Postdoctoral training, Samuel Lunenfeld Research Institute, Mount Sinai Hospital,Toronto

Ph.D., Biochemistry, University of California, San Francisco

B.A., Biochemistry, Northwestern University

Genetic and biochemical analysis of cell signaling during hematopoiesis

We are interested in how cells integrate the different signals they receive and make correct developmental decisions. To explore this question we are studying murine hematopoiesis. There are several advantages to this model system. Hematopoietic differentiation relies for the most part on extracellular signals (growth factors and cytokines) to stimulate proliferation, survival, and maturation of progenitors, which makes it an ideal system to explore the how different signaling pathways interact during differentiation. Our approach is to combine genetic analysis of mouse mutants that have hematopoietic defects with biochemical analysis of cell signaling in primary progenitor cell cultures. The mouse mutants we are studying are not targeted mutations, but rather spontaneous or mutagen induced mutations that were identified by their hematopoietic phenotypes. Thus the characterization of the genes mutated at these loci will identify new genes that play key roles in the regulation of hematopoiesis. In the laboratory we have been concentrating on the analysis of three mutations that affect erythropoiesis, the Friend virus susceptibility gene 2, (Fv2), Polycythemia, (Pcm) and flexed-tail, (f).

Fv2/Stk, a host gene involved in Friend virus induced erythroleukemia

Fv2 as its name suggests was first identified because it controls susceptibility to infection by Friend erythroleukemia virus. Our interest in Fv2 is in the mechanism by which it restricts Friend virus infection. Fv2 is thought to accomplish this role in two ways. 1) It regulates the cell cycling of BFU-E, the early erythroid progenitor that is the target for Friend virus infection. A productive retroviral infection requires that the target cell be actively cycling. 2) Early in infection, Friend virus activates the Epo receptor (EpoR) signaling pathway to expand the population of infected cells. Fv2 appears to negatively regulate EpoR signaling, which blocks the expansion of infected cells. Thus, Fv2 appears to be a regulator of cell cycling and EpoR signaling in erythroid cells. Our initial genetic analysis of the Fv2 locus lead to a surprising result. It demonstrated that linked genes, one that controls cell cycling and the second that regulates EpoR signaling and determines susceptibility of Friend virus infection, encode the two functions of Fv2. Current work in the laboratory has identified a naturally occurring mutation in the cell cycle regulator Cdc25A that segregates with the altered cell cycling of erythroid progenitors. Furthermore, in collaboration with Pam Correll at Penn State and Paul Ney at St. Judes Hospital in Memphis we have demonstrated that Fv2 encodes the receptor tyrosine kinase Stk. We are currently investigating the mechanism by which Stk interacts with Friend virus and the EpoR to regulate Friend erythroleukemia.

Polycythemia (Pcm) a murine model for human Polycythemia rubra vera

Human myeloproliferative disorders are a group of related diseases that are characterized by the chronic over-proliferation of multiple hematopoietic lineages. One of these disorders, Polycythemia rubra vera is characterized by the overproduction of erythroid, granulocytic and megakaryocytic cells. The murine Polycythemia (Pcm) mutation causes a myeloproliferative disease that closely resembles human Polycythemia rubra vera. Current work in the laboratory has demonstrated that bone marrow cells from Pcm mice exhibit hypersensitivity to Epo, IL-3, SCF and Tpo. The Pcm mutation appears to alter the threshold for activation of signaling pathways downstream of cytokine receptors. Presently we are trying to characterize this defect biochemically. In addition we are generating a high-resolution genetic linkage map of the Pcm locus as a prelude to cloning the gene.

flexed-tail (f), a regulator of fetal liver and stress erythropoiesis

flexed-tail mutant mice exhibit a severe fetal-neonatal anemia that resolves shortly after birth. Adult f/f mice are normal except when placed under severe erythropoietic stress where they are very slow to respond. Current work in the laboratory has identified an early defect in the development of f/f fetal liver hematopoietic progenitors. In addition we have also identified a defect in the erythroid response to acute anemia. Our work has suggested that f/f mice may have a defect in an erythroid progenitor that is specific to the fetal liver and in the adult responds to erythropoietic stress.

Anamaria Craici, Michele Yon, Li Hao and Robert Paulson. The murine Polycythemia(Pcm) mutation causes alterations in Epo and IL-3 signaling pathways and results in myeloproliferative disease. Manuscript in preparation.

Edward Melkun, Alan Bernstein and Robert Paulson. A naturally occurring point substitution in Cdc25A and not Fv2 is responsible for altered cell cycling of erythroid progenitors. Manuscript submitted.

Lisa Finkelstein, Robert Paulson, and Pam Correll. Expression of Sf-Stk confers Friend virus sensitivity in vitro: Requirement for kinase activity and the Grb2 binding site. 2001 Blood submitted

D. A. Persons, R. F. Paulson, M. R. Loyd, M. T. Herley, S. M. Bodner, A. Bernstein, P. H. Correll, and P. A. Ney. 1999. Fv2-mediated susceptibility to Friend erythroleukemia virus is conferred by a truncated form of the Stk/RON receptor tyrosine kinase. Nature Genetics 23:159–165.

R. Paulson and A. Bernstein. 1998. A genetic linkage map of the mouse chromosome 9 region encompassing the Friend virus susceptibility gene 2 (Fv2). Mammalian Genome 9:381–384.

X. Piao, R. Paulson, P. van der Geer, T. Pawson, and A. Bernstein. 1996. Oncogenic mutation in the Kit receptor tyrosine kinase alters substrate specificity and induces the degradation of the protein tyrosine phosphatase Shp1. Proceedings of the National Academy of Sciences, USA 93:14665–14669.

R. Paulson, S. Vesily, K. Siminovitch, and A. Bernstein. 1996. Signaling by the W/Kit receptor tyrosine kinase is negatively regulated in vivo by the protein tyrosine phosphatase Shp1. Nature Genetics 13:309–315.