Gomez Alberto, Perez Jaqueline, Reyes Abigail, Duran Angel, Roncero Cesar
Instituto de Microbiología Bioquímica, Departamento de Microbiología y Genética, CSIC/Universidad de Salamanca, Salamanca, Spain.
Eukaryot Cell. 2009 Sep;8(9):1449-59. doi: 10.1128/EC.00153-09. Epub 2009 Jul 24.
In Saccharomyces cerevisiae, the simultaneous absence of Slt2 and Rim101 prevents growth in nonosmotically stabilized media (F. Castrejon et al., Eukaryot. Cell 5:507-517, 2006). The double mutant slt2Delta rim101Delta displays altered chitin rings, together with a significant reduction in the overall levels of chitin. Cultures of this mutant lyse upon transfer to nonosmotically stabilized media, mostly through the bud, and such lysis is partially prevented by deletion of the chitinase gene (CTS1). Growth of the slt2Delta rim101Delta double mutant was restored by the overexpression of the GFA1 or CCT7 genes, which code for two biologically unrelated proteins. Further characterization of the mutant and its suppressors indicated that both Slt2 and Rim101 were independently required for the correct assembly of the septum machinery and that their concomitant absence reduced Chs3 accumulation at the neck, leading to lower levels of chitin. GFA1 overexpression, as well as the addition of glucosamine to the growth medium, specifically suppressed the growth defects by activating chitin synthesis at the neck and restoring the normal assembly of the chitin ring. In contrast, overexpression of CCT7, a Cct chaperonin subunit, alleviated the defect in the septum machinery without affecting chitin synthesis. Both suppressors thus act by reducing neck fragility through different mechanisms and allow growth in nonstabilized media. This work reports new roles for Slt2 and Rim101 in septum formation in budding yeast and confirms the homeostatic role of the chitin ring in the maintenance of neck integrity during cell division.
在酿酒酵母中，同时缺失Slt2和Rim101会阻碍其在非渗透稳定培养基中的生长（F. Castrejon等人，《真核细胞》5:507 - 517，2006年）。双突变体slt2Δrim101Δ显示出几丁质环改变，同时几丁质的总体水平显著降低。将该突变体的培养物转移到非渗透稳定培养基后会裂解，主要通过芽部，而几丁质酶基因（CTS1）的缺失可部分防止这种裂解。GFA1或CCT7基因的过表达恢复了slt2Δrim101Δ双突变体的生长，这两个基因编码两种生物学上不相关的蛋白质。对该突变体及其抑制子的进一步表征表明，Slt2和Rim101对于隔膜机制的正确组装都是独立必需的，它们同时缺失会减少Chs3在颈部的积累，导致几丁质水平降低。GFA1的过表达以及在生长培养基中添加葡糖胺，通过激活颈部的几丁质合成并恢复几丁质环的正常组装，特异性地抑制了生长缺陷。相比之下，Cct伴侣蛋白亚基CCT7的过表达减轻了隔膜机制的缺陷，而不影响几丁质合成。因此，这两种抑制子都通过不同机制降低颈部脆弱性，从而使细胞能够在非稳定培养基中生长。这项工作报道了Slt2和Rim101在芽殖酵母隔膜形成中的新作用，并证实了几丁质环在细胞分裂过程中维持颈部完整性的稳态作用。
