Cathy Clarke

Catherine F. Clarke
Department of Chemistry and Biochemistry, UCLA

Professor of Biochemistry

B.S. and Ph.D., University of California, Los Angeles

Phone: (310) 825-0771
Fax: (310) 206-5213
E-mail:cathy@chem.ucla.edu

UCLA Department of Chemistry & Biochemistry
5072B Young Hall
Box 951569 (post)
607 Charles E. Young Drive East (courier)
Los Angeles, CA 90095-1569

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   Research Summary

Introduction – Ubiquinone (coenzyme Q or Q) functions in cells as a redox-active coenzyme of mitochondrial and plasma membrane electron transport, as well as an essential lipid soluble antioxidant. Human dietary supplementation with Q appears to have beneficial effects in slowing the progression of neuro- and muscle-degenerative diseases. However, Q is also involved in generating reactive oxygen species, through the adventitious reduction of dioxygen to superoxide by the ubisemiquinone radical, normally generated during mitochondrial electron transport. Thus, it is not clear how dietary supplementation with Q impacts the Dr. Jekyll/Mr. Hyde aspects of Q function. Cells are capable of synthesizing Q, but much remains to be learned about the sites of its synthesis, mechanisms of inter- and intra-cellular transport, and the regulation and enzymology of its biosynthesis. Research in my laboratory identified eight of the eleven polypeptides required for Q biosynthesis. The goals of my research are to characterize the Coq polypeptides responsible for production of Q and to determine how their activity can be modulated for optimal health.

Research Overview -

(1) We discovered a new Q biosynthetic pathway and showed that yeast are able to synthesize Q from a new aromatic ring precursor, para-aminobenzoic acid (pABA) [77]. This pathway operates in addition to the classic Q biosynthetic pathway that emanates from 4-hydroxybenzoic acid (4HB). We suggest a mechanism where Schiff base mediated deimination forms DMQ6 quinone, thereby eliminating the nitrogen contributed by pABA. This scheme results in the convergence of the 4HB and pABA pathways in eukaryotic Q biosynthesis and has implications regarding the action of pABA-based antifolates. This work is funded by a grant from NSF.

(2) The findings we made in the yeast model have shed much light on diseases resulting from Q deficiency [82, 85]. We showed that human Coq8 and Coq6 polypeptide homologues function to rescue the respective yeast coq mutants [80, 81]. We discovered that yeast Coq8p, and its human homologue ADCK3 function in the phosphorylation of Coq3p, Coq5p, and Coq7p [80]. The kinase function of Coq8p is proposed to be important in the maintenance or formation of a high molecular mass Coq polypeptide complex, which is required for Q biosynthesis.

(3) Over-expression of Coq8p in many of the yeast coq null mutants restores steady state levels of Coq polypeptides. This has been a particularly important discovery, because the yeast coq null mutants (coq3-coq9) previously accumulated the same early Q-intermediate. Coq8 over-expression has enabled us to detect late-stage Q-intermediates that were previously elusive [86]. Finally, we used synthetic analogues of 4-hydroxybenzoic acid to bypass deficient biosynthetic steps and showed that 2,4-dihydroxybenzoic acid is able to restore Q6 biosynthesis and respiratory growth in a coq7 null mutant over-expressing Coq8. The over-expression of Coq8 and the use of analogues of 4-HB represent new tools to elucidate the Q biosynthetic pathway. These recent discoveries are depicted in Figure 1.

Figure 1

Figure 1. S. cerevisiae Q6 biosynthetic pathway: accumulation of Q6 biosynthetic intermediates caused by the over-expression of Coq8 in Dcoq strains. The classic Q biosynthetic pathway is shown in path 1 emanating from 4-hydroxybenzoic acid (4-HB). Coq1 (not shown) synthesizes the hexaprenyl-diphosphate tail which is transferred by Coq2 to 4-HB to form 3-hexaprenyl-4-hydroxybenzoic acid (HHB).  R represents the hexaprenyl tail present in all intermediates from HHB to Q6. Alternatively, path 2 shows para-aminobenzoic acid (pABA) is prenylated by Coq2 to form 3-hexaprenyl-4-aminobenzoic acid (HAB). Both HHB and HAB are early Q-intermediates, readily detected in each of the coq null strains (Dcoq3-Dcoq9). The numbering of the aromatic carbon atoms used throughout this study is shown on the reduced form of Q6, Q6H2. Coq8 over-expression in certain Dcoq strains leads to the accumulation of the following compounds: 4-AP, 3-hexaprenyl-4-aminophenol; 4-HP, 3-hexaprenyl-4-hydroxyphenol; HHAB, 3-hexaprenyl-4-amino-5-hydroxybenzoic acid; HMAB, 3-hexaprenyl-4-amino-5-methoxybenzoic acid; DDMQ6, the oxidized form of demethyl-demethoxy-Q6H2; IDMQ6, 4-imino-demethoxy-Q6; DMQ6H2, demethoxy-Q6H2. IDDMQ6H2, 2-demethyl-4-amino-demethoxy-Q6H2 and DHHB, 3-hexaprenyl-4,5-dihydroxybenzoic acid are shown but have not been detected in this study. Black dotted arrows (from path 2 to path 1) designate the replacement of the C4-amine with a C4-hydroxyl and correspond to the C4-deamination reaction. A putative mechanism to replace the C4-imino group with the C4-hydroxy group is shown in bracket in blue on IDMQ6 but could also occur on IDDMQ6 (not shown). 4-AP and 4-HP which are formed upon inhibition of the C5-hydroxylation catalyzed by Coq6 are shown in red. Analogues of 4-HB and pABA allowing the bypass of certain steps in Q biosynthesis are indicated in green. Steps defective in the Dcoq9 strain are designated with a red asterisk.

4) In collaboration with Retrotope, we have employed novel isotope-reinforced polyunsaturated fatty acids (PUFAs) that reveal the importance of Q as an essential antioxidant [79, 87]. Polyunsaturated fatty acids (PUFA) undergo autoxidation and generate reactive carbonyl compounds that are toxic to cells and associated with apoptotic cell death, age-related neurodegenerative diseases, and atherosclerosis. PUFA autoxidation is initiated by the abstraction of bis-allylic hydrogen atoms. Replacement of the bis-allylic hydrogen atoms with deuterium atoms (termed site-specific isotope-reinforcement) arrests PUFA autoxidation due to the isotope effect. We investigate the effects of different isotope-reinforced PUFAs and natural PUFAs on autoxidation kinetics, and on the viability of coenzyme Q-deficient Saccharomyces cerevisiae coq mutants and wild-type yeast subjected to copper stress. Cells treated with a C11-BODIPY fluorescent probe to monitor lipid oxidation products show that lipid peroxidation precedes the loss of viability due to H-PUFA toxicity. We show that replacement of just one bis-allylic hydrogen atom with deuterium is sufficient to arrest lipid autoxidation. In contrast, PUFA reinforced with two deuterium atoms at mono-allylic sites remain susceptible to autoxidation. Surprisingly, yeast treated with a mixture of approximately 20%:80% isotope-reinforced D-PUFA: natural H-PUFA are profoundly protected from lipid autoxidation-mediated cell killing. Our findings show that inclusion of only a small fraction of PUFA deuterated at the bis-allylic sites is sufficient to profoundly inhibit the chain reaction of non-deuterated PUFA. .

Figure 2

Fig. 2. Replacement of bis-allylic H atoms with deuterium (D) arrests autoxidation of PUFA. A theoretical chain reaction is depicted where a single initiation event producing a lipid peroxyl radical (denoted by –OO•) starts a chain reaction of lipid autoxidation that in the presence of O2, may continue indefinitely and produce many molecules of lipid peroxides; susceptible phospholipid molecules containing a PUFA acyl chain are designated by a red dot. The presence of 20% isotope-reinforced PUFA (denoted by a black dot) inhibit (or slow) chain propagation. Propagation is inhibited for PUFA neighboring the D-PUFA.

 

 Representative Publications

From my Group 1996-2000
From my Group 2001-2005

From my Group 2006-2009

From my Group 2010-Present

76. Brajcich, B. C., Iarocci, A. L., Johnstone, L. A. G., Morgan, R. K., Lonjers, Z. T., Dickson, E. L., Hotchko, M. J., Muhs, J. D., Kieffer, A., Reynolds, B. J., Mandel, S. M., Marbois, B. N., Clarke, C. F., and Shepherd, J. N. (2010) "Evidence that ubiquinone is a required intermediate for rhodoquinone biosynthesis in Rhodospirillum rubrum." J. Bacteriol. 192, 436-445. PMCID: PMC2805321.

77. Marbois, B., Xie, L. X., Choi, S., Hirano, K., Hyman, K., and Clarke, C. F. (2010) “Para-aminobenzoic acid is a precursor in coenzyme Q6 biosynthesis in Saccharomyces cerevisiae.” J. Biol. Chem. 285, 27827-27838. PMCID: PMC2934650.

78. Devarajan, A., Bourquard, N., Hama, S., Navab, M., Grijalva, V.R., Morvaridi, S., Clarke, C. F., Vergnes, L., Reue, K., Tieber, J. F., and Reddy, S. T. (2010) “Paraoxanase 2 deficiency alters mitochondrial function and exacerbates the development of atherosclerosis.” Antioxid Redox Signal 14, 341-351. PMCID: PMC3011913.

79. Hill, S., Hirano, K., Shmanai, V. V., Marbois, B. N., Vidovic, D., Bekish, A. V., Kay, B., Tse, V., Fine, J., Clarke, C. F., and Shchepinov, M. S., (2011) “Isotope-reinforced polyunsaturated fatty acids strongly protect yeast cells from oxidative stress.” Free Radical Biol. Med. 50, 130-138. PMCID: PMC3014413.

80. Xie, L. X., Hsieh, E. J., Watanabe, S., Allan, C. M., Chen, J. Y., Tran, U. C., and Clarke, C. F. (2011) “Expression of the human atypical kinase ADCK3 rescues coenzyme Q biosynthesis and phosphorylation of Coq polypeptides in yeast coq8 mutants.” Biochim. Biophys. Acta (2011) 1811, 348-360. PMCID: PMC3075350.

81. Heeringa, S. F., Chernin, G., Chaki, M., Zhou, W., Sloan, A. J., Ji, Z., Xie, L. X., Salviati, L.,  Hurd, T. W., Vega-Warner, V., Killen, P. D., Raphael Y., Ashraf, S., Ovunc, B., Schoeb, D. S., McLaughlin, H. M., Airik, R., Vlangos, C. N., Gbadegesin, R., Hinkes, B., Saisawat, P., Trevisson, E., Doimo, M., Casarin, A., Pertegato, V., Giorgi, G., Prokisch, H., Rötig, A.,  Antignac, C., Nurnberg, G., Becker, C., Wang, S.,  Ozaltin, F., Topaloglu, R., Bakkaloglu, A., Bakkaloglu, S. A., Müller, D., Beissert, A., Mir, S., Berdeli, A., Özen, S.,  Zenker, M., Verena Matejas, V., Ocana, C. S., Navas, P., Kusakabe, T., Kispert, A., Akman, S., Soliman, N. A., Krick, S., Mundel, P., Reiser J., Peter Nurnberg, P., Clarke, C. F., Wiggins, R. C., Faul, C., and Hildebrandt, F. (2011) “COQ6 mutations in human patients produce nephrotic syndrome with sensorineural deafness.” J. Clin. Invest. (2011) 121, 2013-2024. PMCID: PMC3083770.


82. Falk, M. J., Polyak, E., Zhang, Z., Peng, M., King, R., Maltzman, J. S., Okwuego, E., Horyn, O., Nakamaru-Ogiso, E., Ostrovsky, J., Xie, L. X., Chen, J. Y., Marbois, B., Nissim, I., Clarke, C. F., Gasser, D. L. (2011) “Probucol ameliorates renal and metabolic sequelae of primary CoQ deficiency in Pdss2 mutant mice.” EMBO Mol. Med. 7, 410-427. PMCID: PMC3394513.


83. Pfeiffer, M., Kayser, E.B., Yang, X., Abramson, E., Kenaston, M. A., Lago, C. U., Lo, H.H., Sedensky, M. M., Lunceford, A., Clarke, C. F., Wu, S. J., McLeod, C., Finkel, T., Morgan, P., Mills, E. M. (2011) “Caenorhabditis elegans UCP4 controls complex II-mediated oxidative phosphorylation through succinate transport.” J. Biol. Chem. 286, 37712-37720. PMCID:PMC3199514.

84. Clarke, C. F. (2011) “Coq6 hydroxylase: Unmasked and bypassed.” Chem. Biol. 18, 1069-1070.
PMCID:PMC3245979. INVITED COMMENTARY

85. Rahman, S., Clarke, C. F., and Hirano, M. (2011) “176th ENMC International Workshop: Diagnosis and treatment of coenzyme Q10 deficiency”. Neuromuscular Disorders. 22, 76-86. PMCID: PMC3222743. REVIEW ARTICLE

86. Xie, L. X., Ozeir, M., Tang, J. Y., Chen, J. Y., Jaquinod, S-K., Fontecave, M, Clarke, C. F., and Pierrel, F. (2012) "Over-expression of the Coq8 kinase in Saccharomyces cerevisiae coq null mutants allows for accumulation of diagnostic intermediates of the coenzyme Q6 biosynthetic pathway." J. Biol. Chem. 287, 23571-23581. PMCID: PMC3390632.

87. Hill, S., Lamberson, C. R., Libin X., To, R., Tsui, H. S., Shmanai, V. V., Bekish, A. V., Awad, A. M., Marbois, B. N., Cantor, C. R., Porter, N. A., Clarke, C. F., and Shchepinov, M. S. (2012) "Small amounts of isotope-reinforced polyunsaturated fatty acids suppress lipid autoxidation." Free Rad. Biol. Med. 53, 893-906. PMCID: PMC3437768.

88. Gomez, F., Saiki, R., Chin, R., Srinivasan, C., and Clarke, C. F. (2012) "Restoring de novo coenzyme Q biosynthesis in Caenorhabditis elegans coq-3 mutants yields profound rescue compared to exogenous coenzyme Q supplementation". Gene 506, 106-116. PMCID: PMC3437764.

89. Nguyen, T. P. T., Clarke, C. F. (2012) "Folate status of gut microbiome affects Caenorhabditis elegans lifespan". BMC Biology 10, 66 PMCID: PMC3409036.

90. Gomez, F., Monsalve, G. C., Tse, V., Saiki, R., Weng, E., Lee, L., Srinivasan, C., Frand, A. R., and Clarke, C. F. (2012) "Delayed accumulation of intestinal coliform bacteria enhance life span and stress resistance in Caenorhabditis elegans fed respiratory deficient E. coli." BMC Microbiol. 12, 300. PMCID: PMC3548685.

91. Allan, C. M., Hill, S., Morvaridi, S., Saiki, R., Johnson, J. S., Liau, W-S., Hirano, K., Kawashima, T., Ziming, J., Loo, J. A., Shepherd, J. N., and Clarke, C. F. (2013) "A conserved START domain coenzyme Q-binding polypeptide is required for efficient Q biosynthesis, respiratory electron transport, and antioxidant function in Saccharomyces cerevisiae". Biochim. Biophys. Acta 1831, 776-791. PMCID: PMC3909687.

92. Gasser, D. L., Winkler, C. A., Peng M., Ping, A., McKenzie, L. M., Kirk, G. D., Shi, Y., Xie, L. X., Marbois, B. N., Clarke, C. F., and Kopp, J. B. (2013) "Focal Segmental Glomerulosclerosis is associated with a PDSS2 haplotype and independently, with a decreased content of coenzyme Q10." Amer. J. Physiol. Renal Physiol. Am J Physiol Renal Physiol. Oct; 305(8):F1228-38. PMCID: PMC3798722.

93. Ashraf, S., Gee, H. Y., Woerner, S., Xie, L. X., Vega-Warner, V., Svjetlana, L., Fang, H., Song, X., Cattran, D. C., Avila-Casado, C., Paterson, A. D., Nitschke, P., Bole-Feysot, C., Cochat, P., Esteve-Rudd, J., Haberberger, B., Allen, S. J., Zhou, W., Airik, R., Otto, E. A., Barua, M., Al-Hamed, M. H., Kari, J. A., Bockenhauer, D., Kleta, R., El Desoky, S., Hacihamdioglu, D. O., Gok, F., Washburn, J., Wiggins, R. C., Choi, M., Lifton, R. P., Levy, S., Han, Z., Salviati, L., Prokisch, H., Williams, D. S., Pollak, M., Clarke, C. F., Pei, Y., Antignac, C., and Hildebrandt, F. (2013) "ADCK4 mutations promote steroid-resistant nephrotic syndrome through CoQ10 biosynthesis disruption." J. Clin. Invest. 123, 5179-5189. PMCID: PMC3859425.

94. Lamberson, C. R., Xu, L., Muchalski, H., Montenegro-Burke, J. R., Shmanai, V. V., Bekish, A. V., McLean, J. A., Clarke, C. F., Shchepinov, M. S., and Porter, N. A. (2014) "Unusual kinetic isotope effects of deuterium reinforced polyunsaturated fatty acids in tocopherol mediated free radical chain oxidations." J. Amer. Chem. Soc. 136, 838-841. PMCID: In Process.

95. He, C.H., Xie, L. X., Allan, C. M., Tran, U. C., and Clarke, C. F. (2014) "Coenzyme Q supplementation or over-expression of the yeast Coq8 kinase stabilizes multi-subunit Coq polypeptides complexes in yeast coq null mutants." Biochim Biophys Acta 1841, 630-644.

 

 Current Members of Clarke Lab
Visiting Faculty
Srinivasan, Chandra

Dr. Chandra Srinivasan

Associate Professor of Biochemistry
Department of Chemistry & Biochemistry
California State University, Fullerton

Ph.D. Students

Chris Allan

B.S., University of California, Los Angeles

Cuiwen He

B.S., University of California, Los Angeles

Theresa Nguyen

B.S., Loyola Marymount University, Los Angeles

Hui Su Tsui

Hui (Sue) Tsui

B.S., University of California, Los Angeles

Letian Xie

B.S., University of California, Los Angeles

Co-Mentored Ph.D. Students
Barry Lee

Barry Lee

B.S., Johns Hopkins

Primary Mentor: Simon Beaven, Division of Digestive Diseases & Pfleger Liver Institute, UCLA School of Medicine

Rotation Research Students
Kelly Quinn

Kelly Quinn

B.A., UC Berkeley

Undergraduate Students
Dylan Black
Dylan Black

UCLA Biophysics
Alice Hsu

Alice Hsu

UCLA Biochemistry

Nikki Lam

HeiTong (Nikki) Lam

UCLA Biochemistry

Yvonne Nong

Yvonne Nong

UCLA Biochemistry

Minhhan Pham
Minhhan Pham

UCLA Biochemistry
Emily Weng
Emily Weng

UCLA Mol. Cell. Dev. Biol.
 
Former Lab Members
Updates? We'd like to know! E-mail cathy@chem.ucla.edu
Researchers
Dr. Grigory Belogrudov Department of Physiology, UCLA School of Medicine
Dr. Deborah Berthold Research Associate, University of Illinois Champagne-Urbana
Dr. Jenna Hutton Research Associate, University of Rutgers
Dr. Pam Larsen University of Texas Health Sciences Center at San Antonio, Dept. of Cellular & Structural Biology
Dr. Beth Marbois Research Scientist, Department of Biology, U North Carolina, Chapel Hill
Postdoctoral Fellows
Dr. Susan Morvaridi B.S. & Ph.D., Lund University, Research Scientist, UCLA School of Medicine
Dr. Ryoichi Saiki B.A., M.S., & Ph.D. University of Shimane, Research Scientist, Funakoshi, Tokyo, Japan
Ph.D. Students
Dr. Thai Do
B.S., Ph.D.,UCLA - Project Scientist, Molecular Express, Rancho Dominguez, CA
Dr. Peter Gin B.S., Ph.D. 2005, UCLA - Project Scientist, Baxter, Los Angeles, CA
Dr. Fernando Gomez B.S., Ph.D. UCLA 2012 - Post-doctoral Researcher, University of Arlington, Virginia
Dr. Melissa Gulmezian B.A., UC Irvine; Ph.D. - UCLA 2006; Scientist, Allergan, Irvine, CA
Dr. Edward Hsieh Ph.D. 2006, UCLA - Research Scientist, Merck Laboratories, San Francisco
Dr. Tanya Kruse B.S., Ph.D., UCLA - Project Scientist, Baxter, Los Angeles, CA

Dr. Adam Lunceford

B.S., Brigham Young University; Ph.D., UCLA 2008 - J.D., UC Davis, Patent Attorney, Cooley LLP, Washington, D.C.
Dr. Wayne Poon
B.S., Ph.D. UCLA - Research Faculty, UC Irvine
Dr. UyenPhuong C. Tran B.S., UCLA, Ph.D., UCLA 2007 - Research Scientist, Department of Biological Chemistry, UC Irvine
Master Students
Suzie Ward Baba B.S.
Diana Davis B.S., North Carolina State University; M.S., UCLA
Edouard Debonneuil M.S., Computational Biology, UCLA
Shota Watanabe B.S., M.S., UCLA, currently at the University of Hawaii Medical School
Undergraduate Students
Agape Awad B.S., Biochemistry, UCLA, currently Biochemistry & Molecular Biology Ph.D. student at UCLA
Jia Yan Chen Class of 2011 - UCLA Biochemistry
Samuel Choi Class of 2010 - UCLA Biochemistry
Peter Dang Class of 2006 - UCLA Biochemistry

Jason Dinoso

Class of 2001 - UCLA Biochemistry, currently a postdoc at the University of Washington, Seattle
Shauna Hill B.S., Biochemistry, UCLA, currently a graduate student at The University of Texas Health Science Center at San Antonio
Kathleen Hirano Class of 2009 - UCLA Biochemistry, currently a graduate student at UC Berkeley
Kyle Hyman Class of 2008 - UCLA Biochemistry
Ziming Ji Class of 2010 - BS Biophysics, UCLA
Jarrett Johnson B.S., Biochemistry, UCLA, currently a graduate student in Chemical Biology at U Michigan
Bradley Kay Class of 2011 - UCLA Biochemistry
Laura Lee Class of 2010 - California State University, Fullerton

Nico Lee

B.S., M.S., UCLA Biochemistry

Mark Lui

Class of 2012 - UCLA Biochemistry
Amjad Nazzal Current Biochemistry major, UCLA
Tin Mai Class of 2009 - UCLA Biochemistry

Sabine Paterson

Class of 2001 - UCLA Biochemistry
An Pham Class of 2010 - UCLA Biochemistry
Quynh Pham Class of 2013 - UCLA Biochemistry
Thuy Anh Pham Class of 2013 - UCLA Biochemistry
Tomer Schwartz Class of 2012 - UCLA Biochemistry
Yuchen Shi Class of 2008 - UCLA Biochemistry
Jennifer Tang Class of 2012 - UCLA Biochemistry
Aye Win Tin Class of 2011 - UCLA Biochemistry
Randy To Class of 2012 - UCLA Biochemistry
Vincent Tse Class of 2012 - UCLA Mol. Cell. Dev. Biol.
Ruiwen Wang Class of 2010 - UCLA Biochemistry
Summer Interns
Jonathan Fine Pali High 2011
Eva Morozko Class of 2012 - Seton Hall University, B.S. in Biochemistry
Jeffery Roth Class of 2010 - Beverly Hills High



     

Updated 03/05/14