Attia Ahmed S, Hansen Eric J
Department of Microbiology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-9048, USA.
J Bacteriol. 2006 Nov;188(22):7840-52. doi: 10.1128/JB.01204-06. Epub 2006 Sep 8.
The UspA2 protein has been shown to be directly involved in the serum-resistant phenotype of Moraxella catarrhalis. The predicted 5'-untranslated regions (UTR) of the uspA2 genes in several different M. catarrhalis strains were shown to contain various numbers (i.e., 6 to 23) of a heteropolymeric tetranucleotide (AGAT) repeat. Deletion of the AGAT repeats from the uspA2 genes in the serum-resistant M. catarrhalis strains O35E and O12E resulted in a drastic reduction in UspA2 protein expression and serum resistance. PCR and transformation were used to construct a series of M. catarrhalis O12E strains that differed only in the number of AGAT repeats in their uspA2 genes. Expression of UspA2 was maximal in the presence of 18 AGAT repeats, although serum resistance attained wild-type levels in the presence of as few as nine AGAT repeats. Increased UspA2 expression was correlated with both increased binding of vitronectin and decreased binding of polymerized C9. Real-time reverse transcription-PCR analysis showed that changes in the number of AGAT repeats affected the levels of uspA2 mRNA, with 15 to 18 AGAT repeats yielding maximal levels. Primer extension analysis indicated that these AGAT repeats were contained in the 5'-UTR of the uspA2 gene. The mRNA transcribed from a uspA2 gene containing 18 AGAT repeats was found to have a longer half-life than that transcribed from a uspA2 gene lacking AGAT repeats. These data confirm that the presence of the AGAT repeats in the 5'-UTR of the uspA2 gene is necessary for both normal expression of the UspA2 protein and serum resistance.
已证明UspA2蛋白直接参与卡他莫拉菌的血清抗性表型。在几种不同的卡他莫拉菌菌株中，uspA2基因预测的5'-非翻译区（UTR）含有不同数量（即6至23个）的异聚四核苷酸（AGAT）重复序列。从血清抗性卡他莫拉菌菌株O35E和O12E的uspA2基因中删除AGAT重复序列，导致UspA2蛋白表达和血清抗性急剧降低。使用PCR和转化构建了一系列卡他莫拉菌O12E菌株，它们仅在uspA2基因中的AGAT重复序列数量上有所不同。在存在18个AGAT重复序列时，UspA2的表达最高，尽管在存在少至9个AGAT重复序列时血清抗性达到野生型水平。UspA2表达的增加与玻连蛋白结合增加和聚合C9结合减少相关。实时逆转录-PCR分析表明，AGAT重复序列数量的变化影响uspA2 mRNA的水平，15至18个AGAT重复序列产生最高水平。引物延伸分析表明这些AGAT重复序列包含在uspA2基因的5'-UTR中。发现从含有18个AGAT重复序列的uspA2基因转录的mRNA半衰期比从缺乏AGAT重复序列的uspA2基因转录的mRNA长。这些数据证实，uspA2基因5'-UTR中AGAT重复序列的存在对于UspA2蛋白的正常表达和血清抗性都是必需的。
