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Membrane Type 1 Matrix Metalloproteinase Cleaves Low Density Lipoprotein Receptor to Regulate Lipid Metabolism
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- Author / Creator
- Wang, Maggie
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Low-density lipoprotein receptor (LDLR) mediates clearance of plasma LDL cholesterol
(LDL-C) and protects against the development of atherosclerosis. Our previous studies
demonstrated that membrane type-1 matrix metalloproteinase (MT1-MMP/MMP14) cleaves LDL
receptor (LDLR) and exacerbates the development of atherosclerosis. In this study, we
investigated the determinants in LDLR and MT1-MMP that were critical for MT1-MMP-induced
LDLR cleavage. We performed site-directed mutagenesis to generate LDLR and MT1-MMP
mutations and observed that deletion of various functional domains in LDLR, including the ligand binding repeats (LBR), the epidermal growth factor precursor homology domain, the clustered O-glycosylation region, and the C-terminal cytoplasmic tail, did not affect MT1-MMP induced
cleavage of the receptor. Furthermore, removal of four predicted MT1-MMP cleavage sites on
LDLR had no effect on cleavage by MT1-MMP. In addition, we observed multiple cleaved
fragments of LDLR in the cell culture medium. In MT1-MMP, deletion of the catalytic domain or
the MT-loop, but not the hemopexin (HPX) domain or the C-terminal cytoplasmic tail of MT1-
MMP, significantly decreased MT1-MMP’s ability to cleave LDLR. Further Ala-scanning
analysis revealed the important role of Ile at position 167 within the MT-loop in MT1-MMP’s
action on LDLR. Replacement of Ile167 with polar residues in MT1-MMP caused a significant
loss in LDLR cleavage, whereas mutation of Ile167 to a non-polar amino acid residue had no effect.
In addition, MT1-MMP cleaves extracellular matrix (ECM) components, such as type I
collagen, and plays a critical role in maintaining homeostasis of ECM. Considering the crucial role
of ECM in the progression of liver fibrosis, we studied MT1-MMP’s effects on this process. To
do so, we fed MT1-MMP hepatocyte specific knockout (Mmp14LKO) mice and the controlMmp14Flox mice with a high fat, high fructose, and high cholesterol Gubra amylin liver NASH
(GAN) diet for 42 weeks. We did not observe any significant difference between Mmp14LKO and
Mmp14Flox mice in body weight, blood glucose levels, plasma levels of total cholesterol (TC), high
density lipoprotein cholesterol (HDL-C), non-HDL-C, and triglyceride (TG). The ratio of liver
weight to body weight, Oil-Red O staining, and hematoxylin and eosin (H&E) staining were also
comparable in the two genotypes. Histologically, the liver samples of both Mmp14LKO and
Mmp14Flox mice did not show notable fibrosis even though they appeared to have serious steatosis.
Analysis of liver lipid levels revealed that lacking hepatic MT1-MMP did not change TC and TG
levels. mRNA levels of genes encoding factors important for lipid metabolism (Srebp1c, Fasn,
Srebp2, Pcsk9, Ppara, and Cpt1a), inflammation (Tnfa, Il1b), and fibrosis (Tgfb, Col1a1) were
also not altered in the liver of Mmp14LKO mice compared to that of Mmp14Flox mice.
In conclusion, our studies indicate that MT1-MMP does not require a specific cleavage site
on LDLR and may cleave LDLR at multiple sites. However, an amino acid residue with a
hydrophobic side chain at position 167 in MT-loop is critical for MT1-MMP-induced LDLR
cleavage. Furthermore, lacking hepatic MT1-MMP does not appear to alter the development of
liver steatosis in mice or promote liver fibrosis in mice fed the GAN diet. These data combined
with our previous findings suggest that MT1-MMP may be a promising therapeutic target to lower
plasma cholesterol levels and pave the way for the design of highly selective MT1-MMP inhibitors. -
- Graduation date
- Fall 2022
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- Type of Item
- Thesis
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- Degree
- Master of Science
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- License
- This thesis is made available by the University of Alberta Library with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.