RAM: A conserved signaling network that regulates Ace2p transcriptional activity and polarized morphogenesis
RAM: A conserved signaling network that regulates Ace2p transcriptional activity and polarized morphogenesis
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DOI：
10.1091/mbc.e03-01-0018
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发表时间：
2003-09-01
影响因子：
3.3
通讯作者：
Luca, FC
中科院分区：
文献类型：
--
作者：
Nelson, B;Kurischko, C;Luca, FC
在酿酒酵母中，极化形态发生对于芽位选择、芽发育和细胞分离至关重要。后者由Ace 2 p转录因子介导，其控制细胞分离基因的子细胞特异性表达。最近，一组蛋白质，包括Cbk 1 p激酶，其结合伙伴Mob 2 p，Tao 3 p（Pag 1 p）和Hym 1 p被证明可以调节Ace 2 p活性和细胞形态发生。这些蛋白质似乎形成了一个信号网络，我们指定RAM为Ace 2 p活性和细胞形态发生的调节。为了找到额外的RAM组件，我们进行了遗传筛选的双边交配和细胞分离突变体，并确定了PAK相关激酶Kic 1 p的等位基因，除了Cbk 1 p，Mob 2 p，Tao 3 p和Hym 1 p。每个RAM基因的缺失导致Ace 2 p功能的丧失，并引起细胞极性缺陷，这与突变体或极性体突变体不同。双杂交和免疫共沉淀实验揭示了RAM蛋白之间相互作用的复杂网络，除了先前记录的Cbk 1 p-Mob 2 p和Cbk 1 p-Tao 3 p相互作用之外，还包括Cbk 1 p-Cbk 1 p，Cbk 1 p-Kic 1 p，Kic 1 p-Tao 3 p和Kdc 1 p-Hym 1 p相互作用。我们还发现了一种新的富含亮氨酸的重复序列的蛋白质Sog 2 p与Hym 1 p和Kic 1 p相互作用。缺乏Sog 2 p的细胞表现出与RAM信号传导中的扰动相关的特征性细胞分离和细胞形态缺陷。每个RAM蛋白定位于皮层网站的增长过程中的萌芽和交配信息素的反应。Hym 1 p是Kic 1 p和Sog 2 p依赖和Sog 2 p和Kic 1 p是相互依赖的本地化，表明这些蛋白质之间的密切功能关系。在有丝分裂末期，在子细胞核中仅检测到Mob 2 p和Cbk 1 p。Mob 2 p-Cbk 1 p复合物的核定位和激酶活性依赖于所有其他RAM蛋白，这表明Mob 2 p-Cbk 1 p在RAM网络中的功能较晚。我们的数据表明，RAM信号的功能结构类似于S。酿酒酵母有丝分裂出口网络和粟酒裂殖酵母分隔起始网络，并且可能在真核生物中保守。
In Saccharomyces cerevisiae, polarized morphogenesis is critical for bud site selection, bud development, and cell separation. The latter is mediated by Ace2p transcription factor, which controls the daughter cell-specific expression of cell separation genes. Recently, a set of proteins that include Cbk1p kinase, its binding partner Mob2p, Tao3p (Pag1p), and Hym1p were shown to regulate both Ace2p activity and cellular morphogenesis. These proteins seem to form a signaling network, which we designate RAM for regulation of Ace2p activity and cellular morphogenesis. To find additional RAM components, we conducted genetic screens for bilateral mating and cell separation mutants and identified alleles of the PAK-related kinase Kic1p in addition to Cbk1p, Mob2p, Tao3p, and Hym1p. Deletion of each RAM gene resulted in a loss of Ace2p function and caused cell polarity defects that were distinct from formin or polarisome mutants. Two-hybrid and coimmunoprecipitation experiments reveal a complex network of interactions among the RAM proteins, including Cbk1p-Cbk1p, Cbk1p-Kic1p, Kic1p-Tao3p, and Kdc1p-Hym1p interactions, in addition to the previously documented Cbk1p-Mob2p and Cbk1p-Tao3p interactions. We also identified a novel leucine-rich repeat-containing protein Sog2p that interacts with Hym1p and Kic1p. Cells lacking Sog2p exhibited the characteristic cell separation and cell morphology defects associated with perturbation in RAM signaling. Each RAM protein localized to cortical sites of growth during both budding and mating pheromone response. Hym1p was Kic1p- and Sog2p-dependent and Sog2p and Kic1p were interdependent for localization, indicating a close functional relationship between these proteins. Only Mob2p and Cbk1p were detectable in the daughter cell nucleus at the end of mitosis. The nuclear localization and kinase activity of the Mob2p-Cbk1p complex were dependent on all other RAM proteins, suggesting that Mob2p-Cbk1p functions late in the RAM network. Our data suggest that the functional architecture of RAM signaling is similar to the S. cerevisiae mitotic exit network and Schizosaccharomyces pombe septation initiation network and is likely conserved among eukaryotes.