Abstract
The B cell-specific expression of immunoglobulin (Ig) genes is controlled by the concerted action of variable (V) region promoters and intronic or 3' enhancers, all of which are active in a lymphoid-specific manner. A crucial highly conserved element of the V region promoters is the octamer site -ATTTGCAT-, which can be bound by ubiquitous (Oct-1) as well as B cell-specific (Oct-2) factors. Another less conserved element found in many Ig promoters is pyrimidine-rich and has been shown to be functionally important, in particular for those Ig promoters that have only an imperfect octamer site. In this study we have analyzed the factors binding specifically to the pyrimidine-rich motif of the V kappa 19 promoter, a light chain gene promoter with an imperfect octamer site. Using nuclear extracts prepared from B cells, we detected two sets of specific complexes in electrophoretic mobility shift experiments. One complex appears to be ubiquitous but enriched in lymphoid cells and represents the binding of a potentially novel factor with an apparent molecular mass of approximately 50 kDa. The other complex was found only with extracts from pre-B or B cells as well as from a macrophage cell line and appears to be caused by the binding of PU.1, a factor of the Ets family. We show that on this Ig promoter Oct factors (Oct-1 or Oct-2) and PU.1 can bind concomitantly but without synergism. By transfection experiments in non-B cells we demonstrate that PU.1 is indeed able to activate this promoter in concert with Oct-2. Furthermore, we show that PU.1 can bind with varying affinities to the pyrimidine-rich elements of several other Ig promoters. These data suggest a more general role for PU.1 or other members of the Ets family in the activation of Ig promoters.
Highlights
The B cell specificity ofIg promoters is largely caused by the octamer element [9], this same element is implicated in the ubiquitous expression of small nuclear RNA genes [23], the cell cycle regulation of the histone H2B gene [24], and the VPl6-dependent expression of herpesvirus intermediate early genes [25,26,27]
The different cellular distribution of these two proteins led to the proposal that Oct-2 rather than Oct-l is responsible for the B cell specificity of Ig promoters
We show that one ofthese factors binding to that site appears to be PD.l, a member of the Ets family that is expressed in B cells and macrophages [35]
Summary
The B cell specificity ofIg promoters is largely caused by the octamer element [9], this same element is implicated in the ubiquitous expression of small nuclear RNA genes [23], the cell cycle regulation of the histone H2B gene [24], and the VPl6-dependent expression of herpesvirus intermediate early genes [25,26,27]. The ability of the octamer element to promote ubiquitous as well as B cell-specific gene expression was initially suggested to reside in its interaction with Oct-l and Oct-2. Analysis of Oct-2 knockout mice has shown that Oct-2 is not required for the generation of Ig-bearing B cells but is crucial for their maturation to Ig-secreting cells, and this implies that Oct-2 plays an essential role only late in B cell differentiation [29] It appears that both octamer factors can activate transcription from Ig promoters. We show that one ofthese factors binding to that site appears to be PD.l, a member of the Ets family that is expressed in B cells and macrophages [35]. Suggest that PD. as well as perhaps other factors of the Ets family might be involved in the regulation of a number of Ig promoters
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