Koseoglu M Murat, Graves Lee M, Marzluff William F
Department of Biology, Program in Molecular Biology and Biotechnology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599, USA.
Mol Cell Biol. 2008 Jul;28(14):4469-79. doi: 10.1128/MCB.01416-07. Epub 2008 May 19.
Histone mRNA levels are cell cycle regulated, and a major regulatory mechanism is restriction of stem-loop binding protein (SLBP) to S phase. Degradation of SLBP at the end of S phase results in cessation of histone mRNA biosynthesis, preventing accumulation of histone mRNA until SLBP is synthesized just before entry into the next S phase. Degradation of SLBP requires an SFTTP (58 to 62) and KRKL (95 to 98) sequence, which is a putative cyclin binding site. A fusion protein with the 58-amino-acid sequence of SLBP (amino acids 51 to 108) fused to glutathione S-transferase (GST) is sufficient to mimic SLBP degradation at late S phase. Using GST-SLBP fusion proteins as a substrate, we show that cyclin A/Cdk1 phosphorylates Thr61. Furthermore, knockdown of Cdk1 by RNA interference stabilizes SLBP at the end of S phase. Phosphorylation of Thr61 is necessary for subsequent phosphorylation of Thr60 by CK2 in vitro. Inhibitors of CK2 also prevent degradation of SLBP at the end of S phase. Thus, phosphorylation of Thr61 by cyclin A/Cdk1 primes phosphorylation of Thr60 by CK2 and is responsible for initiating SLBP degradation. We conclude that the increase in cyclin A/Cdk1 activity at the end of S phase triggers degradation of SLBP at S/G(2).
组蛋白mRNA水平受细胞周期调控，一种主要的调控机制是将茎环结合蛋白（SLBP）限制在S期。S期末期SLBP的降解导致组蛋白mRNA生物合成停止，阻止组蛋白mRNA积累，直到在下一个S期进入前SLBP重新合成。SLBP的降解需要一个SFTTP（58至62）和KRKL（95至98）序列，这是一个假定的细胞周期蛋白结合位点。一个将SLBP的58个氨基酸序列（氨基酸51至108）与谷胱甘肽S-转移酶（GST）融合的融合蛋白足以模拟S期末期SLBP的降解。以GST-SLBP融合蛋白作为底物，我们发现细胞周期蛋白A/Cdk1使Thr61磷酸化。此外，通过RNA干扰敲低Cdk1可在S期末期使SLBP稳定。在体外，Thr61的磷酸化是CK2随后使Thr60磷酸化所必需的。CK2抑制剂也可防止S期末期SLBP的降解。因此，细胞周期蛋白A/Cdk1使Thr61磷酸化引发了CK2使Thr60磷酸化，并导致SLBP降解的起始。我们得出结论，S期末期细胞周期蛋白A/Cdk1活性的增加触发了S/G2期SLBP的降解。
