Huang He, Rastegar Mojgan, Bodner Caroline, Goh Siew-Lee, Rambaldi Isabel, Featherstone Mark
McGill Cancer Centre, McGill University, 3655 Promenade Sir William Osler, Montréal, Québec H3G 1Y6, Canada.
J Biol Chem. 2005 Mar 18;280(11):10119-27. doi: 10.1074/jbc.M413963200. Epub 2005 Jan 15.
MEIS proteins form heteromeric DNA-binding complexes with PBX monomers and PBX.HOX heterodimers. We have shown previously that transcriptional activation by PBX.HOX is augmented by either protein kinase A (PKA) or the histone deacetylase inhibitor trichostatin A (TSA). To examine the contribution of MEIS proteins to this response, we used the chromatin immunoprecipitation assay to show that MEIS1 in addition to PBX1, HOXA1, and HOXB1 was recruited to a known PBX.HOX target, the Hoxb1 autoregulatory element following Hoxb1 transcriptional activation in P19 cells. Subsequent to TSA treatment, MEIS1 recruitment lagged behind that of HOX and PBX partners. MEIS1A also enhanced the transcriptional activation of a reporter construct bearing the Hoxb1 autoregulatory element after treatment with TSA. The MEIS1 homeodomain and protein-protein interaction with PBX contributed to this activity. We further mapped TSA-responsive and CREB-binding protein-dependent PKA-responsive transactivation domains to the MEIS1A and MEIS1B C termini. Fine mutation of the 56-residue MEIS1A C terminus revealed four discrete regions required for transcriptional activation function. All of the mutations impairing the response to TSA likewise reduced activation by PKA, implying a common mechanistic basis. C-terminal deletion of MEIS1 impaired transactivation without disrupting DNA binding or complex formation with HOX and PBX. Despite sequence similarity to MEIS and a shared ability to form heteromeric complexes with PBX and HOX partners, the PREP1 C terminus does not respond to TSA or PKA. Thus, MEIS C termini possess transcriptional regulatory domains that respond to cell signaling and confer functional differences between MEIS and PREP proteins.
MEIS蛋白与PBX单体以及PBX.HOX异二聚体形成异源DNA结合复合物。我们之前已经表明，PBX.HOX的转录激活可被蛋白激酶A（PKA）或组蛋白脱乙酰酶抑制剂曲古抑菌素A（TSA）增强。为了研究MEIS蛋白对这种反应的作用，我们使用染色质免疫沉淀试验表明，在P19细胞中Hoxb1转录激活后，除了PBX1、HOXA1和HOXB1之外，MEIS1也被募集到一个已知的PBX.HOX靶标，即Hoxb1自调控元件。TSA处理后，MEIS1的募集落后于HOX和PBX伙伴。用TSA处理后，MEIS1A也增强了携带Hoxb1自调控元件的报告构建体的转录激活。MEIS1同源结构域以及与PBX的蛋白质-蛋白质相互作用促成了这种活性。我们进一步将TSA反应性和依赖于CREB结合蛋白的PKA反应性反式激活结构域定位到MEIS1A和MEIS1B的C末端。对56个残基的MEIS1A C末端进行精细突变，揭示了转录激活功能所需的四个离散区域。所有损害对TSA反应的突变同样降低了PKA的激活作用，这意味着存在共同的机制基础。MEIS1的C末端缺失损害了反式激活，而不破坏DNA结合或与HOX和PBX的复合物形成。尽管PREP1与MEIS序列相似，并且具有与PBX和HOX伙伴形成异源复合物的共同能力，但其C末端对TSA或PKA无反应。因此，MEIS的C末端具有转录调节结构域，可对细胞信号作出反应，并赋予MEIS和PREP蛋白之间的功能差异。
