| The erythropoietin receptor (EpoR) is required for the proliferation and survival of committed erythroid lineage cells. Previous studies have utilized receptor mutations to show the requirement for the distal half of the cytoplasmic domain of the EpoR and receptor tyrosines for activation of signaling pathways potentially critical to Epo function. To extend these studies to in vivo erythropoiesis, two mutant strains of mice were created. One strain (H) contains a truncation of the distal-half of the cytoplasmic domain while the second strain (HM) contains the same truncation as well as the mutation of the residual tyrosine (Y343) to a phenylalanine. Strikingly, both strains of mice are viable but with alterations in constitutive erythropoiesis or in in vitro assays of red cell lineage function. Challenging H mutant mice with continuous injections of Epo results in an erythrocytosis that is not seen in HM mice. The results demonstrate that neither the distal region nor the remaining tyrosine are essential for in vivo EpoR function but contribute in a subtle manner to receptor function.; To further investigate signal transduction pathways that are dependent on the EpoR, DNA microarray analysis was performed. RNAs isolated from JAK2-/-, EpoR-/-, EpoR-H or EpoR-HM fetal livers were compared to the control RNAs obtained from wild type littermates using either Affymetrix gene chips or Incyte Genomics cDNA microarray chips. In this analysis, genes affected by the H or HM mutations were identified in fetal livers.; Recently, a family of proteins was identified that could downregulate a variety of cytokine receptors. One of the family members, SOCS3, was implicated in the iv downregulation of EpoR and/or JAK2. To gain insights into the SOCS3-mediated inhibitory mechanisms, X-ray crystallography method was utilized to determine the 3-dimensional structure of SOCS3. The purification and crystallization conditions were successfully established throughout the experiments. Diffraction data were collected and analyzed. But processing of the collected reflection data revealed a need for further analysis to build a 2.8 A resolution model of SOCS3. |
