Zhang Hongming, Marconi Richard T
Department of Microbiology and Immunology, Richmond, VA 23298-0678, USA.
J Bacteriol. 2005 Dec;187(23):7985-95. doi: 10.1128/JB.187.23.7985-7995.2005.
The Borrelia genome is comprised of linear and circular elements, including a group of 32-kb circular plasmids (cp32s). Earlier analyses identified a bacteriophage, varphiBB-1, that may package cp32s, suggesting that these plasmids are prophages. cp32-8, cp32-9, and cp32-1 (plasmids L, N, and P, respectively) encode virulence factors such as the factor H binding, OspE proteins (BBL39, BBN38, and BBP38). Here the expression patterns of cp32-8 open reading frames (ORFs) in in vitro-cultivated 1-methyl-3-nitroso-nitroguanidine (MNNG)-treated and untreated spirochetes and during infection were assessed. ORFs BBL42 through BBL28, which encode several bacteriophage protein homologs, were found to be cotranscribed and expression was upregulated by MNNG. Immunoblotting revealed that MNNG-induced transcription led to increased protein production. The expression of several genes that reside outside of the BBL42-BBL28 operon was not affected by MNNG. Some of these genes, including OspE (BBL39), appear to represent morons. Real-time reverse transcription-PCR of spirochetes in mouse tissue revealed that although the phage operon was not induced during infection, transcription of BBL23 (previously designated BlyA), a putative holin, was upregulated. This observation indicates that some genes within the operon can be independently transcribed from internal promoters. Additional transcriptional analyses of the operon identified multiple transcriptional start sites and provided evidence for the expression of a homologous operon from other cp32s. The data support the hypothesis put forth by C. Eggers and D. S. Samuels (J. Bacteriol. 181:7308-7313, 1999) that the cp32s are prophages, a finding with broad implications for our understanding of Borrelia pathogenesis and Borrelia genome evolution.
伯氏疏螺旋体基因组由线性和环状元件组成，包括一组32 kb的环状质粒（cp32s）。早期分析鉴定出一种噬菌体，即φBB - 1，它可能包裹cp32s，这表明这些质粒是前噬菌体。cp32 - 8、cp32 - 9和cp32 - 1（分别为质粒L、N和P）编码诸如因子H结合蛋白、OspE蛋白（BBL39、BBN38和BBP38）等毒力因子。在此，评估了cp32 - 8开放阅读框（ORF）在体外培养的经1 - 甲基 - 3 - 亚硝基胍（MNNG）处理和未处理的螺旋体中的表达模式以及在感染过程中的表达模式。发现编码几种噬菌体蛋白同源物的ORF BBL42至BBL28是共转录的，并且其表达在MNNG处理后上调。免疫印迹显示MNNG诱导的转录导致蛋白质产量增加。位于BBL42 - BBL28操纵子之外的几个基因的表达不受MNNG影响。其中一些基因，包括OspE（BBL39），似乎代表了基因块。对小鼠组织中螺旋体的实时逆转录PCR分析表明，虽然噬菌体操纵子在感染过程中未被诱导，但推定的成孔蛋白BBL23（先前称为BlyA）的转录上调。该观察结果表明操纵子内的一些基因可以从内部启动子独立转录。对该操纵子的进一步转录分析确定了多个转录起始位点，并为来自其他cp32s的同源操纵子的表达提供了证据。这些数据支持了C. Eggers和D. S. Samuels（《细菌学杂志》181:7308 - 7313，1999）提出的假设，即cp32s是前噬菌体，这一发现对我们理解伯氏疏螺旋体的发病机制和伯氏疏螺旋体基因组进化具有广泛的意义。
