James E. Metherall, Ph.D.
Associate Professor, Department of Human Genetics

Contribution to Society

Inherited abnormalities of cholesterol metabolism lead to considerable morbidity and mortality. Mutations resulting in increased cholesterol levels contribute to Coronary Heart Disease which accounts for more than 500,000 deaths per year in the United States. In addition, mutations that drastically lower cholesterol levels lead to mental retardation; more than 1 in every 32 Americans is heterozygous for one of these mutations. We study these and other mutations so that we can better understand the normal processes and identify new ways of diagnosing and treating these very common disorders.

Research Summary

Our lab studies the cellular and molecular processes that control cholesterol metabolism. We use a wide range of approaches including; biochemical and metabolic assays, human genetics, somatic cell genetics, and mammalian expression cloning. Our work is divided into three separate areas: 1) the study of genes involved in cellular cholesterol homeostasis and transport, 2) the identification and characterization of mutations affecting cholesterol biosynthesis and their contributions to inherited forms of mental retardation, and 3) the development and use of robotics to automate large scale biochemical and genetic screens in a wide range of functional genomics screens.

Cholesterol is an essential component of mammalian cell membranes. Cells maintain exquisite control over the level of free cholesterol within the cell by a classic end-product feedback mechanism. Elevated sterol levels suppress transcription of the genes for the first two enzymes of the cholesterol biosynthetic pathway: HMG CoA synthase and HMG CoA reductase. In addition, sterols suppress transcription of the gene for the LDL receptor, a cell surface receptor that mediates the uptake of cholesterol-rich lipoprotein particles. A DNA binding protein (SREBP) interacts with the promoter of each of these genes to regulate transcription. The DNA binding protein is originally synthesized as a transmembrane precursor anchored to the endoplasmic reticulum (ER). In the absence of sterols, the precursor is protealized by site 1 protease (S1P) which removes one of the transmembrane domains and the ER-retention signal. The protein is then free to flow to the Golgi where a second protease, S2P, removes the second transmembrane domain. The released protein then enters the nucleus and promotes transcription. Sterol cleavage activating protein (SCAP) is an essential cofactor for S1P. SCAP binds cholesterol and when cholesterol is bound it no longer functions as a cofactor. Consequently when cholesterol levels rise, SCAP is inactivated, SREBP is retained in the ER and the transcription of sterol-regulated genes decreases.

We have isolated mutant Chinese Hamster Ovary (CHO) cell lines that have defects in these processes. The study of these mutants helped define the pathway and complementation of the mutants was used to isolate the genes for S2P, S1P and SCAP. SRD -2 cells harbor a gene rearrangement in SREBP that truncates the protein, thereby eliminating the transmembrane domains. The truncated protein is free to enter the nucleus and activate transcription even in the presence of sterols. We have also isolated a mutant cell line (SRD-6) that lacks S2P protease. In SRD-6 cells, the DNA binding protein remains trapped in the Golgi and therefore, fails to activate transcription even in the absence of sterols. Similarly, another cell line lack S1P activity and SREBP is retained in the ER as a full length protein. In addition, we have isolated a mutant cell line (SRD-7) that fails to deliver cholesterol to the ER. While cholesterol is found predominantly in the plasma membrane, the triggering event for proteolysis and many other important processes involved in cholesterol metabolism occur in the ER. SRD-4 cells have a dominant mutation in SCAP which prevents it from acting as a cofactor for S1P. Consequently, SRD-4 cells fail to activate transcription through SREBP. SRD-4 cells also have a mutation in acyl CoA:cholesterol acyltransferase (ACAT), an important enzyme involved in coronary heart disease. SRD-7 cells accumulate cholesterol-rich vesicles in the cytoplasm suggesting that vesicular trafficking normally delivers cholesterol to the ER. We have demonstrated that this transport process also requires the activity of a known protein, one or more members of the multiple-drug resistance (MDR) family of ABC cassette transporters.

Recent Publications

Scuderi, A., Simin, K., Kazuko, S. G., Metherall, J.E., Letsou, A. (2006) scylla and charybde Homologues of the Human Apototic Gene RTP801, are Required for Head Involution in Drosophila. Dev. Biol. 291, 110-22.

Metherall, J. E., Nash, E. and Warnick, D. C.(2005) Cholesterol Metabolism Regulation. Nature Encyclopedia of Life Sciences, John Wiley & Sons, Ltd, London.

Hoshijima, K., Metherall, J.E., Grunwald, D.J. (2002) A Protein Disulfide Isomerase Expressed in the Embryonic Midline is Required for Left/Right Asymmetries. Genes and Development 16, 2518-2529.

Simin, K., Scuderi, A., Reamey, J., Dunn, D., Weiss, R., Metherall, J.E., Letsou, A. (2002) Profiling Patterned Transcripts in Drosophila Embryos. Genome Research 12, 1040-1047. (see commentary: Oliver, B. (2002) Fly Factory. Genome Research 12, 1017-1018.)

Gao, Z.-H., Metherall, J.E., Virshup, D. M. (1999) Library Screening to Identify Casein Kinase I Substrates. Identification of Casein Kinase I Substrates by in vitro Expression Cloning (IVEC) Screening. Biochem. Biophys. Res. Com. 268, 562-566.

Elsea, S. H., Mykytyn, K., Ferrell, K., Das, P., Dubiel, W., Patel, P. I., Metherall, J. E. (1999) The COP9 Signalosome Subunit Gene, SGN3, Maps within the Smith-Magenis Syndrome Critical Interval. Am. J. Med. Gen. 87, 342-348.

Neklason, D. W., Kelley, R., Metherall, J. E. (1999) Biochemical Variants of 7-Dehydrocholesterol Reductase Deficiency. Am. J. Med. Genetics 85, 517-523.

Metherall, J. E. and Nash, E. (1999) Cholesterol Metabolism Regulation. Encyclopedia of Life Sciences, Nature Publishing Group, London.

DeBry, P., Nash, E. A., Neklason, D., Metherall, J. E. (1997) Role of Multidrug Resistance (MDR) P-Glycoproteins in Cholesterol Esterification. J. Biol. Chem. 272, 1026-1031.

Jackson, S. M., Ericsson, J., Goto, A., Metherall, J. E., and Edwards, P. A. (1996) Sterol Regulatory Element Binding Protein is Involved in the Regulation of Farnesyl Diphosphate Synthase and Cholesterol and Fatty Acid Synthesis: Evidence from Sterol Regulation-Defective Cell Lines. J. Lipid Research 37, 1712-1721.

Metherall, J. E., Waugh, K., Li, H. (1996) Progesterone Inhibits Cholesterol Biosynthesis: Accumulation of Sterol Precursors J. Biol. Chem. 271, 2627-2633.

Metherall, J. E., Li, H., Waugh, K. (1996) Role of Multidrug Resistance (MDR) P-Glycoproteins in Cholesterol Biosynthesis. J. Biol. Chem. 271, 2634-2640.

Evans, M. J., Metherall, J. E. (1993) Loss of Transcriptional Repression of Three Sterol‑regulated Genes in Mutant Hamster Cell Lines. Mol. Cell. Biol. 13, 5175-5185.

Naglich, J., Metherall, J. E., Russell, D. W., Eidels, L. (1992) Expression Cloning of a Diphtheria Toxin Receptor : Identity with a Heparin-Binding EGF-Like Growth Factor Precursor. Cell 69, 1051-1061.

Metherall, J. E., Ridgway, N. D., Dawson, P. A., Goldstein, J. L., Brown, M. S. (1991) A 25-Hydroxycholesterol Resistant Cell Line Deficient in Acyl CoA:Cholesterol Acyltransferase J. Biol. Chem. 266, 12734-12740.

Dawson, P. A., Metherall, J. E., Ridgway, N. D., Brown, M. S., Goldstein, J. L. (1991) Separation of Transcriptional and Post‑transcriptional Regulation of 3‑hydroxy‑3‑methylglutaryl Coenzyme A Reductase in Mutant Hamster Cells. J. Biol. Chem. 266, 9128-9134.

Metherall, J. E., Goldstein, J. L., Luskey, K. L., Brown, M. S. (1989) Loss of Transcriptional Repression of Three Sterol‑regulated Genes in Mutant Hamster Cell Lines. J. Biol. Chem. 264, 15634‑15641.

Metherall, J. E., Gillespie, F. P. and Forget, B. G. (1989) Nuclear Proteins of a Human Erythroleukemic Cell Line That Bind to the Promoter Region of Normal and Nondeletion HPFH ‑Globin Genes. Prog. in Clin. & Biol. Res. 316A, 247-260.

Glazer, P. M., Greggio, N. A., Metherall, J. E., Summers, W. C. (1989) UV‑induced DNA‑binding Proteins in Human Cells. Proc. Natl. Acad. Sci. 86, 1163‑1167.

Metherall, J. E., Gillespie, F. P. and Forget, B.G. (1988) Analysis of Linked - Globin Genes Suggests that Non‑deletion forms of Hereditary Persistence of Fetal Hemoglobin are bona fide Switching Mutants. Am. J. of Human Genetics 42, 476‑481.

Metherall, J. E., Stoeckert, C. J., Gillespie, F. P., Weissman, S. M. and Forget, B.G. (1987) Support for the Causal Relationship Between ‑globin Gene Promoter Mutations and Non-deletion Forms of Hereditary Persistence of Fetal Hemoglobin. Prog. Clin. Biol. Res. 251, 347‑361.

Hoshijima, K., Metherall, J.E., Grunwald, D.J. (2002) A Protein Disulfide Isomerase Expressed in the Embryonic Midline is Required for Left/Right Asymmetries. Genes and Development 16, 2518-2529.

Simin, K., Scuderi, A., Reamey, J., Dunn, D., Weiss, R., Metherall, J.E., Letsou, A. (2002) Profiling Patterned Transcripts in Drosophila Embryos. Genome Research 12, 1040-1047. (see commentary: Oliver, B. (2002) Fly Factory. Genome Research 12, 1017-1018.)

Gao, Z.-H., Metherall, J.E., Virshup, D. M. (1999) Library Screening to Identify Casein Kinase I Substrates. Identification of Casein Kinase I Substrates by in vitro Expression Cloning (IVEC) Screening. Biochem. Biophys. Res. Com. 268, 562-566.

Elsea, S. H., Mykytyn, K., Ferrell, K., Das, P., Dubiel, W., Patel, P. I., Metherall, J. E. (1999) The COP9 Signalosome Subunit Gene, SGN3, Maps within the Smith-Magenis Syndrome Critical Interval. Am. J. Med. Gen. 87, 342-348.