Peritoneal B-1 cells constitute a unique B cell subset, distinguished by numerous phenotypic and functionalcharacteristics. B-1 cells are an important subset to study because they are responsible for the majority ofnon-immune serum immunoglobulin (Ig), which provides serological protection against a range ofmicroorganisms prior to adaptive immunity. B-1 cells have been implicated in the pathogenesis ofautoimmunity and malignant transformation. Notably, B-1 cells represent the cell of origin for human chroniclymphocytic leukemia (CLL). In adult animals B-1 cells are self-renewing, whereas conventional splenic B(B-2) cells are not. This suggests that the regulation of G0/GI-S phase progression differs between B-1 andB-2 cells. We have reported that dramatic functional differences exist between B-1 and B-2 cells in thesignals required for S-phase entry. B-1 cells fail to proliferate to anti-lg stimulation, which drives B-2 cells intoS phase. Conversely, B-1 cells proliferate in response to phorbol ester (PMA) and do so unusually rapidly,whereas B-2 cell proliferation requires PMA plus calcium ionophore. We have studied the cell cycle responseto PMA as a means to understand the regulation of G0/G1-S phase progression in B-1 and, more generally,all B cells. Our work has led to several key findings: (i) cyclin D3 is uniquely positioned to mediate S-phaseentry in PMA stimulated B-1 cells; (ii) PMA induces cyclin D3-cdk4 complex assembly in B-1 and B-2 cells,but complexes in the latter are inactive. Similarly,IL-4induces the assembly of inactive cyclin D3-cdk4complexes in B-2 cells; and (iii) cdk4 exhibits a distinct phosphorylation profile coincident with inhibition of itskinase activity in PMA-stimulated B-2 cells. The long term objective of this project is to understand theregulation and function of cyclin D3 (and cyclinD3-cdk4 complexes) in B-1 and B-2 cells as an entry point fordissecting important elements that govern restriction point progression and commitment to S-phase entry ofall B cells. This will be accomplished through three specific aims. 1. Elucidate the biological role of cyclin D3in peritoneal B-1 cells; 2. Map the phosphorylation sites on cdk4 in B-2 and B-1 cells; and 3. Elucidate thebiological role and regulation of individual phosphoacceptor sites in cdk4. The information developed fromthis project will elucidate novel principles that govern the regulation of G0/GI-to-S progression in B cells andwill be of general applicability to the regulation of cell cycle progression in all mammalian cell types.
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