Freitag N E, Seifert H S, Koomey M
Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor 48109-0620, USA.
Mol Microbiol. 1995 May;16(3):575-86. doi: 10.1111/j.1365-2958.1995.tb02420.x.
Expression of Type IV pili by the bacterial pathogen Neisseria gonorrhoeae appears to be essential for colonization of the human host. Several N. gonorrhoeae gene products have been recently identified which bear homology to proteins involved in pilus assembly and protein export in other bacterial systems. We report here the isolation and characterization of transposon insertion mutants in N. gonorrhoeae whose phenotypes indicate that the N. gonorrhoeae pilF and pilD gene products are required for gonoccocal pilus biogenesis. Mutants lacking the pilD gene product, a pre-pilin peptidase, were unable to process the pre-pilin subunit into pilin and thus were non-piliated. pilF mutants processed pilin but did not assemble the mature subunit. Both classes of mutants released S-pilin, a soluble, truncated form of the pilin subunit previously correlated with defects in pilus assembly. In addition, mutants containing transposon insertions in pilD or in a downstream gene, orfX, exhibited a severely restricted growth phenotype. Deletion analysis of pilD indicated that the poor growth phenotype observed for the pilD transposon mutants was a result of polar effects of the insertions on orfX expression. orfX encodes a predicted polypeptide of 23 kDa which contains a consensus nucleotide-binding domain and has apparent homologues in Pseudomonas aeruginosa, Pseudomonas putida, Thermus thermophilus, and the eukaryote Caenorhabditis elegans. Although expression of orfX and pilD appears to be transcriptionally coupled, mutants containing transposon insertions in orfX expressed pili. Unlike either pilF or pilD mutants, orfX mutants were also competent for DNA transformation.
细菌病原体淋病奈瑟菌IV型菌毛的表达似乎对于在人类宿主中定殖至关重要。最近已鉴定出几种淋病奈瑟菌基因产物，它们与其他细菌系统中参与菌毛组装和蛋白质输出的蛋白质具有同源性。我们在此报告淋病奈瑟菌中转座子插入突变体的分离和表征，其表型表明淋病奈瑟菌菌毛F（pilF）和菌毛D（pilD）基因产物是淋球菌菌毛生物合成所必需的。缺乏菌毛D基因产物（一种菌毛前体肽酶）的突变体无法将菌毛前体亚基加工成菌毛蛋白，因此没有菌毛。菌毛F突变体加工了菌毛蛋白，但没有组装成熟亚基。这两类突变体都释放了S-菌毛蛋白，这是一种可溶性的、截短形式的菌毛蛋白亚基，以前与菌毛组装缺陷相关。此外，在菌毛D或下游基因orfX中含有转座子插入的突变体表现出严重受限的生长表型。菌毛D的缺失分析表明，菌毛D转座子突变体观察到的生长不良表型是插入对orfX表达的极性效应的结果。orfX编码一个预测的23 kDa多肽，其包含一个共有核苷酸结合结构域，并且在铜绿假单胞菌、恶臭假单胞菌、嗜热栖热菌和真核生物秀丽隐杆线虫中具有明显的同源物。尽管orfX和菌毛D的表达似乎是转录偶联的，但在orfX中含有转座子插入的突变体表达菌毛。与菌毛F或菌毛D突变体不同，orfX突变体也具有DNA转化能力。
