Andrey Shaw, MD
Professor, Pathology and Immunology
Our work is focused in three major areas:
Immune Cell Activation
Our current focus is to develop real-time sensors that would allow us to monitor the activation of signaling pathways in vivo. Our approach is to use mouse knock-in or BAC recombineering technology to generate reporters that would allow us to monitor gene expression in vivo using two-photon microscopy or to generate novel reporters that would read-out the activation of various signaling pathways.
Our current projects involve generating a calcium FRET reporter mouse that would allow us to monitor calcium signaling in real-time in a live animals. Our strategy involves manipulating the well-studied Cameleon FRET reporter, generate an inducible transgene, and then monitor calcium signaling in any cell of interest in a variety of differentinflammation and disease models. We are also generating a reporter for the important MAP kinase, ERK2. ERK is a very important kinase activated by a wide variety of different types of stimuli. Our current strategy is to replace endogenous ERK with a GFP-fusion protein of ERK. Our rationale is that since ERK translocates to the nucleus when it is activated, we could use the sub-cellular localization of ERK2 as a surrogate read-out for its state of activation.
Podocyte biology and glomerular disease
The podocyte is one of three kinds of cells that make up the glomerulus, a collection of capillaries that constitute the primary blood filtration apparatus. The podocyte has a fascinating morphology with multiple long foot processes that cover the entire outside surface of the capillary. The exact function of the podocyte is unknown but it has been implicated as a component of the filtration barrier.
Mutations in CD2AP are involved in the pathogenesis of the human disease focal segmental glomerulosclerosis (FSGS). We are currently using NextGen sequencing as a method of identifying candidate genes that are involved in FSGS epistatic to CD2AP. We have identified about 2500 podocyte specific genes and plan to sequence 1000 patients in the next two years. Our preliminary analysis has identified about 60 genes that are implicated in the human disease focal segmental glomerulosclerosis.
We are currently using mouse genetics to develop a system to test candidate genes in mice. Our system involves the generation of a sensitized ES cell (heterozygous for both CD2AP and synaptopodin), a method for efficient and rapid knock-in of RNAi constructs(into the hgprt locus) and a method of generating pure mice from ES cells (eliminates having to breed animals). We anticipate that our genetic system will allow us to rapidly screen candidate genes, identified by humansequencing, for their role in the progression of kidney failure.
KSR and its role in ERK activation
Kinase suppressor of Ras (KSR) was originally identified in the fly and the worm as a gene that is important in the Ras/ERK signaling pathway. The current model is that KSR is a pseudokinase that functions as a scaffold by binding to Raf, MEK and ERK. Recently, we and others provided evidence to suggest that KSR may function also as a kinase. Our current work focuses on the role that the phosphorylation and kinase activity KSR might play in RAF/MEK/ERK activation and whether KSR would be a good target for cancer therapies. We are also working with Dr. Lynn Cornelius (Dermatology) to understand why BRAF inhibitors canparadoxically activate the Ras/MAP kinase pathway in certain situations.
Cemerski S, Das J, Giurisato E, Markiewicz MA, Allen PM, Chakraborty AK, Shaw AS. Evidence for signaling from the center of the immune synapse. Immunity 2008;29:414-422.
Kao H, Lin J, Littman DR, Shaw AS, Allen PM. Regulated movement of CD4 in and out of the immunological synapse. J Immunol 2008;181:8248-8257.stub
Giurisato E, Lin J, Harding A, Cerutti E, Cella M, Lewis RE, Colonna M, Shaw AS. The MAP kinase scaffold KSR1 is required for recruitment of ERK to the immunological synapse. Mol Cell Biol 2009;29:1554-1564.
Markiewicz MA, Wise EL, Buchwald ZS, Cheney EE, Hansen TH, Suri A, Cemerski S, Allen PM, Shaw AS. IL-12 enhances CTL synapse formation and induces self-reactivity. J Immunol 2009;182:1351-1361.
Lin J, Harding A, Giurisato E, Shaw AS. KSR1 modulates the sensitivity of the MAPK pathway activation in Tcells without altering fundamental system outputs. Mol Cell Biol. 2009;29:2082-2091.
Otero K, Turnbull IR, Poliani PL, Vermi W, Cerutti E, Aoshi T, Tassi I, Takai T, Stanley SL, Miller M, Shaw AS, Colonna M. Macrophage colony-stimulating factor induces the proliferation and survival of macrophages via a pathway involving DAP12 and beta-catenin. Nat Immunol 2009;10:734-743.
Lin J, Hou KK, Piwnica-Worms H, Shaw AS. The polarity protein Par1b/EMK/MARK2 regulated T cell receptor-induced microtubule-organizing center polarization. J Immunol 2009, 183:1215-1221.
Hartleben B, Godel M, Meyer-Schwesinger C, Liu S, Ulrich T, Kobler S, Wiech T, Grahammer F, Arnold SJ, Lindenmeyer MT, Cohen CD, Pavenstadt H, Kerjaschki D, Mizushima N, Shaw AS, Walz G, Huber TB. Autophagy influences glomerular disease susceptibility and maintains podocyte homeostasis in aging mice. J Clin Invest 2010;120:1084-1096.
Filbert EL, Nguyen A, Markiewicz MA, Fowlkes BJ, Huang YH, Shaw AS. Kinase suppressor of Ras 1 is required for full ERK activation in thymocytes but not for thymocyte selection. Eur J Immunol 2010;40:3226-3234.
Hu J, Yu H, Kornev A, Zhao J, Filbert EL, Taylor SS, Shaw AS. Mutation that blocks ATP binding creates a pseudokinase stabilizing the scaffolding function of the kinase suppressor of Ras, CRAF and BRAF. Proc Natl Acad Sci 2011;108:6067-6072.
Akilesh S, Suleiman H, Yu H, Stander MC, Lavin P, Gbadegesin R, Antignac C, Pollak M, Kopp JB, Winn MP, Shaw AS. Arhgap24 inactivates Rac1 in mouse podocytes, and a mutant form is associated with familial focal segmental glomerulosclerosis. J Clin Invest 2011;121:4127-4137.
Markiewicz MA, Wise EL, Buchwald ZS, Pinto AK, Zafirova B, Polic B, Shaw AS. RAE1-epsilon ligand expressed on pancreatic islets recruits NKG2D receptor expressing cytotoxic T cell independent of T cell receptor recognition. Immunity 2012; 36:132-141.
Filbert EL, LeBorgne M, Lin J, Heiser JE, Shaw AS. Stathmin regulates microtubule dynamics and microtubule organizing center polarization in activated T cells. J Immunol 2012; 188:5421-5427.
Cheney, E.E., et al., A dual function of NKG2D ligands in NK-cell activation. Eur J Immunol, 2012. 42(9): p. 2452-8.
Zhao, J., et al., CD2AP Links Cortactin and Capping Protein at the Cell Periphery to Facilitate Lamellipodia Formation. Mol Cell Biol, 2012.