The goal of these studies is to examine the molecular properties of three genes we have cloned, BP1, BP2, and HMG-1(Y), which data from our laboratory and others suggest are repressors of the b-globin gene. Data presented in this proposal points to the potential importance of BP1 in hematopoiesis: the expression of BP1 is restricted to hematopoietic cells after birth, and sequencing has revealed that BP1 contains a homeobox gene, placing it in a class of genes central to development. BP2 is ubiquitously expressed, suggesting a more general role in globin regulation. The investigators have also cloned a CDNA encodng HMG-1(Y), a known protein which we demonstrate binds to and bends Silencer 1 DNA, making it a strong candidate for involvement in repression of the b-globin gene. They will define the role(s) of BP1, BP2 and HMG-1(Y) in normal erythroid development and determine their potential to ameliorate the complications seen in sickle cell anemia (SCA). Benefits of reduction of bs globin expression by repressors may be indirect, since repression of b-globin may lead to reciprocal activation of g -globin. In addition repression of the endogenous bs globin gene would reduce the amount of HbS, favoring the formation of Hb F or Hb A after introduction of a normal g -globin gene or b-globin gene, respectively. BP1 and/or BP2 may directly activate g -globin gene expression, as well: both transcription factors bind upstream of g -globin genes, and there is a direct correlation of reactivation of fetal genes and increased binding activity of BP1 in patients. The ability of these genes to repress b-globin will be measured in cultured cells and in erythroid progenitors grown in a liquid culture system. Expression of BP1, BP2 and HMG-1(Y) will be decreased in erythroid progenitors from cord blood to determine whether this will cause a switch from fetal to adult hemoglobin synthesis. To further examine the functions of BP1 and BP2, transgenic mice will be created overexpressing BP1 and BP2 in their erythroid cells. The proposed studies will firmly establish the role of cloned BP1, BP2 and HMG-1(Y) CDNAS in the repression of the b-globin gene and will determine whether these genes may be useful in the therapy of sickle cell anemia.
