Molecular Analyses of transcriptional factors in filamentous fungi and their applications

以米曲霉为代表的丝状真菌在日本发酵工业中扮演着重要的角色。为了更有效地利用各种生殖资源，特别是淀粉、纤维素和木聚糖等碳水化合物，我们对从米曲霉中分离的Taka-淀粉酶A基因的表达特性进行了研究。在诱导条件下，CCAAT序列及其结合复合体ANCP在促进Taka-AmylaseA基因表达方面发挥了重要作用。此外，分别从根瘤拟青霉和纤细拟青霉中分离出纤维素酶和木聚糖酶基因，并对它们的基因结构进行了研究。在过去两年中，我们取得了以下成果。1.克隆了水稻曲霉CCAAT结合复合体的编码亚基基因Aohap B、C和D，并进行了序列测定。发现AohapC基因与从根结线虫中分离的hapC缺失突变体互补，表明AoHapC与根结线虫HapC在功能上是可以互换的。2.淀粉反应元件SRE及其结合蛋白SREB在Taka-AmylaseA基因的诱导中起重要作用。从多核拟青霉中分离到amyR基因，并对其进行了序列测定。利用Aidulans AmyR缺失突变体，建议SREB与AmyR相同。3.对EglA基因的启动子区域进行了初步分析，以寻找与EglA诱导有关的元件。4.确定了米曲霉木聚糖酶基因xynFI的诱导元件。从米曲霉中分离到诱导木聚糖酶基因的xlnR基因，并进行了序列测定。

Filamentous fungi represented by Aspergillus oryzae have been played important roles in Japanese fermentation industries. We have examined the expression properties of Taka-amylase A gene isolated from A. oryzae in order to utilize efficiently various reproductive natural resources, especially carbohydrates such as starch, cellulose and xylan. CCAAT sequence and its binding complex, AnCP, have been shown to play important roles in enhancement of Taka-amylaseA gene expression under the inducible conditions. Furthermore, cellulase and xylanase genes were isolated from A. nidulans and C. gracile, respectively and their gene structures have been elucidated.In this project, cellulase and xylanase genes have been characterized as to their regulation of gene expression in addition to the Taka-amylase gene. We have obtained the following results during last two years. #1. Genes, Aohap B, Cand D, encoding subunits comprising the A. oryzae CCAAT binding complex were cloned and sequenced. The AohapCgene was found to compliment the hapC deletion mutant isolated from A. nidulans, suggesting AoHapC is functionally interchangeable with A. nidulans HapC. #2.SRE (Starch Responsive Element) and its binding protein, SREB, were suggested to play a vital role in induction of Taka-amylaseA gene. AmyR gene was isolated from A. nidulans and sequenced. Utilizing A. nidulans amyR deletion mutant, SREB was suggested to be the same as AMYR. #3. Preliminary analysis as to the promoter region of the eglA gene was conducted to find any elements involved in induction of eglA. #4. The A. oryzae xylanase gene, xynFI, was determined for its inducible element. The xlnR gene responsible for the induction of xylanase gene was isolated from A. oryzae and sequenced.