Overview

Cell and molecular biology, lipoprotein receptors, lipoprotein and cholesterol metabolism, intracellular protein sorting, golgi function, somatic cell genetics, atherosclerosis, scavenger receptors, pathogen receptors, macrophage physiology, pattern recognition in vertebrate and invertebrate immune systems.

Research Summary

We are using genetic, biochemical, physiologic, chemical genetics and cell and molecular biological methods to study cell surface receptor structure and function. We are focused on two categories of lipoprotein receptors because of their relevance to human biology and medicine. They are the low density lipoprotein (LDL) receptor and three classes of scavenger receptors, including the HDL receptor SR-BI. The analysis of these systems should help provide insights into basic biological processes and contribute to our understanding of atherosclerosis and coronary heart disease (CHD). The risk of developing atherosclerosis is directly related to plasma levels of LDL cholesterol and inversely related to those of high density lipoprotein (HDL) cholesterol.

The LDL Receptor: LDL is the principal cholesterol carrier in human plasma. Plasma concentrations of LDL are critically regulated by LDL receptors, which mediate the sterol regulated endocytosis of LDL via coated pits, coated vesicles, endosomes and lysosomes. LDL cholesterol is released into the cell after the LDL is degraded by hydrolytic enzymes in lysosomes. Genetic defects in the LDL receptor gene lead to hypercholesterolemia and premature CHD.

We hope to define in molecular detail the pathways of receptor synthesis and processing, membrane protein sorting and endocytic recycling, with special focus on the structure and function of the Golgi apparatus, by isolating and characterizing mammalian cells carrying mutations in structural and regulatory genes responsible for receptor function. These mutants will help define the gene products and functions required for LDL receptor activity. We have isolated a large collection of mutant Chinese hamster ovary (CHO) cells which defines 9 genetic complementation groups (ldlA-ldlI) containing both constitutive and temperature-sensitive, conditional-lethal mutants. By employing biochemical, immunological, and recombinant DNA techniques, we hope to gain further insight into the mechanisms underlying membrane protein structure and function. For example we have used ldlB and ldlC mutants to identify and characterize the COG (conserved oligomeric Golgi) complex – an octomer – that is important for determining the structure and function of the Golgi apparatus. We have also identified a set of COG-sensitive Golgi-associated proteins, called GEARs, the analysis of which is likely to provide new insights into resident Golgi protein localization mechanisms. Based on our analysis of the CHO cell mutants, we proposed and helped show that mutations in COG subunits (Cog1 and Cog7) are responsible for some cases of the inherited human disease Congenital Disorders of Glycosylation. In addition, analysis of the ldlG mutant has forced a re-evaluation of the role of the function of the Golgin GM130 in Golgi function and membrane trafficking.

Scavenger Receptors: Scavenger receptors (SR) are multiligand cell surface receptors defined by their ability to bind modified lipoproteins (e.g., acetylated [Ac] or oxidized [Ox] LDL) with high affinity; however, they bind many other classes of ligands. We have used biochemical and molecular genetic methods to identify and characterize three structurally distinct classes of SR: SR-A, SR-B and SR-C. There are three subtypes of SR-A, which are expressed primarily in mammalian macrophages, and 4 subtypes of SR-C, one of which is expressed in Drosophila melanogaster macrophages. SR-As and SR-CI can both bind a wide variety of polyanions, including chemically modified proteins, some sulfated polysaccharides, the microbial toxins endotoxin and lipoteichoic acid, asbestos, and certain polynucleotides (e.g., poly I, but not poly C). The broad binding specificity of these macrophage scavenger receptors suggest that they may play a role in a variety of macrophage-related physiologic and pathologic systems, e.g., macrophage-associated immune responses (self vs. nonself recognition) and inflammation. Studies of SR-AI/II knockout mice support this suggestion.

The SR-Bs, including SR-BI and CD36, are members of the CD36 superfamily of proteins and are expressed on a variety of cell types and tightly bind anionic phospholipids. SR-BI can bind LDL and HDL. Indeed, it was the first physiologically relevant HDL receptor to be described. SR-BI is expressed primarily in liver and steroidogenic tissues. SR-BI, which in some cells is clustered in lipid rafts (e.g., caveolae) rather than coated pits, mediates selective cholesterol uptake from HDL by a mechanism distinct from the classic LDL receptor endocytic pathway.

After HDL binds to SR-BI, cholesterol is transferred to the cells and the cholesterol-depleted particle is released into the extracellular space – there is no lysosomal degradation. We are using biochemical, biophysical, physiologic, molecular genetic and ‘chemical genetic’ approaches to study the structure and mechanism of action of SR-BI. For example, we have isolated small molecules that specifically block SR-BI-mediated lipid transfer without decreasing lipoprotein binding and a variety of mutant forms of the receptor (e.g., ones that bind LDL but not HDL). We have shown that SR-BI expression is coordinately regulated with cholesterol metabolism (e.g., steroidogeneis) and that genetic manipulation of its expression can significantly influence plasma HDL and biliary cholesterol concentrations. It also influences the development of atherosclerosis coronary heart disease, female fertility and red blood cell maturation by mechanisms currently under study. We have generated SR-BI homozygous null (knockout) mice and exploited their characteristics to generate several novel murine models of fatal occlusive atherosclerotic coronary heart disease (CHD). These new models appear to be especially attractive for the analysis of the molecular and cellular mechanisms underlying CHD and for developing new pharmacologic and genetic therapies for the prevention and/or treatment of cardiovascular disease. If SR-BI’s functions in humans are similar to those in mice, SR-BI will become an attractive target for therapeutic intervention in a variety of diseases.

Selected Publications

Foulquier F, Ungar D, Reynders E, Zeevaert R, Mills P, Garcia-Silva MT, Briones P, Winchester B, Morelle W, Krieger M, Annaert W, Matthijs G. A new inborn error of glycosylation due to a Cog8 deficiency reveals a critical role for the Cog1-Cog8 interaction in COG complex formation. Hum Mol Genet. Apr 1;16(7):717-30 (2007)

Choi MY, Romer AI, Hu M, Lepourcelet M, Mechoor A, Yesilaltay A, Krieger M, Gray PA, Shivdasani RA.A dynamic expression survey identifies transcription factors relevant in mouse digestive tract development. Development. Oct;133(20):4119-29. (2006)

Yesilaltay A, Kocher O, Pal R, Leiva A, Quinones V, Rigotti A, Krieger M. PDZK1 is required for maintaining hepatic scavenger receptor, class B, type I (SR-BI) steady state levels but not its surface localization or function. J Biol Chem. Sep 29;281(39): 28975-80. Epub 2006 Jul 25. (2006)

Vasile E, Oka T, Ericsson M, Nakamura N, Krieger M. IntraGolgi distribution of the Conserved Oligomeric Golgi (COG) complex. Exp Cell Res.Oct 1;312(16):3132-41. (2006)

B. Liu and M. Krieger Highly Purified Scavenger Receptor Class B, Type I reconstituted into phosphatidylcholine/ cholesterol liposomes mediates high affinity high density lipoprotein binding and selective lipid uptake. J. Biol. Chem. 277, 34125-34135. (2002)

Ungar D, Oka T, Krieger M, Hughson FM. Retrograde transport on the COG railway. Trends Cell Biol. Feb;16(2):113-20. (2006)

Foulquier F, Vasile E, Schollen E, Callewaert N, Raemaekers T, Quelhas D, Jaeken J, Mills P, Winchester B, Krieger M, Annaert W, Matthijs G. Conserved oligomeric Golgi complex subunit 1 deficiency reveals a previously uncharacterized congenital disorder of glycosylation type II. Proc Natl Acad Sci U S A. Mar 7;103(10):3764-9. (2006)

Yesilaltay A, Morales MG, Amigo L, Zanlungo S, Rigotti A, Karackattu SL, Donahee MH, Kozarsky KF, Krieger M. Effects of hepatic expression of the high-density lipoprotein receptor SR-BI on lipoprotein metabolism and female fertility. Endocrinology. Apr;147(4):1577-88 (2006)

Karackattu SL, Trigatti B, Krieger M. Hepatic lipase deficiency delays atherosclerosis, myocardial infarction, and cardiac dysfunction and extends lifespan in SR-BI/apolipoprotein E double knockout mice. Arterioscler Thromb Vasc Biol. Mar 26(3):548-54. (2006)

S. Zhang, M.H. Picard, E. Vasile, Y. Zhu, R.L. Raffai, K.H. Weisgraber, and M. Krieger Diet-Induced occlusive coronary atherosclerosis, myocardial infarction, cardiac dysfunction and premature death in SR-BI-deficient, hypomorphic apolipoprotein ER61 mice. Circulation 111, 3457-64. (2005)

Oka T, Vasile E, Penman M, Novina CD, Dykxhoorn DM, Ungar D, Hughson FM, Krieger M. Genetic analysis of the subunit organization and function of the conserved oligomeric golgi (COG) complex: studies of COG5- and COG7-deficient mammalian cells. J Biol Chem. Sep 23;280(38):32736-45 (2005)

X. Wu, R. A. Steet, O. Bohorov, J. Bakker, J. Newell, M. Krieger, L. Spaapen, S. Kornfeld and H. H. Freeze. Mutation of the COG complex subunit gene COG7 causes a lethal congenital disorder. Nature Medicine 10, 518-523. (2004)

T. Oka, D. Ungar, F. M. Hughson, and M. Krieger. The COG and COPI complexes interact to control the abundance of GEARs, a subset of Golgi integral membrane proteins. Molecular Biology of the Cell 15, 2423-2435. (2004)

O. Kocher, A. Yesilaltay, C. Cirovic, R. Pal, A. Rigotti, and M. Krieger. Targeted disruption of the PDZK1 gene in mice causes tissue-specific depletion of the HDL Receptor SR-BI and altered lipoprotein metabolism. J. Biol. Chem. 278, 52820-52825. (2003)

Braun, A., Zhang, S., Miettinen, H.E., Ebrahim, S., Holm, T.M., Vasile, E., Post, M.J., Yoerger, D.M., Picard, M.H., Krieger, J.L., Andrews, N.C., Simons, M., and Krieger, M. Probucol prevents early coronary heart disease and death in the high-density lipoprotein receptor SR-BI/apolipoprotein E double knockout mouse. PNAS 100 (12): 7283-7288. (2003)

Rigotti, A., Miettinen, H.E., and Krieger, M. The role of the high-density lipoprotein receptor SR-BI in the lipid metabolism of endocrine and other tissues. Endocrine Reviews 24(3):357-387. (2003)

S. D. Covey, M. Krieger, W. Wang, M. Penman, and B. L. Trigatti Scavenger Receptor Class B Type I-Mediated Protection Against Atherosclerosis in LDL Receptor-Negative Mice Involves Its Expression in Bone Marrow-Derived Cells. Arterioscler Thromb Vasc Biol. 23, 1589-1594. (2003)

Vasile, E., Perez, T., Nakamura, N., and Krieger, M. Structural Integrity of the Golgi is Temperature Sensitive in Conditional-Lethal Mutants with No Detectable GM130. Traffic 4(4): 254-272 (2003)

Ungar, D., Oka, T., Brittle, E. E.,Vasile, E., Lupashin, V.V., Chatterton, J. E., Heuser, J.E., Krieger, M., and Waters, M.G. Characterization of a mammalian Golgi-localized protein complex, COG, that is required for normal Golgi morphology and function. J. Cell Biology 157: 405-415 (2002)

T.J. Nieland, M. Penman, L. Dori, M. Krieger, and T. KirchhausenDiscovery of chemical inhibitors of the selective transfer of lipids mediated by the HDL receptor SR-BI. PNAS 99, 15422-15427. (2002).

B. Liu and M. KriegerHighly Purified Scavenger Receptor Class B, Type I reconstituted into phosphatidylcholine/ cholesterol liposomes mediates high affinity high density lipoprotein binding and selective lipid uptake. J. Biol. Chem. 277, 34125-34135. (2002)

Gu, X., Lawrence, R., and Krieger, M. Dissociation of the high density lipoprotein and low density lipoprotein binding activities of murine scavenger receptor class B type I (mSR-BI) using retrovirus library-based activity dissection. J. Biol. Chem. 275: 9120-9130. (2000)

See PubMed for Krieger Lab publications.