The overall goal of our project is to understand transcriptional mechanisms that control differentiation in early B lymphocytes. The differentiation of B lymphocytes from committed precursor cells to antibody-secreting plasma cells proceeds through multiple steps defined by changes in the expression pattern of lineage-specific genes. As a paradigm for a temporally regulated B cell-specific gene, the mb-1 (MB- 1; CD79alpha) gene encodes a trans-membrane protein that is required for cell surface display and signaling functions of membrane-bound immunoglobulin (mIg) on early B cells. We hypothesize that the early B cell-specific transcription factor Pax-5 is an important regulator of mb-1 transcription. Recombinant Pax-5 binds specifically to the mb-1 promoter by itself in vitro, and Pax-5 recruits proteins of the Ets protooncogene family to a site that is essential for promoter function in transfected pre-B cells. These observations represent the first demonstration of protein:protein interactions involving a member of the paired domain (Pax) protein family. Moreover, we describe a novel mechanism governing DNA binding by Ets proteins. Therefore, our experiments will address how Pax-5 and Ets proteins regulate mb-1 gene transcription, and potentially, other genes in early B cells (and potentially neuronal cells, which also express these proteins). We will examine the molecular basis for Pax-5:Ets interactions at the protein:protein and protein:DNA levels and determine how different Ets proteins affect the functional properties of Pax-5:Ets complexes. We will test whether the down-regulation of mb-1 gene expression in terminally differentiated plasma cells is due to the absence of early B cell specific factors in these cells (e.g., Pax-5). We will also address the regulation of functional Pax-5 activity in resting vs. activated B cells. In summary, our goal is to understand how these proteins interact to regulate target genes in B lymphocytes and thus, control differentiation. Regulation of the developmentally important mb-1 gene by Pax-5 suggests a mechanism for the absence of mature B cells in genetically altered mice lacking functional pax-5 genes. Ultimately, our study may define a novel mechanism for control of normal and abnormal cell proliferation (including B lymphomas and leukemias), because both Pax-5 and Ets proteins have been implicated as regulators of proliferation in B cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI037574-03
Application #
2882192
Study Section
Allergy and Immunology Study Section (ALY)
Program Officer
Ridge, John P
Project Start
1997-03-01
Project End
2002-02-28
Budget Start
1999-03-01
Budget End
2000-02-29
Support Year
3
Fiscal Year
1999
Total Cost
Indirect Cost
Name
National Jewish Health
Department
Type
DUNS #
City
Denver
State
CO
Country
United States
Zip Code
80206
Fitzsimmons, Daniel; Lukin, Kara; Lutz, Ryan et al. (2009) Highly cooperative recruitment of Ets-1 and release of autoinhibition by Pax5. J Mol Biol 392:452-64
Maier, Holly; Ostraat, Rachel; Parenti, Sarah et al. (2003) Requirements for selective recruitment of Ets proteins and activation of mb-1/Ig-alpha gene transcription by Pax-5 (BSAP). Nucleic Acids Res 31:5483-9
Maier, Holly; Colbert, Jeff; Fitzsimmons, Daniel et al. (2003) Activation of the early B-cell-specific mb-1 (Ig-alpha) gene by Pax-5 is dependent on an unmethylated Ets binding site. Mol Cell Biol 23:1946-60
Maier, Holly; Hagman, James (2002) Roles of EBF and Pax-5 in B lineage commitment and development. Semin Immunol 14:415-22
Sigvardsson, Mikael; Clark, Dawn R; Fitzsimmons, Daniel et al. (2002) Early B-cell factor, E2A, and Pax-5 cooperate to activate the early B cell-specific mb-1 promoter. Mol Cell Biol 22:8539-51
Johnson, A D; Fitzsimmons, D; Hagman, J et al. (2001) EGL-38 Pax regulates the ovo-related gene lin-48 during Caenorhabditis elegans organ development. Development 128:2857-65
Fitzsimmons, D; Lutz, R; Wheat, W et al. (2001) Highly conserved amino acids in Pax and Ets proteins are required for DNA binding and ternary complex assembly. Nucleic Acids Res 29:4154-65
Hagman, J; Wheat, W; Fitzsimmons, D et al. (2000) Pax-5/BSAP: regulator of specific gene expression and differentiation in B lymphocytes. Curr Top Microbiol Immunol 245:169-94
Wheat, W; Fitzsimmons, D; Lennox, H et al. (1999) The highly conserved beta-hairpin of the paired DNA-binding domain is required for assembly of Pax-Ets ternary complexes. Mol Cell Biol 19:2231-41