Deleting an E-box-GATA intronic cis-element (+9.5) reduces GATA-2 expression and inactivates the hematopoietic stem cell (HSC) generator in the mouse embryo by disrupting GATA-2-dependent genetic networks. Many of the essential constituents of this genetic network have not been defined. To discover components of this network, the +9.5 sequence/molecular properties were compared to thousands of +9.5-like cis-elements genome-wide. This innovative training method yielded a genome-wide library of +9.5-like sites, containing GATA-2 target genes that may inform unique modes of controlling hematopoietic stem and progenitor cells (HSPCs). I discovered that GATA-2 regulates expression of the unstudied Sterile Alpha Motif Domain 14 (Samd14) gene, and Samd14 increased fetal liver HSPC levels and promoted SCF/c-Kit signaling. Initial data demonstrates that the +9.5-like Samd14-Enhancer mediates Samd14 expression, is required for stress erythropoiesis, and confers stress-dependent Samd14 upregulation in the spleen.
These aims will rigorously establish the mechanism by which Samd14 promotes SCF/c-Kit signaling (Aim 1), the function of the GATA-2-regulated Samd14-Enh in an innovative mouse model (Aim 2), and define a cohort of stress- dependent genes and their interconnectivity (Aim 3). As a K award recipient, I will follow a detailed training plan with specific research and career development milestones to be achieved over the course of the award.
The Aims are designed to expand and distinguish my research expertise from my mentor, and we have agreed that I can use the Samd14 work as a foundation for developing my independent research program. The career development activities outlined in the proposal will provide training in writing/presentation skills, laboratory management, and grantsmanship. I have organized a mentoring committee comprised of well-established and successful scientists, with research programs relevant to my work, who are committed to my success and will play an active role in facilitating my career transition. A key element of this plan entails direct training in bone marrow transplantation techniques to study stress hematopoiesis. UW-Madison is a top-tier research institution with outstanding NIH-sponsored training programs, core facilities, and research laboratories, which makes it an ideal training environment for me to further develop as an independent scientist. I have strong support from UW-Madison, the Cell and Regenerative Biology Department and Dr. Bresnick to pursue these career goals and compete for a faculty position here or at another research-intensive academic institution.
The research Aims, career development training plan, meetings with mentoring committee and collaborators, educational opportunities, and training environment will ensure the development of a solid foundation to launch a productive academic career.
I discovered a new constituent of the quintessential SCF/c-Kit signaling pathway, Samd14. This project will advance our mechanistic understanding of the relationship between Samd14 and SCF/c-Kit signaling, with potentially-important therapeutic implications.
|Katsumura, Koichi R; Mehta, Charu; Hewitt, Kyle J et al. (2018) Human leukemia mutations corrupt but do not abrogate GATA-2 function. Proc Natl Acad Sci U S A 115:E10109-E10118|
|Bresnick, Emery H; Hewitt, Kyle J; Mehta, Charu et al. (2018) Mechanisms of erythrocyte development and regeneration: implications for regenerative medicine and beyond. Development 145:|
|McIver, Skye C; Hewitt, Kyle J; Gao, Xin et al. (2018) Dissecting Regulatory Mechanisms Using Mouse Fetal Liver-Derived Erythroid Cells. Methods Mol Biol 1698:67-89|
|Hewitt, Kyle J; Katsumura, Koichi R; Matson, Daniel R et al. (2017) GATA Factor-Regulated Samd14 Enhancer Confers Red Blood Cell Regeneration and Survival in Severe Anemia. Dev Cell 42:213-225.e4|
|Katsumura, Koichi R; Bresnick, Emery H; GATA Factor Mechanisms Group (2017) The GATA factor revolution in hematology. Blood 129:2092-2102|