Specificity/Metal Inhibition of Interaction Gata-1/DNA

Hope College, Holland MI

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Abstract

[unreadable] DESCRIPTION (provided by applicant): This is a continuation of the study described in the proposal "Specificity of the interaction between Gata-1 and DNA." The purpose of the ongoing investigation is to probe 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 as 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. Thus, 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 factor contributes to blood cell production and maturation. Specifically, this work examines the role of 5 amino acid side chains in direct contact with the DNA major groove when Gata binds DNA. Several mutations to these structurally central amino acids have been found which preserve DNA binding in Gala's minimal DNA binding domain. Because these represent mutants not found in naturally-occurring Gata-1, current plans are to introduce these changes back into full- length Gata. Additionally, a strongly inhibitory effect of the toxic heavy metal leads on Gata DNA binding and function has been found. Since lead is known to exert pathological effects on the hematopoietic system, a goal of this proposal to test whether Gata is a molecular target for lead toxicity in cell culture systems. [unreadable] [unreadable] [unreadable]