Kihm A J, Hershey J C, Haystead T A, Madsen C S, Owens G K
Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, VA 22908-0011, USA.
Proc Natl Acad Sci U S A. 1998 Dec 8;95(25):14816-20. doi: 10.1073/pnas.95.25.14816.
rRNA synthesis by RNA polymerase I requires both the promoter selectivity factor 1, which is composed of TATA binding protein (TBP) and three TBP-associated factors, and the activator upstream binding factor (UBF). Whereas there is strong evidence implicating a role for phosphorylation of UBF in the control of growth-induced increases in rRNA transcription, the mechanism of this effect is not known. Results of immunoprecipitation studies with TBP antibodies showed increased recovery of phosphorylated UBF from growth-stimulated smooth muscle cells. Moreover, using an immobilized protein-binding assay, we found that phosphorylation of UBF in vivo in response to stimulation with different growth factors or in vitro with smooth muscle cell nuclear extract increased its binding to TBP. Finally, we demonstrated that UBF-TBP binding depended on the C-terminal 'acidic tail' of UBF that was hyperphosphorylated at multiple serine sites after growth factor stimulation. Results of these studies suggest that phosphorylation of UBF and subsequent binding to TBP represent a key regulatory step in control of growth-induced increases in rRNA synthesis.
RNA聚合酶I合成rRNA需要启动子选择性因子1（由TATA结合蛋白（TBP）和三个TBP相关因子组成）以及激活因子上游结合因子（UBF）。虽然有强有力的证据表明UBF磷酸化在生长诱导的rRNA转录增加的调控中起作用，但其作用机制尚不清楚。用TBP抗体进行免疫沉淀研究的结果显示，从生长刺激的平滑肌细胞中回收的磷酸化UBF增加。此外，使用固定化蛋白结合试验，我们发现，体内响应不同生长因子刺激或体外使用平滑肌细胞核提取物时UBF的磷酸化增加了其与TBP的结合。最后，我们证明UBF-TBP结合依赖于UBF的C末端“酸性尾巴”，该尾巴在生长因子刺激后在多个丝氨酸位点发生高度磷酸化。这些研究结果表明，UBF的磷酸化以及随后与TBP的结合代表了控制生长诱导的rRNA合成增加的关键调节步骤。
