MOLECULAR-CLONING AND EXPRESSION OF GENETIC-DETERMINANTS FOR THE IRON UPTAKE SYSTEM MEDIATED BY THE VIBRIO-ANGUILLARUM PLASMID PJM1
MOLECULAR-CLONING AND EXPRESSION OF GENETIC-DETERMINANTS FOR THE IRON UPTAKE SYSTEM MEDIATED BY THE VIBRIO-ANGUILLARUM PLASMID PJM1
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DOI：
10.1128/jb.160.3.860-866.1984
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发表时间：
1984-01-01
影响因子：
3.2
通讯作者：
CROSA, JH
中科院分区：
文献类型：
--
作者：
TOLMASKY, ME;CROSA, JH
鳗弧菌侵袭菌株的质粒pJM1编码一个铁摄取系统，该系统介导铁载体和铁-铁载体复合体的膜受体的生物合成。这一系统与鳗弧菌引起海水鱼出血性败血症的能力有关。将含有pJM1介导的铁吸收系统必需区域的重组衍生物克隆到粘粒载体pVK102中，通过三因子交配的方法将其导入低毒力铁吸收缺陷鳗弧菌中。3个重组克隆pJHC-T7、pJHC-T11和pJHC-T2612具有受体活性的遗传决定因素。受体活性的产生与OM2的存在有关，OM2是一种86千道尔顿的外膜蛋白，是在铁限制条件下诱导的。两个克隆pJHC-T7和pJHC-T2612也编码铁载体的活性，尽管水平比野生型低得多。含有这两个克隆的菌株仍然不能在铁限制的条件下生长。只有当细胞中除了重组粘粒外还存在其他本土的pJM1衍生物时，这种能力才能被克服。这种高铁载体产量和在铁限制条件下生长的能力的恢复是实现的，即使在涉及铁载体活性的基因中或在铁载体和受体产生中都存在损伤的情况下也是如此。因此，pJM1介导的另一种功能，可能是一种反式作用因子，可能在铁载体的产生调节中发挥作用。实验感染结果表明，通过将重组克隆导入1株低毒力鳗弧菌，恢复其在铁限制条件下的生长能力，与细菌致病力的增加有关。
Plasmid pJM1 from an invasive strain of V. anguillarum encodes an Fe uptake system which mediates the biosynthesis of a siderophore and a membrane receptor for the Fe-siderophore complex. This system has been associated with the ability of V. anguillarum to cause hemorrhagic septicemic disease in marine fish. Recombinant derivatives containing essential regions of the pJM1-mediated Fe uptake system cloned into cosmid vector pVK102 were introduced into low-virulence Fe uptake-deficient V. anguillarum strains by using a trifactor mating procedure with helper plasmid pRK2013. Three recombinant clones, pJHC-T7, pJHC-T11, and pJHC-T2612, possessed genetic determinants for receptor activity. Production of receptor activity was correlated in all 3 cases with the presence of OM2, an 86-kilodalton outer membrane protein which was induced under Fe-limiting conditions. Two clones, pJHC-T7 and pJHC-T2612, also coded for the production of siderophore activity, although at a much lower level than the wild type. Strains harboring either of these 2 clones were still unable to grow under Fe-limiting conditions. This inability was overcome only when other indigenous pJM1 derivatives were present in the cells in addition to the recombinant cosmids. This restoration of high siderophore production and ability to grow under Fe-limiting conditions was achieved even when the indigenous plasmids possessed lesions in genes involved in siderophore activity or in both siderophore and receptor production. Thus, another function mediated by plasmid pJM1, possibly a trans-acting factor, may play a role in the regulation of siderophore production. Results of experimental infections demonstrated that restoration of the ability to grow under conditions of Fe limitation by introduction of a recombinant clone into 1 of the low-virulence V. anguillarum strains was correlated with an increase in bacterial pathogenicity.