Project Description: |
The Pikaart lab focuses on several aspects of transcription regulation. First, we study the molecular basis for the specific binding of Gata-1, a hematopoietic transcription factor, to DNA. Gata-1 is required for the maturation of bone marrow stem cells into functional circulating blood cells. Gata-1 functions in the regulation of cell growth, allowing for continuing cell division during hematopoietic differentiation, as well as activating transcription of
blood cell-specific proteins, such a hemoglobin.
Interplay between the several Gata protein family members and with other transcription factors, both general and tissue-specific, is complex and involves subtle distinctions among various regulatory outcomes depending on context. We seek a more thorough characterization of the means by which Gata recognizes regulatory binding sites in DNA will enhance our understanding of how this key hematopoietic transcription factors contributes to blood cell production and maturation.
Specifically, this work has examined the role of five amino acid side chains in direct contact with the DNA major groove when Gata binds DMA. We have found that several mutations to these structurally central amino acids can preserve DNA binding in Gala's minimal DNA binding domain. Additionally, we have discovered a strongly inhibitory effect of the toxic heavy metal lead on Gata DNA binding and function.
During the coming summer, most of the focus will be on measuring direct effect of lead on Gata-1 function in cell culture transactivation models as well as in mouse bone marrow cultures.
As a second area of focus, we have started a new study on the role of histone modification (particularly ubiquitylation) as a mechanism of chromatin mediated gene regulation. This are of work as begun in collaboration with Dr. Aaron Best of Hope's biology department. We have prepared stains of Giardia, a single-celled eukaryotic organism, containing modified histone H2B genes which we are analyzing for ubiquitylation.
Work on the Giardia chromatin project will involve preparing and sequencing gene constructs based on the H2B gene and culture of Giardia cells expressing those constructs. Giardia grown in this way will be subjected to protein extraction and analysis by western blotting. |
