Enforced expression of CUL-4A interferes with granulocytic differentiation and exit from the cell cycle

The cullin family of proteins is involved in the ubiquitin-mediated degradation of cell cycle regulators. Relatively little is known about the function of the CUL-4A cullin, but its overexpression in breast cancer suggests CUL-4A might also regulate the cell cycle. In addition, since other cullins are required for normal development, we hypothesized that CUL-4A is involved in regulating cell cycle progression during differentiation. We observed that CUL-4A mRNA and protein levels decline 2.5-fold during the differentiation of PLB-985 myeloid cells into granulocytes. To examine the significance of this observation, we overexpressed CUL-4A in these cells and found that modest (< 2-fold), enforced expression of CUL-4A attenuates terminal granulocytic differentiation and instead promotes proliferation. This overexpression similarly affects the differentiation of these cells into macrophages. We recently reported that nearly one half of CUL-4A+/- mice are nonviable, and in this report, we show that the viable heterozygous mice, which have reduced CUL-4A expression, have dramatically fewer erythroid and multipotential progenitors than normal controls. Together these results indicate that appropriate CUL-4A expression is essential for embryonic development and for cell cycle regulation during granulocytic differentiation and suggest this gene plays a broader role in hematopoiesis. Since enforced CUL-4A expression does not alter the cell cycle distribution of uninduced cells but dramatically increases the proportion of induced cells that remains in S-phase and reduces the proportion that accumulates in G0/G1, our results show that this CUL-4A regulatory function is interconnected with differentiation, a novel finding for mammalian cullins.