Cell Biology, Neurobiology & Anatomy

Joseph C. Besharse, PhD Marvin Wagner Professor and Chair   Department of Cell Biology, Neurobiology & Anatomy Medical College of Wisconsin 8701 Watertown Plank Road Milwaukee, WI 53226-0509 Phone: (414) 456-8261 FAX: (414) 456-6517 email: jbeshars@mcw.edu

Education:
PhD, Southern Illinois University, 1973
Postdoctoral, Columbia University College of Physicians and Surgeons

Graduate Programs:
Program in Cell and Developmental Biology
Program in Neuroscience

Positions Available:
Positions are currently available for PhD students and Postdoctoral Fellows. Please contact Dr. Joseph Besharse at jbeshars@mcw.edu or visit our Postdoctoral Positions web page.

Research Area: Cellular and molecular basis of circadian rhythmicity in peripheral oscillators; molecular/cellular basis of trafficking of the transduction machinery in retinal photoreceptors

Circadian Clocks in the Eye and Liver: We are studying the cellular and molecular basis of circadian rhythmicity in the retina and other peripheral oscillators such as liver. Many features of photoreceptor and liver metabolism occur in a rhythmic pattern, and a central feature of this regulation is a "circadian clock" that provides endogenous timing signals independent of external cues. We have shown that retinal photoreceptors are the site of a circadian clock and others have shown that the liver hepatocytes are also clocks. Current work is derived from genetic models such as Drosophila in which rhythmically expressed "central clock genes" interact in coupled feedback loops to generate self-sustained circadian oscillations of clock gene expression. This central "clockwork", in turn, controls the rhythmic expression of downstream "clock-controlled" genes that are critical for circadian rhythms of cell function (see Figure 1).

The current major focus of the laboratory is to use mouse genetics to study the role of the central "clockwork" genes, Period and Clock in the retina and the role of the clock regulated genes Nocturnin and Usp2 downstream of the circadian clock. Because circadian organization is widespread in the retina and controls fundamental pathways such as turnover of the phototransduction machinery, it has long been assumed that disruption of circadian clock organization would have a major impact on retinal function. We are capitalizing on targeted mutations in mice of "central clockwork" genes to directly test this assumption. We have also developed targeted mutations of the "clock-regulated genes", nocturnin (an mRNA deadenylase) and Usp2 (a ubiquitin specific protease). We are studying the role of period genes (Per1, Per2, and Per3), Clock, Nocturnin, and Usp2 in rhythmic retinal function and testing the hypothesis that disruption of these genes causes functional deficits that can impair vision and lead to retinal degeneration.

Trafficking of Phototransduction Components in Photoreceptors: Turnover of photosensitive membrane throughout the life of a photoreceptor depends on maintenance of a delicate balance between photosensitive membrane assembly and degradation. These events are controlled by circadian clocks (see above). Both protein and lipid components are synthesized in the cell body and transported vectorially to the region of the sensory cilium where phototransduction organelle is assembled. The cilium is important in the transport of macromolecules from sites of synthesis in the cell body to the region of membrane assembly and in the morphogenesis of flattened discs. Our current work is directed at a model called intraflagellar transport (IFT) in which microtubule based motors (dynein and kinesin) move protein complexes along microtubule tracks into and out of the cilium (see Figure 2). The IFT model is thought to apply to all motile and sensory cilium structures among the eukaryotes; IFT proteins are required for cilium assembly and are found widely in both motile and sensory cilia. Recent work in mice carrying mutations in genes encoding the kinesin II motor and the IFT complex protein, IFT88, demonstrate that this pathway is required for assembly of the phototransduction system in photoreceptors (see Figure 3). Research in this laboratory is directed at organization of the IFT protein complex and its physical interaction with both cell specific cargo and with microtubule based motor proteins.

Figure 1. Diagram illustrating the relationship of the central clockwork (left) and clock regulated genes (right). Only the Per/Cry loop of the clock is illustrated. Clock regulated genes are downstream of the clock and involved in circadian function within cells.

Figure 2. The components of intraflagellar transport as revealed in the green alga, Chlamydomonas.

Figure 3. A model for intrafalgellar transport in photoreceptors based on work of Pazour, et al., 2002.

Selected Publications:

Insinna, C., Pathak, N., Perkins, B., Drummond, I., and J. C. Besharse.  The Homodimeric Kinesin, Kif17, Is essential for Photoreceptor Outer Segment Development. Developmental Biology. 2008.

Luby-Phelps, K., Fogerty, J., Baker, S. A., Pazour, G. J., and J. C. Besharse.  Spatial Distribution of Intraflagellar Transport Proteins in Vertebrate Photoreceptors. Vision Research. Feb;48(3):413-23. 2007.

Green, C.B., Douris, N., Kojima, S, Strayer, C.A., Fogerty, J., Lourim, D., Keller, S.A., and J. C. Besharse. Loss of nocturnin, a circadian deadenylase, confers resistance to hepatic steatosis and diet-induced obesity. Proceedings National Academy Sciences. 104: 9888-9893. 2007.

Gabarino-Pico, E., S. Niu, M. D. Rollag, C. A. Strayer, J.C. Besharse, and C.B. Green.  Immediate early responses of the circadian poly A ribonuclease nocturnin to two extracellular stimuli. RNA. 13: 745-755. 2007.

Besharse JC, M Zhuang, K Freeman and J Fogerty. Regulation of Photoreceptor Per1 and Per2 by Light, Dopamine, and a Circadian Clock. European Journal of Neuroscience 20:167-174, 2004.

Peet J, A Bragin, PS Calvert, SS Nikonov, S Mani, J Zhao, JC Besharse, E Pierce, B Knox and EN Pugh, Jr: Quantification of the cytoplasmic spaces of living cells with EGFP reveals arrestin-EGFP to disequilibrium in dark adapted rod photoreceptors. J Cell Science 117:3049-3059, 2004.

Green CB and JC Besharse: Retinal Circadian Clocks and Control of Retinal Physiology. J Biological Rhythms 9:91-102, 2004.

Besharse JC, SA Baker, K Luby-Phelps and GJ Pazour. 2003. Photoreceptor intersegmental transport and retinal degeneration, a conserved pathway common to motile and sensory cilia. Adv Exp Med Biol 533:157-64, 2003.

Baker SA, K Freeman, K Luby-Phelps, GJ Pazour and JC Besharse: IFT20 links kinesin II with a mammalian intraflagellar transport complex that is conserved in motile flagella and sensory cilia. J Biological Chemistry 278:34211-34218, 2003.

Luby-Phelps K, G Ning, J Fogerty and JC Besharse: Visualization of identified GFP-expressing cells at the light and electron microscopic level. J Histochem. and Cytochem 51:271-274, 2003.

Mikami A, SH Tynan, T Hama, K Luby-Phelps, T Saito, JE Crandall, JC Besharse and RB Vallee: Molecular structure of cytoplasmic dynein 2 and its distribution in neuronal and ciliated cells. J Cell Science 120:4801-4808, 2002.

Pazour GL, SA Baker, JA Deane, DG Cole, BA Dickert, JL Rosenbaum, GB Witman and JC Besharse: The intraflagellar transport protein, IFT88, is essential for photoreceptor assembly and maintenance. J Cell Biology 157:103-114, 2002. Pharmacogentics 12:55-65, 2002.

Wang Y, DL Osterbur, PL Megaw, G Tosini, C Fukuhara, CB Green and JC Besharse: Rhythmic expression of nocturnin mRNA in multiple tissues of the mouse. BMC Developmental Biology 1:9, 2001.

Besharse JC: Preview: Coupling an activated MAP kinase pathway to circadian output clock. Neuron 29:3-4, 2001.

Zhuang M, Y Wang, BM Steenhard and JC Besharse: Differential regulation of two period genes in the Xenopus eye. Molecular Brain Research 82: 52-64, 2000.

Steenhard BM and JC Besharse: Phase shifting the retinal circadian clock: xPer2 mRNA induction by light and dopamine. Journal of Neuroscience 20: 8572-8577, 2000.

Hollander BA, M-Y Liang and JC Besharse: Linkage of a nucleolin-related protein and casein kinase II with the detergent stable, photoreceptor cytoskeleton. Cell Motility and the Cytoskeleton 43:114-127, 1999.

Mani SS, JC Besharse and BE Knox: Immediate upstream sequence of arrestin directs rod specific expression in Xenopus. Journal Biological Chemistry 274: 15590-15597, 1999.

Knox BE, C Schlueter, BM Sanger, CB Green and JC Besharse: Transgene expression in Xenopus rods. FEBS Letters 423:117-121, 1998.

Muresan V, E Bendala-Tufanisco, BA Hollander and JC Besharse: Evidence for kinesin-related proteins associated with the axoneme of retinal photoreceptors. Exp. Eye Research 64:895-903, 1997.

Green CB and JC Besharse: Identification of a novel vertebrate circadian clock regulated gene encoding the protein nocturnin. Proc. Nat. Acad. Sci. 93:14884-14888, 1996.

Green CB and JC Besharse: A high stringency differential display screen for circadian clock regulated retinal mRNAs. Molecular Brain Research 37:157-165, 1996.

Besharse JC and MG Wetzel: Immunocytochemical localization of opsin in rod photoreceptors during periods of rapid disc assembly. J Neurocytology 24:371-388, 1995.

Cahill GM and JC Besharse: Circadian clock functions localized in Xenopus retinal photoreceptors. Neuron 10:573-577, 1993.