 2002, CD44 directs membrane-type 1 matrix metalloproteinase to lamellipodia by associating with its hemopexin-like domain., EMBO J. source

      The section “The PEX domain of MT1-MMP binds directly to the stem region of CD44H” (page unavailable) shows that a domain outside the catalytic core binds CD44H; the discussion also says that domain is indispensable for CD44H shedding. Our copy does not establish that exosites are less conserved or that they generally govern substrate selection across MMPs. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • It may state two separate claims as one.

    Premisecitation

    MT1-MMP promotes ectodomain shedding of LDLR, thereby increasing plasma LDL-C levels in mice.

    “In our previous studies, we found that membrane type 1-matrix metalloproteinase (MT1-MMP) promotes ectodomain shedding of LDLR, thereby increasing plasma LDL-C levels and exacerbating the development of atherosclerosis in mice (18, 19).”
    Quote in the paperSubstantial claim2 of 2 sources checked
    • cites [18] Alabi, 2021, Membrane type 1 matrix metalloproteinase promotes LDL receptor shedding and accelerates the development of atherosclerosis., Nat Commun. source
    • cites [19] Xia, 2021, Membrane-type I matrix metalloproteinase (MT1-MMP), lipid metabolism and therapeutic implications., J Mol Cell Biol. source

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

    Premisecitation

    The NPVY motif within the C-terminus is critical for internalization of LDLR into clathrin-coated pits.

    “The NPVY motif within the C-terminus is critical for the internalization of the receptor into clathrin-coated pits (2–4).”
    Quote in the paperNot a substantial claim1 of 3 sources checked
    • cites [2] Rudenko, 2002, Structure of the LDL receptor extracellular domain at endosomal pH., Science. source

      The ‘Extracellular Domain at Endosomal pH’ section says LDLR mediates endocytosis, but it does not address what directs the receptor into clathrin-coated pits. Searches for NPVY, the C-terminal motif, and clathrin-coated pits found no passage establishing the motif’s role; page numbers are unavailable in our copy. (LLM, conf=0.72)

      Experimental — accuracy not yet measured
    • cites [3] Davis, 1986, The J.D. mutation in familial hypercholesterolemia: Amino acid substitution in cytoplasmic domain impedes internalization of LDL receptors., Cell. source
    • cites [4] Davis, 1987, The low density lipoprotein receptor. Identification of amino acids in cytoplasmic domain required for rapid endocytosis., J Biol Chem. source

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

    Premisecitation

    An antibody specifically against the MT-loop has been developed and can block binding of pro-MMP2 to MT1-MMP and inhibit pro-MMP2 activation.

    “Indeed, an antibody specifically against the MT-loop has been developed and can block binding of pro-MMP2 to MT1-MMP and inhibit pro-MMP2 activation (67).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [67] Shiryaev, 2013, A monoclonal antibody interferes with TIMP-2 binding and incapacitates the MMP-2-activating function of multifunctional, pro-tumorigenic MMP-14/MT1-MMP., Oncogenesis. source

      The Abstract says antibody 9E8 recognizes the MT-loop and inhibits MT1-MMP-dependent MMP-2 activation. The Discussion explains that it blocks TIMP-2 binding to MT1-MMP; pro-MMP-2 binds TIMP-2 to form the activation complex. Thus the paper supports impaired pro-MMP-2 recruitment, but not the claimed direct blockade of pro-MMP-2 binding to MT1-MMP. (LLM, conf=0.88)

      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

    Weaver et al. reported that the MT-loop, together with the hemopexin domain, is required for the translocation of MT1-MMP from the apical to the basal membrane in polarized epithelial cells during tubulogenesis, but deletion of the entire MT-loop has no significant effect on the expression, trafficking, processing, or the proteolytic activity of MT1-MMP.

    “Weaver et al. reported that the MT-loop, together with the hemopexin domain, is required for the translocation of MT1-MMP from the apical to the basal membrane in polarized epithelial cells during tubulogenesis (65), but deletion of the entire MT-loop has no significant effect on the expression, trafficking, processing, or the proteolytic activity of MT1-MMP (61, 66).”
    Quote in the paperNot a substantial claim3 of 3 sources checked
    • cites [65] Weaver, 2014, Basal localization of MT1-MMP is essential for epithelial cell morphogenesis in 3D collagen matrix., J Cell Sci. source
    • cites [61] Lang, 2004, Crystal structure of the catalytic domain of MMP-16/MT3-MMP: Characterization of MT-MMP specific features., J Mol Biol. source

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

      Experimental — accuracy not yet measured
    • cites [66] English, 2001, Characterization of the role of the “MT-loop”: An eight-amino acid insertion specific to progelatinase A (MMP2) activating membrane-type matrix metalloproteinases., J Biol Chem. source

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

      Experimental — accuracy not yet measured

  • Probably not a substantial claim.

    Premisecitation

    The catalytic domain is highly conserved among MMP family members.

    “The catalytic domain is highly conserved among MMP family members (19, 24).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [19] Xia, 2021, Membrane-type I matrix metalloproteinase (MT1-MMP), lipid metabolism and therapeutic implications., J Mol Cell Biol. source
    • cites [24] Castro-Castro, 2016, Cellular and molecular mechanisms of MT1-MMP-dependent cancer cell invasion., Annu Rev Cell Dev Biol. source

  • It may state two separate claims as one.

    Findingown data

    The relative cell numbers migrated through collagen type I-coated inserts were comparable in cells transfected with wild-type MT1-MMP and MT1-MMPI167A; both significantly promoted cell migration compared to control.

    “We observed that the relative cell numbers migrated through collagen type I-coated inserts were comparable in cells transfected with the wild-type MT1-MMP and MT1-MMPI167A; both significantly promoted cell migration compared to the control (Figure 5B).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Ile167 is situated in the middle of a small α-helix in the MT-loop with its hydrophobic side chain positioning inside the loop.

    “Ile167 is situated in the middle of a small α-helix in the MT-loop with its hydrophobic side chain positioning inside the loop (Figure 7).”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    Mutations I167E, I167K and I167T significantly impaired MT1-MMP-induced LDLR cleavage, whereas substitution of Ile with a hydrophobic residue (I167V, I167L, I167M, and I167F) retained the ability to cleave LDLR.

    “As shown in Figures 6A,B, like I167A, mutations I167E, I167K and I167T significantly impaired MT1-MMP-induced LDLR cleavage, whereas substitution of Ile with a hydrophobic residue (mutations I167V, I167L, I167M, and I167F) retained the ability to cleave LDLR.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: Ile within the MT-loop of MT1-MMP is highly conserved in different species except for alligators and turtles, which have valine instead.

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

    Findingnone given

    The MT-loop consists of seven amino acid residues and is present in all four membrane type MMPs (MT1-, MT2-, MT3-, and MT5-MMP), except the two glycosyl phosphatidylinositol-anchored membrane-associated MMPs (MT4- and MT6-MMP).

    “The MT-loop consists of seven amino acid residues and is present in all four membrane type MMPs (MT1-, MT2-, MT3-, and MT5-MMP), except the two glycosyl phosphatidylinositol-anchored membrane-associated MMPs (MT4- and MT6-MMP) (Figure 4A).”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    Substitution of Ile167 with Ala (MT1-MMPI167A) significantly impaired MT1-MMP's ability to promote LDLR degradation, while mutation of other MT-loop amino acid residues had no significant impact.

    “We found that substitution of Ile167 with Ala (MT1-MMPI167A), like E240A, significantly impaired MT1-MMP’s ability to promote LDLR degradation, while mutation of other amino acid residues had no significant impact.”
    • Also stated in: § Mutational analysis of the MT-loop · plain
      “Consistently, biotinylation experiments showed that I167A and E240A could not effectively reduce the levels of the mature form of LDLR when compared to the wild-type and other mutant MT1-MMP (Figure 4D).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. The paper may give no reason why it holds.

    Findingnone given

    Pro163 and Tyr164 in the MT-loop are completely conserved.

    “Pro163 and Tyr164 in the MT-loop are completely conserved.”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Premisecitation

    Deletion of the cytoplasmic tail did not affect the trafficking of MT1-MMP to the plasma membrane nor its ability to activate proMMP2, but the mutant proteins displayed a different distribution pattern on the cell surface and an impaired ability to mediate cell invasion compared to the wild-type protein.

    “Deletion of the cytoplasmic tail did not affect the trafficking of MT1-MMP to the plasma membrane nor its ability to activate proMMP2, but the mutant proteins displayed a different distribution pattern on the cell surface and an impaired ability to mediate cell invasion compared to the wild-type protein (59, 60).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [59] Lehti, 2000, Regulation of membrane-type-1 matrix metalloproteinase activity by its cytoplasmic domain., J Biol Chem. source

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

    • cites [60] Rozanov, 2004, Aberrant, persistent inclusion into lipid rafts limits the tumorigenic function of membrane type-1 matrix metalloproteinase in malignant cells., Exp Cell Res. source

  • Probably not a substantial claim.

    Premisecitation

    Plasma levels of soluble LDLR (sLDLR) are positively correlated with circulating LDL-C levels.

    “It has been reported that plasma levels of sLDLR are positively correlated to circulating LDL-C levels (15–17).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [15] Mayne, 2007, Plasma PCSK9 levels correlate with cholesterol in men but not in women., Biochem Biophys Res Commun. source
    • cites [16] Shimohiro, 2014, Association between serum soluble low-density lipoprotein receptor levels and metabolic factors in healthy Japanese individuals., J Clin Lab Anal. source
    • cites [17] Mayne, 2018, Associations between soluble LDLR and lipoproteins in a Caucasian cohort and the effect of PCSK9 loss-of-function., J Clin Endocrinol Metab. source

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

    Premisecitation

    No consensus cleavage site for MMPs has been identified, but MMPs are believed to preferentially cleave specific substrates at certain locations containing specific amino acid residues.

    “While no consensus cleavage site for MMPs has been identified, it is believed that MMPs preferentially cleave specific substrates at certain locations containing specific amino acid residues (53).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [53] Eckhard, 2016, Active site specificity profiling of the matrix metalloproteinase family: Proteomic identification of 4300 cleavage sites by nine MMPs explored with structural and synthetic peptide cleavage analyses., Matrix Biol. 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

    Mutations in LDLR cause familial hypercholesterolemia (FH).

    “Mutations in LDLR cause familial hypercholesterolemia (FH), characterized by elevated levels of plasma LDL-C and increased risk of coronary heart disease (1).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [1] Goldstein, 2015, A century of cholesterol and coronaries: From plaques to genes to statins., Cell. source

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

    Premisecitation

    Targeting HPX with a small inhibitor can selectively inhibit MT1-MMP activity and suppress cancer cell growth.

    “Targeting HPX with a small inhibitor can selectively inhibit MT1-MMP activity and suppress cancer cell growth (58).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [58] Fields, 2015, New strategies for targeting matrix metalloproteinases., Matrix Biol. source

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

    • Rests on an assumption: Exosites are a promising target for the development of specific MMP inhibitors.

  • It may state two separate claims as one.

    Interpretationown data

    Substitution of Ile167 with Ala does not appear to affect MT1-MMP trafficking or its ability to promote cell migration.

    “Therefore, substitution of Ile167 with Ala does not appear to affect MT1-MMP trafficking or its ability to promote cell migration.”
    • Also stated in: § Discussion · plain
      “We also did not observe a significant difference in cell migration and MT1-MMP trafficking between the wild-type and I167A mutant MT1-MMP.”
    Quote in the paperSubstantial claim

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

    Study design and conductcitation

    The antibody can recognize the full-length and self-cleaved MT1-MMP.

    “The antibody can recognize the full-length and self-cleaved MT1-MMP, consistent with a previous report (47).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [47] Rozanov, 2003, Membrane type-1 matrix metalloproteinase functions as a proprotein self-convertase. Expression of the latent zymogen in Pichia pastoris, autolytic activation, and the peptide sequence of the cleavage forms., J Biol Chem. source

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

    Premisecitation

    MT1-MMP in vascular smooth muscle cells regulates extracellular matrix homeostasis and plays an important role in maintaining normal VSMC function.

    “On the other hand, MT1-MMP in vascular smooth muscle cells (VSMC) regulates extracellular matrix homeostasis and plays an important role in maintaining normal VSMC function (49).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [49] Barnes, 2017, Membrane-tethered metalloproteinase expressed by vascular smooth muscle cells limits the progression of proliferative atherosclerotic lesions., J Am Heart Assoc. 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.

    Study design and conductnone given

    Values were mean ± S.D. of ≥3 experiments.

    “Values were mean ± S.D. of ≥3 experiments.”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Interpretationown data

    Each of the LDLR regions and endocytosis of LDLR are not required for MT1-MMP's action on the receptor.

    “Therefore, each of these regions and endocytosis of LDLR are not required for MT1-MMP’s action on the receptor.”
    Quote in the paperSubstantial claim

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

    Premisecitation

    Macrophage MT1-MMP degrades collagen in atherosclerotic plaques and consequently increases plaque vulnerability.

    “In addition, it has been reported that macrophage MT1-MMP degrades collagen in atherosclerotic plaques and consequently increases plaque vulnerability (48).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [48] Schneider, 2008, Matrix-metalloproteinase-14 deficiency in bone-marrow-derived cells promotes collagen accumulation in mouse atherosclerotic plaques., Circulation. source

  • Probably not a substantial claim. It may be too vague to check against evidence.

    Interpretationreasoning

    The MT-loop represents a potential target for the development of selective MT1-MMP inhibitors due to its specificity and structurally easy accessibility.

    “Nevertheless, the MT-loop represents a potential target for the development of selective MT1-MMP inhibitors due to its specificity and structurally easy accessibility.”
    Quote in the paperNot a substantial claim

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

    Study design and conductown data

    Similar results were observed in three different experiments.

    “Similar results were observed in three different experiments.”
    Quote in the paperNot a substantial claim
32 of 32

  • The paper relies on this without support.

    The MT-loop is specific for MT1-MMP.

    “Given that the MT-loop is specific for MT1-MMP, our findings provide a foundation for the future design of inhibitors that can specifically target MT1-MMP’s action on LDLR.”

  • The paper relies on this without support.

    The ectodomain of LDLR can be cleaved by proteases to generate a soluble form of LDLR (sLDLR), which inactivates LDLR-mediated LDL clearance.

    “The ectodomain of LDLR can be cleaved by proteases to generate a soluble form of LDLR (sLDLR), which inactivates LDLR-mediated LDL clearance.”

  • The paper relies on this without support.

    The five selected predicted cleavage sites are the relevant ones for MT1-MMP cleavage on LDLR.

    “Based on the position weight matrix score and proximity to the transmembrane domain, which has been suggested to be the cleavage region of MT1-MMP on LDL receptor related protein 1 (LRP1) (46), we selected 5 locations A521, G529, G565, and N645 within the YWTD region and A789 within the transmembrane domain near the outer layer of plasma membrane.”

  • The paper relies on this without support.

    Ile within the MT-loop of MT1-MMP is highly conserved in different species except for alligators and turtles, which have valine instead.

    “Ile within the MT-loop of MT1-MMP is highly conserved in different species except for alligators and turtles. Instead of Ile, they have another hydrophobic residue, valine.”

  • The paper relies on this without support.

    The catalytic core region is highly conserved among 23 MMP family members, making specific inhibition of MT1-MMP proteolytic activity by targeting this domain very challenging.

    “It is of note that the catalytic core region is highly conserved among 23 MMP family members, which makes it very challenging to specifically inhibit the proteolytic activity of MT1-MMP by targeting this functional domain.”

  • The paper relies on this without support.

    Exosites are a promising target for the development of specific MMP inhibitors.

    “Exosites have been shown to be a promising target for the development of specific MMP inhibitors.”
6 of 6

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?

  • Two zinc metalloproteases, MT1-MMP and BMP1, also cleave the LDLR.

    “In addition to the proteolytic cleavage of the LDLR by γ-secretase, two zinc metalloproteases also cleave the receptor, MT1-MMP and BMP1.”

    From Post-translational regulation of the low-density lipoprotein receptor provides new targets for cholesterol regulation (2024) · cites this paper as [56]

    The Introduction and Results identify MT1-MMP as a zinc-dependent metalloproteinase that cleaves LDLR. Our copy does not identify BMP1 as another LDLR-cleaving protease, so it supports only the MT1-MMP part of the claim. (LLM, conf=0.87)

    Experimental — accuracy not yet measured

  • MT1-MMP cleaves the LDLR adjacent to the plasma membrane.

    “MT1-MMP cleaves the LDLR adjacent to the plasma membrane (Figures 1H and 2) [56].”

    From Post-translational regulation of the low-density lipoprotein receptor provides new targets for cholesterol regulation (2024) · cites this paper as [56]

    The Introduction reports that MT1-MMP promotes shedding of the LDLR ectodomain. In Results, the authors examine a predicted cleavage site near the outer layer of the plasma membrane, but MT1-MMP still reduces LDLR when that site is mutated; the paper does not establish that cleavage occurs adjacent to the membrane. Page numbers are unavailable in our copy. (LLM, conf=0.87)

    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.

  • MT-loop is called MT1-MMP-specific but also present in other MT-MMPs

    Major

    The proposed specificity of targeting the MT-loop rests in part on calling it specific to MT1-MMP. The results instead state that the loop is present in MT2-, MT3-, and MT5-MMP as well.

  • I556A and I566A are used for the same described mutant

    Minor

    The site-disruption description specifies I566A together with T567A, but the results name the paired mutant I556AT567A. The two residue positions differ.

  • The stated MT-loop range contains eight positions, not seven

    Minor

    The inclusive residue range 163-170 contains eight positions, whereas the results describe the MT-loop as consisting of seven amino acid residues.

  • The stated HA-tag position conflicts with the ΔHPX tag assignment

    Minor

    The methods place the MT1-MMP HA-tag between residues 115 and 116, whereas the ΔHPX deletion is described as spanning residues 312 to 535. That deletion would not remove the stated tag, contrary to the claim that ΔHPX lacked it.

4 of 4

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Metadata

Authors
Wang, M., Alabi, A., Gu, H.-M., Gill, G., Zhang, Z., Jarad, S., Xia, X.-D., Shen, Y., Wang, G.-Q., & Zhang, D.-W.
Affiliations
  • University of Alberta (CA)
  • Guangzhou Medical University (CN)
ORCID
Venue
Frontiers in Cardiovascular Medicine
Publisher
Frontiers Media SA
Published
2022-08-25
Volume
9
Pages
917238-917238
ISSN
2297-055X (ISSN-L)
DOI
10.3389/fcvm.2022.917238
PMC
PMC9452735
OpenAlex
W4293222313
Cited by
13
References
68
Citations by year
2026: 1 · 2025: 1 · 2024: 7 · 2023: 1 · 2022: 3
FWCI
1.06
Type
journal article
Language
en
Open access
Licence
Locations
  • Frontiers in Cardiovascular Medicine (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • Canadian Institutes of Health Research (PS 155994)
  • Natural Sciences and Engineering Research Council of Canada
  • National Natural Science Foundation of China
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Protease and Inhibitor Mechanisms
  • Health Sciences › Medicine › Hematology › Blood Coagulation and Thrombosis Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biochemistry › Cholesterol and Lipid Metabolism
Keywords
MT1-MMP, LDL receptor, receptor cleavage, atherosclerosis, hemopexin domain, catalytic domain

About these checks

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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