SPI-B activates transcription via a unique proline, serine, and threonine domain and exhibits DNA binding affinity differences from PU.1

Abstract

SPI-B is a B lymphocyte-specific Ets transcription factor that shares a high degree of similarity with PU.1/SPI-1. In direct contrast to PU.1(-/-) mice that die in utero and lack monocytes, neutrophils, B cells, and T cells, Spi-B(-/-) mice are viable and exhibit a severe B cell proliferation defect. Since PU.1 is expressed at wild type levels in Spi-B(-/-) B cells, the mutant mice provide genetic evidence that SPI-B and PU.1 have at least some non- redundant roles in B lymphocytes. To begin to understand the molecular basis for these defects, we delineated functional domains of SPI-B for comparison to those of PU.1. By using a heterologous co-transfection system, we identified two independent transactivation domains in the N terminus of SPI- B. Interestingly, only one of these domains (amino acids 31-61), a proline/serine/threonine-rich region, unique among Ets proteins, is necessary for transactivation of the immunoglobulin λ light chain enhancer. This transactivation motif is in marked contrast to PU.1, which contains acidic and glutamine-rich domains. In addition, we describe a functional PU.1 site within the c-FES promoter which SPI-B fails to bind efficiently and transactivate. Finally, we show that SPI-B interacts with the PU.1 cofactors Pip, TBP, c-Jun and with lower affinity to nuclear factor interleukin-6β and retinoblastoma. Taken together, these data suggest that SPI-B binds DNA with a different affinity for certain sites than PU.1 and harbors different transactivation domains. We conclude that SPI-B may activate unique target genes in B lymphocytes and interact with unique, although currently unidentified, cofactors.