Department of Molecular Genetics, The Ohio State University, Columbus, OH 43210, USA.
J Cell Sci. 2013 Mar 1;126(Pt 5):1218-26. doi: 10.1242/jcs.118521. Epub 2013 Jan 23.
Cells of the budding yeast Saccharomyces cerevisiae select a site for polarized growth in a specific pattern that depends on their cell type. Haploid a and α cells bud in the axial budding pattern, which requires assembly of a landmark that includes the Bud4 protein. To understand how an axial bud site is established, we performed a structure-function analysis of Bud4. Bud4 contains DUF1709 (domain of unknown function), which is similar to a part of the anillin-homology domain, and a putative Pleckstrin homology (PH) domain near to its C terminus. Although its localization depends on septins, a conserved family of GTP-binding proteins, Bud4 is necessary for the stable inheritance of septin rings during cell division. Although some anillins interact directly with septins, we find that neither DUF1709 nor the PH domain is necessary for targeting Bud4 to the mother-bud neck. Instead, this C-terminal region is crucial for association of Bud4 with Bud3 and other components of the axial landmark. Remarkably, septins colocalize with Bud4 mutant proteins that lack these C-terminal domains, forming an arc or a single ring instead of a double ring during and after cytokinesis. Interestingly, overexpression of Bud4 also induces formation of extra Bud4 rings and arcs that are associated with septins. Analyses of a series of bud4 truncation mutants suggest that at least two domains in the central region play a redundant role in targeting Bud4 to the mother-bud neck and are thus likely to interact with septins. Taken together, these results indicate that Bud4 functions as a platform that links septins to the axial landmark.
出芽酵母酿酒酵母的细胞以特定模式选择极化生长的位点，该模式取决于其细胞类型。 单体 a 和 α 细胞以轴状出芽模式出芽，这需要组装一个地标，其中包括 Bud4 蛋白。 为了了解轴向芽位点是如何建立的，我们对 Bud4 进行了结构功能分析。 Bud4 包含 DUF1709（未知功能域），类似于肌球蛋白同源结构域的一部分，以及靠近其 C 末端的假定的 Pleckstrin 同源（PH）结构域。 尽管它的定位依赖于 septin，一种保守的 GTP 结合蛋白家族，但 Bud4 对于在细胞分裂过程中 septin 环的稳定遗传是必需的。 尽管一些肌球蛋白直接与 septin 相互作用，但我们发现 DUF1709 或 PH 结构域对于将 Bud4 靶向母芽颈都不是必需的。 相反，这个 C 末端区域对于 Bud4 与 Bud3 和轴向地标其他成分的关联是至关重要的。 值得注意的是，在细胞分裂过程中和之后，septin 与缺乏这些 C 末端结构域的 Bud4 突变蛋白共定位，形成一个弧形或单个环，而不是双环。 有趣的是，Bud4 的过表达也会诱导额外的 Bud4 环和弧形的形成，这些环和弧形与 septin 相关。 对一系列 bud4 截断突变体的分析表明，中央区域的至少两个结构域在将 Bud4 靶向母芽颈方面发挥冗余作用，因此可能与 septin 相互作用。 总之，这些结果表明 Bud4 作为一个平台，将 septin 与轴向地标连接起来。
