EFFECT OF PU.1 PHOSPHORYLATION ON INTERACTION WITH NF-EM5 AND TRANSCRIPTIONAL ACTIVATION
EFFECT OF PU.1 PHOSPHORYLATION ON INTERACTION WITH NF-EM5 AND TRANSCRIPTIONAL ACTIVATION
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DOI：
10.1126/science.8456286
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发表时间：
1993-03-12
期刊：
影响因子：
56.9
通讯作者：
ATCHISON, ML
中科院分区：
文献类型：
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作者：
PONGUBALA, JMR;VANBEVEREN, C;ATCHISON, ML
PU.1 募集第二个 B 细胞限制性核因子 NF-EM5 与免疫球蛋白 kappa 3' 增强子中的 DNA 位点结合。 NF-EM5 的 DNA 结合需要与 PU.1 和特定的 DNA 接触进行蛋白质-蛋白质相互作用。去磷酸化的 PU.1 与 DNA 结合，但不与 NF-EM5 相互作用。 PU.1 中丝氨酸到丙氨酸突变的分析表明，丝氨酸 148 (Ser148) 是蛋白质与蛋白质相互作用所必需的。细菌中产生的 PU.1 不与 NF-EM5 相互作用。纯化的酪蛋白激酶 II 将细菌产生的 PU.1 磷酸化，将其修饰为与 NF-EM5 相互作用并招募 NF-EM5 与 DNA 结合的形式。对细菌产生的 PU.1 的磷酸肽分析表明 Ser148 被酪蛋白激酶 II 磷酸化。该位点在体内也被磷酸化。野生型 PU.1 的表达使含有 PU.1 和 NF-EM5 结合位点的报告构建体的表达增加了近六倍，而 Ser148 突变体仅形成弱激活的转录。这些结果表明 PU.1 在 Ser148 处的磷酸化对于与 NF-EM5 的相互作用是必需的，并表明这种磷酸化可以调节转录活性。
PU.1 recruits the binding of a second B cell-restricted nuclear factor, NF-EM5, to a DNA site in the immunoglobulin kappa 3' enhancer. DNA binding by NF-EM5 requires a protein-protein interaction with PU.1 'and specific DNA contacts. Dephosphorylated PU.1 bound to DNA but did not interact with NF-EM5. Analysis of serine-to-alanine mutations in PU.1 indicated that serine 148 (Ser148) is required for protein-protein interaction. PU.1 produced in bacteria did not interact with NF-EM5. Phosphorylation of bacterially produced PU.1 by purified casein kinase II modified it to a form that interacted with NF-EM5 and that recruited NF-EM5 to bind to DNA. Phosphopeptide analysis of bacterially produced PU.1 suggested that Ser148 is phosphorylated by casein kinase II. This site is also phosphorylated in vivo. Expression of wild-type PU.1 increased expression of a reporter construct containing the PU.1 and NF-EM5 binding sites nearly sixfold, whereas the Ser148 mutant form only weakly activated transcription. These results demonstrate that phosphorylation of PU.1 at Ser148 is necessary for interaction with NF-EM5 and suggest that this phosphorylation can regulate transcriptional activity.