Russ P. Carstens, M.D.
University of Pennsylvania
575 Clinical Research Building
415 Curie Boulevard
Philadelphia, PA 19104
Undergraduate: Johns Hopkins University, Baltimore, MD
Medical: Yale University School of Medicine, New Haven, CT
Residency: Duke University Medical Center, Durham, NC
Fellowship: Duke University Medical Center, Durham, NC
Board Certification: Internal Medicine and Nephrology
Study of the molecular mechanisms of alternative splicing. Identification of global changes in splicing during the epithelial-mesenchymal transition (EMT).
Description of Research:
The focus of my laboratory is investigation of alternative splicing, whereby a single gene transcript can generate numerous spliced mRNAs, thereby greatly expanding ribonomic and proteomic diversity. We are particularly interested in the regulation of cell and tissue-specific splicing choices that are important during development of numerous organs including the kidney. Our previous studies focused on alternative splicing of fibroblast growth factor receptor 2 (FGFR2) as a model system. Mutually exclusive splicing of two exons, IIIb and IIIc, gives rise to two functionally different receptors, FGFR2-IIIb and FGFR2-IIIc, in epithelial and mesenchymal cells, respectively. The exquisite cell type-specific expression of these epithelial or mesenchymal specific splice variants is essential during vertebrate development and a switch in FGFR2 splicing occurs during the epithelial to mesenchymal transition (EMT). We recently used luciferase-based splicing reporter assays to carry out a genome-wide, high throughput cDNA screen for factors that regulate FGFR2 splicing. These studies identified a number of novel mammalian splicing regulators, including two epithelial-specific factors that we named Epithelial Splicing Proteins 1 and 2 (ESRP1 and ESRP2). Expression of these splicing regulators is required for expression of the epithelial FGFR2 splice variant and ongoing work has shown that they regulate the splicing of an extensive epithelial-specific splicing program. These regulated targets include functionally relevant splicing switches that are implicated in the EMT and both ESRPs are transcriptionally inactivated during the EMT. We are currently carrying out massively parallel high throughput sequencing (RNA-seq) to identify an even more comprehensive epithelial splicing regulatory network (SRN). We predict that the proteins encoded by transcripts comprising this epithelial splicing signature will also define a protein interaction network that underlies important epithelial cell properties. Furthermore, we also have strong reasons to believe that, like FGFR2, many of these proteins will exhibit previously unrecognized isoform-dependent differences in function between epithelial and mesenchymal cells. These proteins and networks are likely to have biologically coherent functions that are relevant for the EMT in development, renal fibrosis, and cancer metastasis as well as epithelial cell differentiation in diverse tissues and organs. In our ongoing studies we will further investigate the molecular mechanisms by which the ESRPs and several additional novel splicing regulators cause switches in splicing. We are also generating mice carrying conditional knockout alleles for Esrp1 and Esrp2 (and both) that will be used to create tissue-specific Esrp-knockout mice. These tools will allow us to clarify the roles of these splicing regulators during development and also allow us to profile ESRP-regulated splicing targets in vivo.
A number of potential projects to investigate the targets and functions of the ESRPs can be discussed. In addition to further characterizing the molecular mechanisms of ESRP function, we are generating mice with conditional knockout of the ESRPs in order to further investigate their functions in vivo. Among several areas of interest, we plan to investigate the role of the ESRPs in renal organogenesis in which we anticipate they regulate splicing events that are required for the mesenchymal to epithelial transition (MET) in metanephric mesenchyme during kidney development.
Hovhannisyan, R.H. and Carstens, R.P.: A novel intronic cis-element, ISE/ISS-3, regulates rat fibroblast growth factor receptor 2 (FGFR2) splicing through activation of an upstream exon and repression of a downstream exon containing a non-canonical branchpoint sequence. Mol. Cell. Biol., 25(1): 250-263 25(1): 250-263, January 2005.
Hovhannisyan, R.H., Warzecha, C. and Carstens, R.P.: Characterization of sequences and mechanisms through which ISE/ISS-3 regulates FGFR2 splicing. Nucl. Acids Res. 34(1): 373-385, January 2006
Newman, E.A., Muh, S.J., Hovhannisyan, R.H., Warzecha, C.C., Jones, R.B., McKeehan, W.L. and Carstens, R.P.: Identification of RNA-binding proteins that regulate FGFR2 splicing through the use of sensitive and specific dual color fluorescence minigene assays. RNA 12(6): 1129-1141, June 2006.
Hovhannisyan, R.H. and Carstens, R.P.: Heterogeneous ribonucleoprotein M (Hnrnp M) is a splicing regulatory protein that can enhance or silence splicing of alternatively spliced exons Journal of Biological Chemistry 282(50): 36265-36274, December 2007.
Warzecha, C.C., Sato, T.K., Nabet, B., Hogenesch, J.B., and Carstens, R.P.: ESRP1 and ESRP2 are epithelial cell type-specific regulators of FGFR2 splicing. Molecular Cell 33(5): 591-601, March 2009.
Warzecha, C.C., Shen, S., Xing, Y, and Carstens, R.P.: The epithelial splicing factors ESRP1 and ESRP2 positively and negatively regulate diverse types of alternative splicing events. RNA Biology 6(5): 546-62, November 2009.
Warzecha, CC, Jiang, P., Amirikian, K., Dittmar, K.A., Lu, H., Shen, S., Guo, W., and Carstens, R.P. An ESRP-regulated splicing programme is abrogated during the Epithelial Mesenchymal Transition. EMBO Journal August (advance online version) 2010.