Roncero C, Valdivieso M H, Ribas J C, Durán A
Instituto de Microbiología Bioquímica, Facultad de Biología, Universidad de Salamanca, Spain.
J Bacteriol. 1988 Apr;170(4):1950-4. doi: 10.1128/jb.170.4.1950-1954.1988.
Calcofluor is a fluorochrome that exhibits antifungal activity and a high affinity for yeast cell wall chitin. We isolated Saccharomyces cerevisiae mutants resistant to Calcofluor. The resistance segregated in a Mendelian fashion and behaved as a recessive character in all the mutants analyzed. Five loci were defined by complementation analysis. The abnormally thick septa between mother and daughter cells caused by Calcofluor in wild-type cells were absent in the mutants. The Calcofluor-binding capacity, observed by fluorescence microscopy, in a S. cerevisiae wild-type cells during alpha-factor treatment was also absent in some mutants and reduced in others. Staining of cell walls with wheat germ agglutinin-fluorescein complex indicated that the chitin uniformly distributed over the whole cell wall in vegetative or in alpha-factor-treated cells was almost absent in three of the mutants and reduced in the two others. Cell wall analysis evidenced a five- to ninefold reduction in the amount of chitin in mutants compared with that in the wild-type strain. The total amounts of cell wall mannan and beta-glucan in wild-type and mutant strains were similar; however, the percentage of beta-glucan that remained insoluble after alkali extraction was considerably reduced in mutant cells. The susceptibilities of the mutants and the wild-type strains to a cell wall enzymic lytic complex were rather similar. The in vitro levels of chitin synthase 2 detected in all mutants were similar to that in the wild type. The significance of these results is discussed in connection with the mechanism of chitin synthesis and cell wall morphogenesis in S. cerevisiae.
荧光增白剂是一种具有抗真菌活性且对酵母细胞壁几丁质有高亲和力的荧光染料。我们分离出了对荧光增白剂具有抗性的酿酒酵母突变体。这种抗性以孟德尔方式分离，并且在所有分析的突变体中表现为隐性性状。通过互补分析确定了五个基因座。在野生型细胞中由荧光增白剂引起的母细胞与子细胞之间异常增厚的隔膜在突变体中不存在。通过荧光显微镜观察，在α因子处理期间酿酒酵母野生型细胞中的荧光增白剂结合能力在一些突变体中也不存在，而在其他突变体中则降低。用小麦胚凝集素 - 荧光素复合物对细胞壁进行染色表明，在三个突变体中，营养细胞或α因子处理细胞中均匀分布在整个细胞壁上的几丁质几乎不存在，在另外两个突变体中则减少。细胞壁分析表明，与野生型菌株相比，突变体中几丁质的含量减少了五到九倍。野生型和突变体菌株中细胞壁甘露聚糖和β - 葡聚糖的总量相似；然而，碱提取后仍不溶的β - 葡聚糖的百分比在突变体细胞中显著降低。突变体和野生型菌株对细胞壁酶解复合物的敏感性相当相似。在所有突变体中检测到的几丁质合酶2的体外水平与野生型相似。结合酿酒酵母中几丁质合成和细胞壁形态发生的机制对这些结果的意义进行了讨论。
