Chung Chih-Ching, Hwang Sheng-Ping L, Chang Jeng
Institute of Marine Biology, National Taiwan Ocean University, Keelung 20224, Taiwan, Republic of China.
Appl Environ Microbiol. 2003 Feb;69(2):754-9. doi: 10.1128/AEM.69.2.754-759.2003.
A high-affinity phosphate transporter gene, TcPHO, was isolated from a growth-dependent subtracted cDNA library of the marine unicellular alga Tetraselmis chui. The full-length cDNA of TcPHO obtained by 5' and 3' rapid amplification of cDNA ends was 1,993 bp long and encoded an open reading frame consisting of 610 amino acids. The deduced amino acid sequence of TcPHO exhibited 51.6 and 49.8% similarity to the amino acid sequences of PHO89 from Saccharomyces cerevisiae and PHO4 from Neurospora crassa, respectively. In addition, hydrophobicity and secondary structure analyses revealed 12 conserved transmembrane domains that were the same as those found in PHO89 and PHO4. The expression of TcPHO mRNA was dependent on phosphate availability. With a low-phosphate treatment, the TcPHO mRNA concentration increased sharply to 2.72 fmol micro g of total RNA(-1) from day 1 to day 2 and remained at this high level from days 2 to 4. Furthermore, rescue treatment with either phosphate or p-nitrophenyl phosphate effectively inhibited TcPHO mRNA expression. In contrast, TcPHO mRNA expression stayed at a low level (range, 0.25 to 0.28 fmol micro g of total RNA(-1)) under low-nitrate conditions. The expression pattern suggests that TcPHO can be used as a molecular probe for monitoring phosphorus stress in T. chui.
从海洋单细胞藻类杜氏盐藻生长依赖型消减cDNA文库中分离出一个高亲和力磷酸盐转运体基因TcPHO。通过cDNA末端的5'和3'快速扩增获得的TcPHO全长cDNA长1993 bp，编码一个由610个氨基酸组成的开放阅读框。推导的TcPHO氨基酸序列与酿酒酵母的PHO89和粗糙脉孢菌的PHO4的氨基酸序列分别具有51.6%和49.8%的相似性。此外，疏水性和二级结构分析揭示了12个保守的跨膜结构域，与PHO89和PHO4中的相同。TcPHO mRNA的表达取决于磷酸盐的可用性。在低磷酸盐处理下，TcPHO mRNA浓度从第1天到第2天急剧增加到2.72 fmol μg总RNA（-1），并在第2天到第4天保持在这个高水平。此外，用磷酸盐或对硝基苯磷酸盐进行挽救处理有效地抑制了TcPHO mRNA的表达。相反，在低硝酸盐条件下，TcPHO mRNA表达保持在低水平（范围为0.25至0.28 fmol μg总RNA（-1））。这种表达模式表明，TcPHO可作为监测杜氏盐藻磷胁迫的分子探针。
