Developmentally-regulated excision of the SPβ prophage reconstitutes a gene required for spore envelope maturation in Bacillus subtilis.
Developmentally-regulated excision of the SPβ prophage reconstitutes a gene required for spore envelope maturation in Bacillus subtilis.
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DOI：
10.1371/journal.pgen.1004636
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发表时间：
2014-10
期刊：
影响因子：
4.5
通讯作者：
Sato T
中科院分区：
文献类型：
--
作者：
Abe K;Kawano Y;Iwamoto K;Arai K;Maruyama Y;Eichenberger P;Sato T
温带噬菌体通过将其DNA注入细菌细胞来感染细菌，在那里它作为噬菌体被纳入宿主基因组。在枯草芽孢杆菌168的基因组中，一个活性的噬菌体SPβ被插入到多糖合成基因spsM中。在这里，我们发现通过精确地从染色体上切除spsM β，在孢子形成过程中发生重排，以重建一个功能性的复合spsM基因。SPβ的切除需要一个假定的位点特异性重组酶SprA和一个辅助蛋白SprB。在最小化的SPβ中，除sprA和sprB外，所有的SPβ基因都被删除，在孢子形成过程中保留了SPβ的切除活性，这表明sprA和sprB是必要的和充分的。sprA在营养生长期间表达，sprB在产孢和丝裂霉素C处理期间被诱导，触发噬菌体裂解周期。我们还证明，sprB的过表达（而不是sprA）导致SPβ前噬菌体切除而不触发裂解周期。这些结果表明，sprB是控制噬菌体切除时间的因素。此外，我们提供的证据表明，spsM是必不可少的多糖添加到孢子包膜。孢子表面多糖的存在使孢子在水中具有亲水性。这种特性可能有利于孢子通过水流在自然环境中传播。类似的重排发生在解淀粉芽孢杆菌FZB42中，其中在孢子形成过程中切除sp β样元件以重建多糖合成基因，这表明这种类型的基因重排在孢子形成细菌中很常见，因为它可以通过噬菌体感染传播。将噬菌体整合到宿主染色体的蛋白质编码序列中通常会导致被中断基因功能的丧失。在胞内形成生物枯草芽孢杆菌菌株168中，SPβ原噬菌体被插入到一个以前未被表征的孢子多糖合成基因spsM中。在营养细胞中，DNA损伤诱导了裂解周期。在此过程中，从基因组中切除SPβ以形成噬菌体颗粒。在这里，我们证明SPβ切除也是一个发育调节事件，在孢子形成过程中系统地发生，以重建功能性spsM基因。在孢子细胞不对称分裂之后，产生了两个细胞室，前孢子和母细胞，前者将成熟为孢子，后者对孢子成熟过程至关重要。由于噬菌体切除仅限于母细胞基因组，而不会发生在前孢子基因组中，因此SPβ是孢子基因组的一个组成部分。因此，孢子萌发后，营养细胞恢复生长，SPβ噬菌体与宿主基因组的其余部分一起垂直繁殖到后代。我们的研究结果表明，SPβ切除的两种途径都支持噬菌体的生命周期和宿主细胞的正常产孢。
Temperate phages infect bacteria by injecting their DNA into bacterial cells, where it becomes incorporated into the host genome as a prophage. In the genome of Bacillus subtilis 168, an active prophage, SPβ, is inserted into a polysaccharide synthesis gene, spsM. Here, we show that a rearrangement occurs during sporulation to reconstitute a functional composite spsM gene by precise excision of SPβ from the chromosome. SPβ excision requires a putative site-specific recombinase, SprA, and an accessory protein, SprB. A minimized SPβ, where all the SPβ genes were deleted, except sprA and sprB, retained the SPβ excision activity during sporulation, demonstrating that sprA and sprB are necessary and sufficient for the excision. While expression of sprA was observed during vegetative growth, sprB was induced during sporulation and upon mitomycin C treatment, which triggers the phage lytic cycle. We also demonstrated that overexpression of sprB (but not of sprA) resulted in SPβ prophage excision without triggering the lytic cycle. These results suggest that sprB is the factor that controls the timing of phage excision. Furthermore, we provide evidence that spsM is essential for the addition of polysaccharides to the spore envelope. The presence of polysaccharides on the spore surface renders the spore hydrophilic in water. This property may be beneficial in allowing spores to disperse in natural environments via water flow. A similar rearrangement occurs in Bacillus amyloliquefaciens FZB42, where a SPβ-like element is excised during sporulation to reconstitute a polysaccharide synthesis gene, suggesting that this type of gene rearrangement is common in spore-forming bacteria because it can be spread by phage infection. Integration of prophages into protein-coding sequences of the host chromosome generally results in loss of function of the interrupted gene. In the endospore-forming organism Bacillus subtilis strain 168, the SPβ prophage is inserted into a previously-uncharacterized spore polysaccharide synthesis gene, spsM. In vegetative cells, the lytic cycle is induced in response to DNA damage. In the process, SPβ is excised from the genome to form phage particles. Here, we demonstrate that SPβ excision is also a developmentally-regulated event that occurs systematically during sporulation to reconstitute a functional spsM gene. Following asymmetric division of the sporulating cell, two cellular compartments are generated, the forespore, which will mature into a spore, and the mother cell, which is essential to the process of spore maturation. Because phage excision is limited to the mother cell genome, and does not occur in the forespore genome, SPβ is an integral part of the spore genome. Thus, after the spores germinate, the vegetative cells resume growth and the SPβ prophage is propagated vertically to the progeny along with the rest of the host genome. Our results suggest that the two pathways of SPβ excision support both the phage life cycle and normal sporulation of the host cells.
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影响因子：
9.2
作者：
McKenney, Peter T.;Driks, Adam;Eskandarian, Haig A.;Grabowski, Paul;Guberman, Jonathan;Wang, Katherine H.;Gitai, Zemer;Eichenberger, Patrick
通讯作者：
Eichenberger, Patrick
影响因子：
11.3
作者：
Higgins D;Dworkin J
通讯作者：
Dworkin J
影响因子：
3.2
作者：
Driks, A
通讯作者：
Driks, A
影响因子：
9.8
作者：
Eichenberger P;Fujita M;Jensen ST;Conlon EM;Rudner DZ;Wang ST;Ferguson C;Haga K;Sato T;Liu JS;Losick R
通讯作者：
Losick R
影响因子：
3.2
作者：
Imamura, Daisuke;Kuwana, Ritsuko;Watabe, Kazuhito
通讯作者：
Watabe, Kazuhito