students

Merkel cell carcinoma is an aggressive skin cancer that affects immunosuppressed and elderly individuals and is thought to arise from mechanoreceptor Merkel cells (neuroendocrine derived). Our laboratory discovered a new virus, which is closely associated with this disease. MCV or MCPyV (Merkel Cell Polyomavirus) is the fifth known polyomavirus to infect humans. MCPyV has a 5387bp genome and shares significant homology to murine polyomavirus and SV40. My projects involve: 1) determining whether other human neoplasms are also associated with MCPyV and 2) investigating the oncogenic potential of MCPyV.

Mycobacteriophages generally have two lysins of different function as well as a holin. The holins initiate lysis of bacteria at a specific point in the viral life cycle. The holins form pores or holes in the cell membrane, allowing lysins access to the bacterial cell wall. The lysins (generally there are two) digest the peptidoglycan wall with one or more enzymatic activities, such as amidase or peptidase activity. The degradation of the cell wall causes the bacteria to lyse due to the greater internal pressure, and the new phage particles are released. Given that lysins act from the outside of bacteria to kill them, they have potential as a treatment for bacterial infections. Other studies with pneumococcal phage lysins have shown lysins abilitiy to clear an infection with high specificity to the target bacteria and little to no developing resistance (Jado et al 2003; Entenza et al 2005). For my dissertation research, I will begin characterizing the enzymatic activity and binding properties of a variety of lysins (beginning with the LysAs) from mycobacteriophages that have been isolated in the Hatfull Lab. Eventually I hope to generate a more potent lysin through mutagenesis, deletions, and domain-swapping of various lysins.

The vertebrate kidney is a complex homeostatic organ that functions to detoxify blood, maintain ion and water equilibrium, and regulate hormone release. The physiological consequences of abnormal kidney formation or function are frequently fatal, with dialysis and organ transplantation the only long-term treatments for kidney disease. Future strategies to fight kidney disease must rely on a fundamental understanding of the earliest events that lead to the formation of the embryonic kidney. Importantly, kidney structure and function is conserved among vertebrates and current evidence suggests that mechanisms of kidney development are similarly conserved. I propose to exploit the embryological and genetic features of Xenopus and zebrafish embryos to (1) identify the earliest cells (progenitors) that give rise to the pronephric kidney and (2) understand the molecular events that initiate and regulate kidney development during embryogenesis.

Membrane proteins are encoded by 30% of the human genome and they constitute more than half of the drug targets in the market. Among them, G-protein coupled receptors (GPCRs), encoded by 1% of the human genome, play a very important role in cell signaling. Rhodopsin, a class A GPCR, is the primary photoreceptor in the mammalian retina. Specific misfolding mutations in rhodopsin cause Retinitis Pigmentosa, a retinal degeneration disease that leads to blindness affecting 1 out of every 3500 persons in the USA. Misfolding of rhodopsin is believed to be a major cause of the disease. My thesis work is aimed at understanding the molecular mechanisms of (mis)folding of rhodopsin. Currently, the most widely accepted model for helical membrane protein folding is the two-stage hypothesis. In this model, the transmembrane helices form first, followed by tertiary contact formation. However, based on preliminary studies of our lab, rhodopsin folding may not conform to this model. It has been proposed that long range interactions between the helical and loop regions play an important role in early stages of folding of rhodopsin. I propose to study the mechanism of folding of rhodopsin by investigating the importance of tertiary interactions preceeding the formation of helices.

Protein polymerization into macromolecular structures

The epithelial sodium channel (ENaC) is a multimeric transmembrane protein involved in sodium transport across the apical membrane of epithelial cells in various tissues. ENaC is a substrate of ER associated degradation (ERAD). The ERAD pathway recognizes misfolded proteins in the endoplasmic reticulum and targets them for proteosomal degradation. Different factors are involved in recognizing different ERAD substrates. I am examining the factors involved in the recognition of ENaC as an ERAD substrate and the mechanism of its degradation.

Kaposi's sarcoma-associated herpesvirus (KSHV; also known as human herpesvirus 8) is the infectious cause of Kaposi's sarcoma (KS), the most common AIDS related malignancy. The KSHV genome contains a remarkable array of cellular gene homologs, encoding proteins involved in cell cycle regulation or cell signaling. Latency-associated nuclear antigen (LANA), encoded by open reading frame (ORF) 73 of the KSHV genome, is a latent protein consistently expressed in all KSHV-associated diseases. LANA is important in viral genome maintenance and is associated with cellular and viral proteins to regulate viral and cellular gene expression. We are interested to investigate the function of LANA and the role of specific KSHV inhibitors of interferon signaling and their role in contributing to KSHV tumorigenesis.

Our lab focuses on the apical membrane trafficking regulation in polarized epithelial cells. My current project is working on the effect of Galectin-3, one of the conserved lectin families on apical membrane trafficking. Endolyn, HA, p75 and Muc1 are used as apical sorting marker proteins for study. Live cell imaging in vivo and biochemical experiments in vitro are used to trace proteins sorting and transport.

The G protein coupled receptor (GPCR) family is with more than 8000 reported sequences the largest group of cell-surface receptors known. The sequences in this family are very diverse and the corresponding receptors respond to a wide range of signals, including light, hormones and neurotransmitters. It is an important and unsolved question to what extent the structures of different GPCR’s are conserved, because the homology between sequences is often very low. Furthermore, the activity of many receptors is modulated by secondary ligands, suggesting that there are two or more ligand binding sites on many GPCR’s. During my dissertation research, I will study the effects of small molecule ligands on the structure and dynamics of different GPCR class members. I will be using a combination of computational and experimental biophysical and biochemical approaches.

The Src Family of Tyrosine Kinases(SFKs) plays a central role in celluar growth, differentiation and function. Our lab discovered that murine Embryonic Stem(ES) cells express seven of eight SFKs.The activity and gene expression profiles change with the differentiation state of ES cells.Pharmacological inhibition of overall SFKs blocks differentiation of ES cells while maintain there pluripotency. It is hypothesised that individual SFK member plays non-redundant roles in ES cell pluripoteny maintenance and differentiation. Rightnow I am working on the roles individual Src Family member plays in ES cell function and fate determination.