Cabib Enrico, Blanco Noelia, Grau Cecilia, Rodríguez-Peña José Manuel, Arroyo Javier
National Institute of Diabetes and Digestive and Kidney Diseases, Laboratory of Biochemistry and Genetics, Bethesda, MD 20892, USA.
Mol Microbiol. 2007 Feb;63(3):921-35. doi: 10.1111/j.1365-2958.2006.05565.x.
In budding yeast, chitin is found in three locations: at the primary septum, largely in free form, at the mother-bud neck, partially linked to beta(1-3)glucan, and in the lateral wall, attached in part to beta(1-6)glucan. By using a recently developed strategy for the study of cell wall cross-links, we have found that chitin linked to beta(1-6)glucan is diminished in mutants of the CRH1 or the CRH2/UTR2 gene and completely absent in a double mutant. This indicates that Crh1p and Crh2p, homologues of glycosyltransferases, ferry chitin chains from chitin synthase III to beta(1-6)glucan. Deletion of CRH1 and/or CRH2 aggravated the defects of fks1Delta and gas1Delta mutants, which are impaired in cell wall synthesis. A temperature shift from 30 degrees C to 38 degrees C increased the proportion of chitin attached to beta(1-6)glucan. The expression of CRH1, but not that of CRH2, was also higher at 38 degrees C in a manner dependent on the cell integrity pathway. Furthermore, the localization of both Crh1p and Crh2p at the cell cortex, the area where the chitin-beta(1-6)glucan complex is found, was greatly enhanced at 38 degrees C. Crh1p and Crh2p are the first proteins directly implicated in the formation of cross-links between cell wall components in fungi.
在出芽酵母中，几丁质存在于三个位置：在初级隔膜处，主要以游离形式存在；在母细胞与芽的颈部，部分与β(1-3)葡聚糖相连；在侧壁，部分与β(1-6)葡聚糖相连。通过使用最近开发的研究细胞壁交联的策略，我们发现与β(1-6)葡聚糖相连的几丁质在CRH1或CRH2/UTR2基因的突变体中减少，而在双突变体中完全缺失。这表明糖基转移酶的同源物Crh1p和Crh2p将几丁质链从几丁质合酶III转运至β(1-6)葡聚糖。CRH1和/或CRH2的缺失加剧了fks1Delta和gas1Delta突变体的缺陷，这些突变体在细胞壁合成方面存在缺陷。温度从30℃转变至38℃会增加与β(1-6)葡聚糖相连的几丁质的比例。CRH1的表达在38℃时也更高，但CRH2的表达并非如此，其表达方式依赖于细胞完整性途径。此外，在38℃时，Crh1p和Crh2p在细胞皮层（即发现几丁质-β(1-6)葡聚糖复合物的区域）的定位大大增强。Crh1p和Crh2p是首批直接参与真菌细胞壁成分之间交联形成的蛋白质。
