Rashid Sajid, Pilecka Iwona, Torun Anna, Olchowik Marta, Bielinska Beata, Miaczynska Marta
Laboratory of Cell Biology, International Institute of Molecular and Cell Biology, Ks. Trojdena 4, 02-109 Warsaw, Poland.
J Biol Chem. 2009 Jul 3;284(27):18115-28. doi: 10.1074/jbc.M109.007237. Epub 2009 May 11.
Canonical Wnt signaling regulates many aspects of cellular physiology and tissue homeostasis during development and in adult organisms. In molecular terms, stimulation by Wnt ligands leads to the stabilization of beta-catenin, its translocation to the nucleus, and stimulation of TCF (T-cell factor)-dependent transcription of target genes. This process is controlled at various stages by a number of regulatory proteins, including transcriptional activators and repressors. Here we demonstrate that the endosomal proteins APPL1 and APPL2 are novel activators of beta-catenin/TCF-mediated transcription. APPL proteins are multifunctional adaptors and effectors of the small GTPase Rab5, which localize to a subpopulation of early endosomes but are also capable of nucleocytoplasmic shuttling. Overexpression of APPL1 or APPL2 protein stimulates the activity of beta-catenin/TCF-dependent reporter construct, whereas silencing of APPL1 reduces it. Both APPL proteins interact directly with Reptin, a transcriptional repressor binding to beta-catenin and HDAC1 (histone deacetylase 1), and this interaction was mapped to the pleckstrin homology domain of APPL1. Moreover, APPL proteins are present in an endogenous complex containing Reptin, beta-catenin, HDAC1, and HDAC2. Overexpression of either APPL protein relieves Reptin-dependent transcriptional repression and correlates with the reduced amounts of HDACs and beta-catenin associated with Reptin as well as with the lower levels of Reptin and HDAC1 on the promoters of beta-catenin target genes. We propose that APPL proteins exert their stimulatory effects on beta-catenin/TCF-dependent transcription by decreasing the activity of a Reptin-containing repressive complex.
经典Wnt信号通路在发育过程和成年生物体中调节细胞生理和组织稳态的许多方面。从分子角度来看，Wnt配体的刺激导致β-连环蛋白的稳定、其向细胞核的转运以及对靶基因的TCF（T细胞因子）依赖性转录的刺激。这一过程在多个阶段受到多种调节蛋白的控制，包括转录激活剂和抑制剂。在这里，我们证明内体蛋白APPL1和APPL2是β-连环蛋白/TCF介导转录的新型激活剂。APPL蛋白是小GTP酶Rab5的多功能衔接子和效应器，它们定位于早期内体的一个亚群，但也能够进行核质穿梭。APPL1或APPL2蛋白的过表达刺激β-连环蛋白/TCF依赖性报告基因构建体的活性，而APPL1的沉默则降低其活性。两种APPL蛋白都直接与Reptin相互作用，Reptin是一种与β-连环蛋白和HDAC1（组蛋白去乙酰化酶1）结合的转录抑制剂，这种相互作用被定位到APPL1的普列克底物蛋白同源结构域。此外，APPL蛋白存在于一个包含Reptin、β-连环蛋白、HDAC1和HDAC2的内源性复合物中。任一APPL蛋白的过表达都能减轻Reptin依赖性转录抑制，并与与Reptin相关的HDACs和β-连环蛋白数量减少以及β-连环蛋白靶基因启动子上Reptin和HDAC1水平降低相关。我们提出，APPL蛋白通过降低含Reptin的抑制复合物的活性，对β-连环蛋白/TCF依赖性转录发挥刺激作用。
