Dai Fangyan, Duan Xueyan, Liang Yao-Yun, Lin Xia, Feng Xin-Hua
Department of Molecular & Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
Methods Mol Biol. 2010;647:125-37. doi: 10.1007/978-1-60761-738-9_7.
In eukaryotes, regulation of signaling mediators/effectors in the nucleus is one of the principal mechanisms that govern duration and strength of signaling. Smads are a family of structurally related intracellular proteins that serve as signaling effectors for transforming growth factor beta (TGF-beta) and TGF-beta-related proteins. Accumulating evidence demonstrates that Smads possess intrinsic nucleocytoplasmic shuttling capacity, which enables them to transmit TGF-beta signals from cell membrane to nucleus. We recently identified two important steps in the termination of nuclear Smad signaling. The first step is initiated by a serine/threonine phosphatase PPM1A that dephosphorylates Smad2/3 in the nucleus, thereby shutting down signaling capacity of phosphorylated Smad2/3. The second step involves nuclear export of dephosphorylated Smad2/3 with the aid of nuclear protein RanBP3 to terminate Smad signaling. This chapter introduces methods for examining nuclear export of Smad2/3 in TGF-beta signaling.
在真核生物中，细胞核内信号转导介质/效应器的调控是控制信号持续时间和强度的主要机制之一。Smad蛋白是一族结构相关的细胞内蛋白，作为转化生长因子β（TGF-β）及TGF-β相关蛋白的信号效应器。越来越多的证据表明，Smad蛋白具有固有的核质穿梭能力，使其能够将TGF-β信号从细胞膜传递至细胞核。我们最近确定了核内Smad信号终止的两个重要步骤。第一步由丝氨酸/苏氨酸磷酸酶PPM1A启动，它使细胞核内的Smad2/3去磷酸化，从而关闭磷酸化Smad2/3的信号传导能力。第二步涉及在核蛋白RanBP3的帮助下，将去磷酸化的Smad2/3输出细胞核，以终止Smad信号。本章介绍检测TGF-β信号中Smad2/3核输出的方法。
