Redkar R J, Herzog R W, Singh N K
Department of Botany and Microbiology, Auburn University, Auburn, Alabama 36849, USA.
Appl Environ Microbiol. 1998 Jun;64(6):2229-31. doi: 10.1128/AEM.64.6.2229-2231.1998.
A differentially expressed gpdA cDNA clone was isolated from NaCl-adapted Aspergillus nidulans (FGSC359) and identified as glyceraldehyde-3-phosphate dehydrogenase (gpdA) on the basis of its nucleotide sequence. The level of gpdA RNA substantially increased in cultures gradually adapted to NaCl but was greatly reduced in cultures exposed briefly to a high concentration of NaCl. A pyrG auxotroph of A. nidulans (A773) was cotransformed with a gpdA-uidA construct and a plasmid containing the Neurospora crassa pyr4 gene as a selectable marker. One pyrG+ beta-glucuronidase-positive (GUS+) transformant was selected, and stable integration of the gpdA-uidA construct into the genome was confirmed by Southern blot analysis. Gradual adaptation to increasing concentrations of NaCl resulted in an increase in GUS activity to 2.7-fold. GUS activity was reduced after a 2-h exposure of an unadapted culture to 2 M NaCl but gradually increased to a maximum of twofold after 24 h. GUS activity also increased by 8.4-fold in Na2SO4-adapted cultures, 4.9-fold in polyethylene glycol-adapted cultures, and 7.5-fold in KCl-adapted cultures. These results are consistent with the hypothesis that the A. nidulans gpdA promoter is transcriptionally activated by osmotic signals.
从适应氯化钠的构巢曲霉(FGSC359)中分离出一个差异表达的gpdA cDNA克隆,并根据其核苷酸序列将其鉴定为甘油醛-3-磷酸脱氢酶(gpdA)。在逐渐适应氯化钠的培养物中,gpdA RNA水平大幅增加,但在短暂暴露于高浓度氯化钠的培养物中则大幅降低。将构巢曲霉的一个pyrG营养缺陷型(A773)与一个gpdA-uidA构建体和一个含有粗糙脉孢菌pyr4基因作为选择标记的质粒共转化。选择了一个pyrG+β-葡萄糖醛酸酶阳性(GUS+)转化体,并通过Southern印迹分析证实了gpdA-uidA构建体稳定整合到基因组中。逐渐适应浓度不断增加的氯化钠导致GUS活性增加到2.7倍。未适应的培养物暴露于2 M氯化钠2小时后,GUS活性降低,但24小时后逐渐增加到最大两倍。在适应硫酸钠的培养物中,GUS活性也增加了8.4倍,在适应聚乙二醇的培养物中增加了4.9倍,在适应氯化钾的培养物中增加了7.5倍。这些结果与构巢曲霉gpdA启动子被渗透信号转录激活的假设一致。