Takeshita Norio, Wernet Valentin, Tsuizaki Makusu, Grün Nathalie, Hoshi Hiro-Omi, Ohta Akinori, Fischer Reinhard, Horiuchi Hiroyuki
Department of Microbiology, Institute for Applied Biosciences, Karlsruhe Institute of Technology (KIT), Karlsruhe, Germany; Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan.
Department of Microbiology, Institute for Applied Biosciences, Karlsruhe Institute of Technology (KIT), Karlsruhe, Germany.
PLoS One. 2015 May 8;10(5):e0125937. doi: 10.1371/journal.pone.0125937. eCollection 2015.
Cell wall formation and maintenance are crucial for hyphal morphogenesis. In many filamentous fungi, chitin is one of the main structural components of the cell wall. Aspergillus nidulans ChsB, a chitin synthase, and CsmA, a chitin synthase with a myosin motor-like domain (MMD) at its N-terminus, both localize predominantly at the hyphal tip regions and at forming septa. ChsB and CsmA play crucial roles in polarized hyphal growth in A. nidulans. In this study, we investigated the mechanism by which CsmA and ChsB accumulate at the hyphal tip in living hyphae. Deletion of kinA, a gene encoding conventional kinesin (kinesin-1), impaired the localization of GFP-CsmA and GFP-ChsB at the hyphal tips. The transport frequency of GFP-CsmA and GFP-ChsB in both anterograde and retrograde direction appeared lower in the kinA-deletion strain compared to wild type, although the velocities of the movements were comparable. Co-localization of GFP-ChsB and GFP-CsmA with mRFP1-KinArigor, a KinA mutant that binds to microtubules but does not move along them, was observed in the posterior of the hyphal tip regions. KinA co-immunoprecipitated with ChsB and CsmA. Co-localization and association of CsmA with KinA did not depend on the MMD. These findings indicate that ChsB and CsmA are transported along microtubules to the subapical region by KinA.
细胞壁的形成和维持对于菌丝形态发生至关重要。在许多丝状真菌中，几丁质是细胞壁的主要结构成分之一。构巢曲霉的几丁质合酶ChsB以及在其N端具有类肌球蛋白运动结构域（MMD）的几丁质合酶CsmA，都主要定位于菌丝顶端区域和正在形成的隔膜处。ChsB和CsmA在构巢曲霉的菌丝极性生长中发挥着关键作用。在本研究中，我们探究了CsmA和ChsB在活菌丝中积累于菌丝顶端的机制。编码传统驱动蛋白（驱动蛋白-1）的基因kinA的缺失，损害了GFP-CsmA和GFP-ChsB在菌丝顶端的定位。与野生型相比，kinA缺失菌株中GFP-CsmA和GFP-ChsB在顺行和逆行方向上的运输频率均较低，尽管运动速度相当。在菌丝顶端区域的后部观察到GFP-ChsB和GFP-CsmA与mRFP1-KinArigor（一种与微管结合但不沿其移动的KinA突变体）共定位。KinA与ChsB和CsmA进行了共免疫沉淀。CsmA与KinA的共定位和结合不依赖于MMD。这些发现表明，ChsB和CsmA通过KinA沿着微管运输到亚顶端区域。
