Regulation Of Erythroid Gene Expression

PU.1是与加塔-1 DNA结合相互作用的转录因子 结构域，并且在决定造血谱系内细胞命运的两种蛋白质之间存在相互抑制。PU.1 据报道，反式激活（transactivation）结构域和DNA结合结构域被 参与互动，我们已经注意到，PU.1 与p53的cDNA具有结构同源性。协同 在Jim Omichinski的实验室中，我们已经表明p53 β在体外和体内也与加塔-1 DNA结合结构域相互作用。加塔-1的接头和C-指是相互作用所必需的。这些蛋白质可抑制 在红系前体细胞系6C 2中，可能需要加塔-1来防止在红细胞形成之前的核浓缩和去核期间或在巨核细胞的多倍化期间的p53诱导。我们与马西山本合作，在GATA 1.05细胞和小鼠的拯救试验中测试不与p53相互作用的加塔-1突变体，以确定这种相互作用在造血过程中的作用。 另一个项目涉及与EKLF转录因子相关的贫血。 EKLF DNA结合域的突变发生在贫血患者中。在CBA患者和Nan（新生儿贫血）小鼠中，EKLF第二锌指中的氨基酸发生突变。在与Jim Bieker的合作中，我们试图确定贫血表型是否基于Nan-EKLF相对于野生型对某些EKLF靶基因的不同DNA结合亲和力。

PU.1 is a transcription factor that interacts with the GATA-1 DNA binding domain, and there is reciprocal inhibition between the two proteins that determines cell fate within the hematopoietic lineages. The PU.1 transactivation (TAD) and DNA binding domains are reported to be involved in the interaction and we have noted that the PU.1 TAD shares structural homology with the TAD of p53. In collaboration with the laboratory of Jim Omichinski we have shown that the p53 TAD also interacts in vitro and in vivo with the GATA-1 DNA binding domain. The linker and C-finger of GATA-1 are required for the interaction. The proteins reciprocally inhibit the transactivation activity of one another in an erythroid precursor cell line, 6C2. GATA-1 may be required to prevent p53 induction during the nuclear condensation and enucleation that precedes erythrocyte formation or during the polyploidization of megakaryocytes. In collaboration with Masi Yamamoto we are testing mutants of GATA-1 that do not interact with p53 in rescue assays in GATA1.05 cells and mice to determine the role of this interaction during hematopoiesis. Another project involves anemias associated with the EKLF transcription factor. Mutations in the EKLF DNA binding domain occur in people with anemias. An amino acid in the second zinc finger of EKLF is mutated in patients with CBA and in the Nan (Neonatal anemia) mouse. In collaboration with Jim Bieker we are trying to determine if the anemic phenotype is based on different DNA binding affinities of Nan-EKLF relative to wild-type for some EKLF target genes.