Abstract
A key feature of chronic lymphocytic leukaemia (CLL) cells is overexpressed protein kinase CβII (PKCβII), an S/T kinase important in the pathogenesis of this and other B cell malignancies. The mechanisms contributing to enhanced transcription of the gene coding for PKCβII, PRKCB, in CLL cells remain poorly described, but could be important because of potential insight into how the phenotype of these cells is regulated. Here, we show that SP1 is the major driver of PKCβII expression in CLL cells where enhanced association of this transcription factor with the PRKCB promoter is likely because of the presence of histone marks permissive of gene activation. We also show how vascular endothelial growth factor (VEGF) regulates PRKCB promoter function in CLL cells, stimulating PKCβ gene transcription via increased association of SP1 and decreased association of STAT3. Taken together, these results are the first to demonstrate a clear role for SP1 in the up regulation of PKCβII expression in CLL cells, and the first to link SP1 with the pathogenesis of this and potentially other B cell malignancies where PKCβII is overexpressed.
Highlights
VEGF-induced stimulation of protein kinase CβII (PKCβII) activity[24]
The results show mean ±SE of n = 3 separate experiments. (b) Effect of 200 nM mithramycin on PKCβII mRNA levels in Chronic lymphocytic leukaemia (CLL) cells taken from 5 patients. (c) Effect of SP1 siRNA compared to negative control siRNA (Neg) on primary CLL cells with respect to SP1 mRNA. (d) Effect of SP1 siRNA compared to negative control siRNA (Neg) on primary CLL cells with respect to PKCβmRNA. (e) Western blot showing the effect of SP1 siRNA and negative control siRNA (Neg) on primary CLL cells with respect to SP1 and PKCβII protein levels (n = 1 experiment)
The results we present are the first to demonstrate a clear role for SP1 in the regulation of PKCβII expression in CLL cells, and we suggest that access of SP1 to the promoter of PRKCB likely results both from a chromosome landscape that is permissive of gene transcription and from VEGF-mediated inhibition of the suppressive effects of STAT3
Summary
VEGF-induced stimulation of PKCβII activity[24] This mechanism is reportedly used in other cell systems[25,26], and may be of particular importance to the pathogenesis of CLL because of the high levels of this cytokine present within tissues where expansion of the malignant clone takes place[27,28]. Treatment with VEGF causes a decrease in STAT3 binding to the PRKCB promoter and maintains elevated binding of SP1 during in vitro culture. Taken together, these results demonstrate a direct relationship between SP1 binding and PRKCB transcription, and further suggest that this TF is a contributor to the pathobiology of CLL and potentially other malignant cells where PKCβII is overexpressed
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