The WW domain protein PRO40 is required for fungal fertility and associates with woronin bodies
The WW domain protein PRO40 is required for fungal fertility and associates with woronin bodies
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DOI：
10.1128/ec.00269-06
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发表时间：
2007-05-01
期刊：
影响因子：
--
通讯作者：
Kueck, Ulrich
中科院分区：
文献类型：
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作者：
Engh, Ines;Wuertz, Christian;Kueck, Ulrich
子囊菌的子实体形成是一个受多基因控制的高度复杂的过程，是真菌有性生命周期的重要组成部分。然而，调控这一细胞过程的分子决定因素在很大程度上是未知的。在这里，我们发现不育的Pro40突变体在一个120 kDa的WW结构域蛋白中存在缺陷，该蛋白在同源子囊菌子实体成熟过程中起着关键作用。虽然WW结构域存在于许多真核蛋白中，但PR040的同源物只存在于丝状子囊菌中。利用野生型Pro40基因的全长或截短版本与不同的Pro40突变株进行互补分析，发现PRO40的C末端对恢复可育表型至关重要。利用差速离心法和蛋白酶保护实验，我们确定了PRO40-FLAG融合蛋白位于细胞器内。进一步对融合蛋白与DsRed或绿色荧光蛋白多肽的显微镜研究表明，PRO40与虫体特异性蛋白HEX-1共定位。然而，荧光显微镜、沉淀法和免疫印迹分析显示，大孢链霉菌和粗糙脉孢菌突变株的虫体完整性没有受到影响。我们讨论了PRO40在子实体形成中的作用。
Fruiting body formation in ascomycetes is a highly complex process that is under polygenic control and is a fundamental part of the fungal sexual life cycle. However, the molecular determinants regulating this cellular process are largely unknown. Here we show that the sterile pro40 mutant is defective in a 120-kDa WW domain protein that plays a pivotal role in fruiting body maturation of the homothallic ascomycete Sordaria macrospora. Although WW domains occur in many eukaryotic proteins, homologs of PR040 are present only in filamentous ascomycetes. Complementation analysis with different pro40 mutant strains, using full-sized or truncated versions of the wild-type pro40 gene, revealed that the C terminus of PRO40 is crucial for restoring the fertile phenotype. Using differential centrifugation and protease protection assays, we determined that a PRO40-FLAG fusion protein is located within organelles. Further microscopic investigations of fusion proteins with DsRed or green fluorescent protein polypeptides showed a colocalization of PRO40 with HEX-1, a Woronin body-specific protein. However, the integrity of Woronin bodies is not affected in mutant strains of S. macrospora and Neurospora crassa, as shown by fluorescence microscopy, sedimentation, and immunoblot analyses. We discuss the function of PRO40 in fruiting body formation.