Kiran Madura Associate Professor UMDNJ-Robert Wood Johnson Medical School Department of Biochemistry New Research Building. Room 383 683 Hoes Lane Piscataway. NJ 08855 (732) 235-5602 FAX - 5417 maduraki@umdnj.edu

Ubiquitin-mediated protein degradation in DNA repair and signal transduction

The major research focus in my laboratory is the investigation of protein ubiquitination and degradation by the proteasome. We discovered that Rad23 is a shuttle-factor that can bind ubiquitinated proteins and deliver them to the proteasome. to initiate degradation. The domains in Rad23 that bind ubiquitinated proteins and the proteasome were identified. We are using molecular. biochemical and genetic methods (in both yeast and cell-culture based systems). to understand the mechanism of intracellular proteolysis. and its significance in DNA repair. stress-response and human neurodegenerative diseases.

DNA repair: Current studies are exploring the role of Rad23 in controlling the stability of the DNA repair protein Rad4. Mutations in the human homolog of Rad4 lead to Xeroderma pigmentosum.. a debilitating disease in which patients are extremely sensitive to sunlight. Rad23 is extensively phosphorylated in vivo. The significance of this post-translation modification. and its significance in DNA repair is under investigation.

Stress-response: We used genetic and biochemical methods to establish a link between Rad23 and the proteasome. Loss of both Rad23 and a specific proteasome subunit results in diverse proteolytic and growth defects. Current studies are focused on characterizing a number of dosage suppressors that can alleviate the defects of this double mutant strain. One of these suppressors is Sts1. and our studies suggest that it is a new regulatory factor in the Ub/proteasome proteolytic system.

Neurodegenerative diseases: We recently described the first biochemical function for the ataxin-3 protein. Mutations in this protein cause Machado-Joseph disease. a fatal neurodegenerative condition. Ataxin-3 interacts with the shuttle-factor Rad23. as well as the chaperone VCP/p97. Our studies suggest that ataxin-3 is a novel component of the proteasome that can bind ubiquitinated substrates. In other studies we are characterizing alpha-synuclein and Parkin. which are causative factors in Parkinson's disease.

Proteomics: We developed affinity matrices to rapidly purify cellular ubiquitinated proteins and the intact proteasome. We have begun to identify targets of ubiquitin-mediated proteolysis. and novel components of the proteasome. by high-resolution two-dimensional gel electrophoresis and mass spectrometry. We have also implemented this approach for characterizing heart disease. breast cancer and colorectal cancer.

Selected Publications

Madura K. (2009) Cell biology: The proteasome assembly line. Nature. 459(7248):787-8.

Hedhli N, Lizano P, Hong C, Fritzky LF, Dhar SK, Liu H, Tian Y, Gao S, Madura K, Vatner SF, Depre C. (2008) Proteasome inhibition decreases cardiac remodeling after initiation of pressure overload. Am J Physiol Heart Circ Physiol. 295(4):H1385-93.

Geetha T, Seibenhener ML, Chen L, Madura K, Wooten MW. (2008) p62 serves as a shuttling factor for TrkA interaction with the proteasome. Biochem Biophys Res Commun. Sep 12;374(1):33-7.

Chen L, Madura K. (2008) Centrin/Cdc31 is a novel regulator of protein degradation. Mol Cell Biol. 28(5):1829-40.

Hedhli N, Wang L, Wang Q, Rashed E, Tian Y, Sui X, Madura K, Depre C. (2008) Proteasome activation during cardiac hypertrophy by the chaperone H11 Kinase/Hsp22. Cardiovasc Res. 77(3):497-505.

Romero-Perez L, Chen L, Lambertson D, Madura K. (2007) Sts1 can overcome the loss of Rad23 and Rpn10 and represents a novel regulator of the ubiquitin/proteasome pathway. J Biol Chem. 282(49):35574-82.

Depre C, Wang Q, Yan L, Hedhli N, Peter P, Chen L, Hong C, Hittinger L, Ghaleh B, Sadoshima J, Vatner DE, Vatner SF, Madura K. (2006) Activation of the cardiac proteasome during pressure overload promotes ventricular hypertrophy. Circulation. 114(17):1821-8.

Chen L. Thiruchelvam MJ. Madura K. Richfield EK. (2006) Proteasome dysfunction in aged human alpha-synuclein transgenic mice. Neurobiol Dis. 23(1):120-6.

Chen L. Madura K. (2006) Evidence for distinct functions for human DNA repair factors hHR23A and hHR23B. FEBS Lett. 580(14):3401-8.

Chuang SM. Madura K. (2005) Saccharomyces cerevisiae Ub-conjugating enzyme Ubc4 binds the proteasome in the presence of translationally damaged proteins. Genetics. 171(4):1477-84.

Chen. L. and Madura. K. (2005). Increased proteasome activity. ubiquitin-conjugating enzymes and eEF1A translation factor detected in breast cancer tissue. Cancer Res. 65:13-20.

Chuang. S-M.. Chen. L.. Lambertson. D.. Anand. M.. Kinzy. T. G. and Madura. K. (2005). Proteasome-mediated degradation of co-translationally damaged proteins involves the translation elongation factor eEF1A. Mol. Cell. Biol. Vol. 25: 403-413.

Doss-Pepe. E.. Chen. L. and Madura. K. (2005). Alpha-Synuclein and Parkin contribute to the assembly of ubiquitin lysine63-linked multiubiquitin chains. J Biol Chem.. 280:16619-24.

Ortolan. T. G.. Chen. L.. Tongaonkar. P. and Madura. K. (2004). Rad23 stabilizes Rad4 from degradation by the Ub/proteasome pathway. Nuc. Acids Res. 34: 1-11.

Madura. K. (2004). Rad23 and Rpn10: Perennial wallflowers join the melee. Trends in Biochem. Sci. 29: 637-40.

Johnston. JA and Madura. K. (2004). Rings. Chains and Ladders; Ubiquitin goes to work in the neuron. Progress in Neurobiology 73: 227-257.

Doss-Pepe. E. W.. Stenroos. E. S.. Johnson. W. G. and Madura. K. (2003). Ataxin-3 interactions with Rad23 and Valosin-containing protein and its associations with ubiquitin chains and the proteasome are consistent with a role in ubiquitin-mediated proteolysis. Mol. Cell. Biol. 23: 6469-6483.

Lambertson. D.. Chen. L. and Madura. K. (2003). Investigating the importance of proteasome-interaction for Rad23 function. Current Genetics 42: 199-208.