? DESCRIPTION (provided by applicant): The ability to generate multiple cell types, including erythrocyte lineage cells, from hematopoietic precursors enables bone marrow transplantation and treatment of a wide variety of diseases. We found that the transcription factor ARID3a is required for normal hematopoietic stem cell development in the mouse. Mice deficient in ARID3a die in utero as the result of failed erythropoiesis. Additional data show ARID3a mediates hematopoietic lineage decisions in human cord blood cells in vitro. Furthermore, we demonstrated that ARID3a has suppressive functions in adult differentiated cell types that regulate expression of pluripotency genes. Together, our data implicate ARID3a as an important DNA-binding protein in hematopoietic stem cells and as a barrier for reprogramming of induced pluripotent stem cells. However, nothing is known regarding the molecular mechanisms by which ARID3a mediates lineage decision effects in these systems. Although our data suggest ARID3a can participate in suppression of gene expression, as well as enhancement of gene transcription, no protein motifs within ARID3a suggest mechanisms which might mediate these functions. We hypothesize that ARID3a interacts with other proteins by serving as a DNA tether to affect gene regulation. In order to understand how ARID3a functions it is critical to identify te proteins with which it is associated. Our data indicate that the human K562 myelomonocytic cell line which can be induced to differentiate into both erythroid and myeloid lineage cells requires ARID3a for differentiation. Existing data from the ENCODE group identify a large number of potential gene targets for ARID3a in K562 cells. Therefore, we will use this cell line as a model system to identify proteins associated with ARID3a by mass spectrometry. Transcriptome analyses of ARID3a knockdown and control K562 cells will identify genes requiring ARID3a for expression. Identified gene targets for ARID3a will be examined for the presence of ARID3a-containing protein complexes. We predict that ARID3a-interacting proteins will include epigenetic mediators, such as histone deacetylases, providing important clues for how ARID3a regulates gene expression. Future studies will utilize these data to develop a better mechanistic understanding of how ARID3a participates in lineage decisions in human hematopoietic stem cells. Ultimately, the ability to manipulate hematopoietic stem cells to generate lineages of choice could have therapeutic use.

R03AI123951

2016-04-01

2019-03-31