Yelton M M, Hamer J E, de Souza E R, Mullaney E J, Timberlake W E
Proc Natl Acad Sci U S A. 1983 Dec;80(24):7576-80. doi: 10.1073/pnas.80.24.7576.
We have cloned the trifunctional trpC gene from Aspergillus nidulans by hybrid phage lambda complementation of an Escherichia coli trpC mutant lacking phosphoribosylanthranilate isomerase activity. Four different phages sharing a 4.3-kilobase region were obtained. Plasmid subclones containing this region also complemented the E. coli trpC mutant. We determined that a 1.8-kilobase DNA fragment was minimally required for complementation. The fragment hybridized with two poly(A)+ RNAs, 3.0 and 3.2 kilobases in length. We infer that these transcripts are Aspergillus trpC mRNAs and that the entire Aspergillus trpC gene is not required for complementation in E. coli. Levels of both trpC transcripts in poly(A)+ RNA are regulated by growth medium composition. They were highest when cells were grown in minimal medium containing nitrate as the nitrogen source and lowest when cells were grown in medium containing yeast extract. The concentrations of the transcripts are also regulated during conidiophore development. Conidiating cultures grown on medium containing yeast extract had significantly higher levels of both transcripts than did hyphae grown in minimal medium containing nitrate. Levels of the transcripts in mature spores were equivalent to those found in hyphae grown in minimal medium containing nitrate. Results from nutritional experiments with an A. nidulans trpC mutant suggest that developmental regulation of trpC mRNA levels may be related to a high requirement for tryptophan or a compound derived from tryptophan during conidiation.
我们通过对缺乏磷酸核糖邻氨基苯甲酸异构酶活性的大肠杆菌trpC突变体进行λ噬菌体杂交互补,从构巢曲霉中克隆了三功能trpC基因。获得了四个共享4.3千碱基区域的不同噬菌体。含有该区域的质粒亚克隆也能互补大肠杆菌trpC突变体。我们确定互补所需的最小DNA片段为1.8千碱基。该片段与两个长度分别为3.0和3.2千碱基的聚腺苷酸加尾(poly(A)+)RNA杂交。我们推断这些转录本是曲霉trpC mRNA,并且在大肠杆菌中互补不需要完整的曲霉trpC基因。聚腺苷酸加尾RNA中两种trpC转录本的水平受生长培养基组成的调节。当细胞在以硝酸盐作为氮源的基本培养基中生长时,它们的水平最高;而当细胞在含有酵母提取物的培养基中生长时,水平最低。在分生孢子梗发育过程中,转录本的浓度也受到调节。在含有酵母提取物的培养基上生长的产分生孢子培养物中,两种转录本的水平均显著高于在以硝酸盐作为氮源的基本培养基中生长的菌丝。成熟孢子中转录本的水平与在以硝酸盐作为氮源的基本培养基中生长的菌丝中的水平相当。对构巢曲霉trpC突变体进行的营养实验结果表明,trpC mRNA水平的发育调节可能与分生孢子形成过程中对色氨酸或色氨酸衍生化合物的高需求有关。