Provence D L, Curtiss R
Department of Biology, Washington University, St. Louis, Missouri 63130.
Infect Immun. 1992 Nov;60(11):4460-7. doi: 10.1128/iai.60.11.4460-4467.1992.
This study determined the role of crl in the production of curli by, the hemagglutination activity of, and fibronectin binding by avian pathogenic Escherichia coli chi 7122. Curli, an extracellular structure that binds fibronectin, was recently described (A. Olsén, A. Jonsson, and S. Normark, Nature [London] 338:652-655, 1989). The crl gene product was hypothesized to be the subunit monomer of curli and to bind fibronectin. E. coli HB101, which does not contain crl, binds fibronectin and produces curli when harboring a plasmid containing the crl gene. We show that HB101 hemagglutinates chicken erythrocytes when harboring the crl gene and that chi 7122 hemagglutinates chicken erythrocytes, binds fibronectin, and produces curli. Hemagglutination activity, fibronectin binding, and curli production are best expressed by chi 7122 and HB101 harboring the crl gene when the bacteria are grown on colonization factor antigen agar at 26 degrees C. The expression of hemagglutination activity, fibronectin binding, and curli production by both strains is decreased by growth on this agar at an increased temperature, of an increased osmolarity, or in an anaerobic atmosphere. This results indicates that the crl gene plays a role in the expression of the three phenotypes in HB101 and possibly in chi 7122 as well. We inactivated crl in chi 7122 by allele replacement in the expectation of abolishing hemagglutination activity, fibronectin binding, and curli production. The mutation was verified by Southern blot analysis and by a polymerase chain reaction, and there was no evidence of a second crl gene in chi 7122. However, the mutant of chi 7122 lacking crl hemagglutinated chicken erythrocytes, bound fibronectin, and produced curli at wild-type levels. This result indicates that crl plays a nonessential role in the expression of these three phenotypes in chi 7122.
本研究确定了卷曲菌毛调控基因（crl）在禽致病性大肠杆菌chi 7122产生卷曲菌毛、血凝活性以及结合纤连蛋白过程中的作用。卷曲菌毛是一种能结合纤连蛋白的细胞外结构，最近已有相关报道（A. 奥尔森、A. 琼森和S. 诺马克，《自然》[伦敦]338:652 - 655, 1989）。推测卷曲菌毛调控基因（crl）的产物是卷曲菌毛的亚基单体，并能结合纤连蛋白。不含卷曲菌毛调控基因（crl）的大肠杆菌HB101，在携带含有卷曲菌毛调控基因（crl）的质粒时，能结合纤连蛋白并产生卷曲菌毛。我们发现，携带卷曲菌毛调控基因（crl）的HB101能凝集鸡红细胞，chi 7122也能凝集鸡红细胞、结合纤连蛋白并产生卷曲菌毛。当细菌在26摄氏度的定居因子抗原琼脂上生长时，chi 7122和携带卷曲菌毛调控基因（crl）的HB101的血凝活性、纤连蛋白结合能力和卷曲菌毛产生能力表现最佳。在温度升高、渗透压增加或厌氧环境下，在这种琼脂上生长会降低两种菌株的血凝活性、纤连蛋白结合能力和卷曲菌毛产生能力。这一结果表明，卷曲菌毛调控基因（crl）在HB101的这三种表型表达中发挥作用，在chi 7122中可能也发挥作用。我们通过等位基因替换使chi 7122中的卷曲菌毛调控基因（crl）失活，期望消除其血凝活性、纤连蛋白结合能力和卷曲菌毛产生能力。通过Southern印迹分析和聚合酶链反应验证了该突变，且没有证据表明chi 7122中存在第二个卷曲菌毛调控基因（crl）。然而，缺失卷曲菌毛调控基因（crl）的chi 7122突变体仍能凝集鸡红细胞、结合纤连蛋白并以野生型水平产生卷曲菌毛。这一结果表明，卷曲菌毛调控基因（crl）在chi 7122的这三种表型表达中发挥非必需作用。
