Haran S, Logendra S, Seskar M, Bratanova M, Raskin I
Biotech Center, Foran Hall, Cook College, Rutgers University, New Brunswick, New Jersey 08901-8520, USA.
Plant Physiol. 2000 Oct;124(2):615-26. doi: 10.1104/pp.124.2.615.
The expression and secretion of acid phosphatase (APase) was investigated in Indian mustard (Brassica juncea L. Czern.) plants using sensitive in vitro and activity gel assays. Phosphorus (P) starvation induced two APases in Indian mustard roots, only one of which was secreted. Northern-blot analysis indicated transcriptional regulation of APase expression. Polymerase chain reaction and Southern-blot analyses revealed two APase homologs in Indian mustard, whereas in Arabidopsis, only one APase homolog was detected. The Arabidopsis APase promoter region was cloned and fused to the beta-glucuronidase (GUS) and green fluorescent protein (GFP) reporter genes. GUS expression was first evident in leaves of the P-starved Arabidopsis plants. In P-starved roots, the expression of GUS initiated in lateral root meristems followed by generalized expression throughout the root. GUS expression diminished with the addition of P to the medium. Expression of GFP in P-starved roots also initiated in the lateral root meristems and the recombinant GFP with the APase signal peptide was secreted by the roots into the medium. The APase promoter was specifically activated by low P levels. The removal of other essential elements or the addition of salicylic or jasmonic acids, known inducers of gene expression, did not activate the APase promoter. This novel APase promoter may be used as a plant-inducible gene expression system for the production of recombinant proteins and as a tool to study P metabolism in plants.
利用灵敏的体外和活性凝胶分析方法,对印度芥菜(Brassica juncea L. Czern.)植株中酸性磷酸酶(APase)的表达和分泌进行了研究。磷(P)饥饿诱导印度芥菜根中产生两种APase,但只有一种被分泌。Northern杂交分析表明APase表达受转录调控。聚合酶链反应和Southern杂交分析显示印度芥菜中有两个APase同源物,而在拟南芥中只检测到一个APase同源物。克隆了拟南芥APase启动子区域,并将其与β-葡萄糖醛酸酶(GUS)和绿色荧光蛋白(GFP)报告基因融合。GUS表达最初在缺磷拟南芥植株的叶片中明显可见。在缺磷根中,GUS表达在侧根分生组织中启动,随后在整个根中普遍表达。向培养基中添加磷后,GUS表达减弱。缺磷根中GFP的表达也在侧根分生组织中启动,带有APase信号肽的重组GFP被根分泌到培养基中。APase启动子在低磷水平下被特异性激活。去除其他必需元素或添加已知的基因表达诱导剂水杨酸或茉莉酸,均不能激活APase启动子。这种新型的APase启动子可作为一种用于生产重组蛋白的植物诱导型基因表达系统,以及研究植物磷代谢的工具。