Molecular analysis of phosphate taxis in bacteria.

A)我们分离到了产碱性磷酸酶(AP)的铜绿假单胞菌PAO1和阴沟肠杆菌IFO3320突变株。对这些突变体的遗传分析表明，突变位于PST操纵子。铜绿假单胞菌突变株在P_i过量条件下对P_i表现出趋化反应。这些结果表明，PST对铜绿假单胞菌P_I趋向性的表达具有负性调控作用。阴沟肠杆菌AP组分突变体即使在P_i限制条件下也不表现出对P_i的趋化反应。这些结果表明，阴沟肠杆菌的PST系统是P_I化学感受所必需的。B)由于目前尚无简便的方法筛选P_i缺陷突变体，我们决定以编码氨基酸MCP的PCTA基因为探针，克隆编码P_i趋向性的MCP基因。在PCTA侧翼区发现了两个MCP基因(命名为pctB和PCTC)。遗传分析表明，PCtB和PCTC分别参与了对7个和2个L氨基酸的趋化作用。铜绿假单胞菌基因敲除突变株PCTA、PCtB和PCTC具有P_I趋向性，提示3个MCP不参与P_I趋向性。我们又分离到了7个与PCTA基因杂交的DNA片段。对这些碎片的调查正在进行中。

a) We isolated P.aeruginosa PAO1 and E.cloacae IFO3320 mutants which constitutively produced alkaline phospgatase (AP). Genetic analysis of these mutants revealed that mutations located in the pst operon. P.aeruginosa mutant strains showed chemotactic response to P_i under conditions of P_i excess. These results indicate that Pst negatively controls the expression of P_i taxis in P.aeruginosa. E.cloacae AP constitutive mutants did not show chemotactic response to P_i even under conditions of P_i limitations. These results suggest that the E.cloacae Pst system is required for P_i chemoreception. b) Since there is no convenient method for selecting P_i defective mutants, we decided to clone a gene encoding a MCP for P_i taxis by using the pctA gene, which encodes a MCP for amino acids, as probe. Two MCP genes (termed pctB and pctC) were found in the pctA-flanking region. Genetic analysis revealed that PctB and PctC were involved in chemotaxis to seven and two L-amino acids, respectively. The P.aeruginosa knock-out mutant of pctA,pctB and pctC showed P_i taxis, suggestig that three MCPs were not involved in P_i taxis. We have isolated seven more DNA fragments hybridized with the pctA gene. Investigation of these fragments is under way.