Lee J H, Isaacson R E
Department of Veterinary Pathobiology, University of Illinois, Urbana 61801, USA.
Infect Immun. 1995 Oct;63(10):4143-9. doi: 10.1128/iai.63.10.4143-4149.1995.
The biogenesis of the pilus adhesin K99 is dependent on the expression of eight contiguous genes, fanA to fanH. Transposon mutants were prepared by using TnlacZ and TnphoA, and selected transposon mutants were used to measure expression of each K99 gene. Expression of the K99 genes is likely controlled at the transcription level, since in general, there were no differences between the results obtained with the two transposons. fanC was the most highly expressed, and fanD was expressed at very low levels. The expression of TnlacZ fusions in fanA and fanB fusions was high. Deletion of fanA, fanB, and part of fanC abolished the expression of fanD but had no effect on the distal genes fanE to fanH. To locate the DNA regions required for expression of fanE to fanH, deletion mutations were prepared and the effects on expression of fanE to fanH were determined. The deletion of a segment between fanD and fanE abolished fanE and fanF expression but did not affect fanG and fanH. The deletion of a portion of fanF (approximately 1 kb proximal to fanG) abolished the expression of fanG and fanH. These results indicate the presence of regulatory elements proximal to fanE and to fanG. Putative promoters were identified in these regions by DNA homology and by primer extension. A stem-loop structure that may act as a transcriptional attenuator of fanF was also found at the beginning of fanF. These data confirm our previous model of K99 transcriptional organization.
菌毛粘附素K99的生物合成依赖于8个相邻基因（fanA至fanH）的表达。利用TnlacZ和TnphoA制备转座子突变体，并使用选定的转座子突变体来测量每个K99基因的表达。K99基因的表达可能在转录水平受到调控，因为总体而言，两种转座子获得的结果之间没有差异。fanC表达量最高，而fanD表达水平极低。fanA和fanB融合体中TnlacZ融合体的表达较高。缺失fanA、fanB和部分fanC会消除fanD的表达，但对远端基因fanE至fanH没有影响。为了定位fanE至fanH表达所需的DNA区域，制备了缺失突变体并确定其对fanE至fanH表达的影响。fanD和fanE之间一段序列的缺失消除了fanE和fanF的表达，但不影响fanG和fanH。缺失部分fanF（fanG近端约1 kb）消除了fanG和fanH的表达。这些结果表明在fanE和fanG近端存在调控元件。通过DNA同源性和引物延伸在这些区域鉴定出了推定的启动子。在fanF起始处还发现了一个可能作为fanF转录衰减子的茎环结构。这些数据证实了我们之前关于K99转录组织的模型。
