Vibrio parahaemolyticus rapidly colonizes surfaces using swarming motility. Surface contact induces the surface-sensing regulon, including lateral flagellar genes, spurring dramatic shifts in physiology and behavior. The bacterium can also adopt a sessile, surface-associated lifestyle and form robust biofilms. These alternate colonization strategies are influenced reciprocally by the second messenger c-di-GMP. Although V. parahaemolyticus possesses 43 predicted proteins with the c-di-GMP-forming GGDEF domain, none have been previously been identified as contributors to surface colonization. We sought to explore this knowledge gap by using a suppressor transposon screen to restore the swarming motility of a nonswarming, high-c-di-GMP strain. Two diguanylate cyclases, ScrJ and ScrL, each containing tetratricopeptide repeat-coupled GGDEF domains, were demonstrated to contribute additively to swarming gene repression. Both proteins required an intact catalytic motif to regulate. Another suppressor mapped in lafV, the last gene in a lateral flagellar operon. Containing a degenerate phosphodiesterase (EAL) domain, LafV repressed transcription of multiple genes in the surface sensing regulon; its repressive activity required LafK, the primary swarming regulator. Mutation of the signature EAL motif had little effect on LafV’s repressive activity, suggesting that LafV belongs to the subclass of EAL-type proteins that are regulatory but not enzymatic. Consistent with these activities and their predicted effects on c-di-GMP, scrJ and scrL but not lafV, mutants affected the transcription of the c-di-GMP-responsive biofilm reporter cpsA::lacZ. Our results expand the knowledge of the V. parahaemolyticus GGDEF/EAL repertoire and its roles in this surface colonization regulatory network.

中文翻译：

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三种新的GGDEF和EAL域蛋白参与副溶血性弧菌Scr表面定殖调控网络的鉴定。

副溶血性弧菌利用群体运动迅速在表面定殖。表面接触诱导包括侧鞭毛基因在内的表面感应调节子，刺激生理和行为的急剧变化。该细菌还可以采取无柄的，与表面相关的生活方式，并形成坚固的生物膜。这些替代定居策略受到第二信使c-di-GMP的相互影响。尽管副溶血性弧菌拥有43个具有c-di-GMP形成GGDEF结构域的预测蛋白，以前没有一个被确定为表面定殖的贡献者。我们试图通过使用抑制子转座子筛选来恢复不易变迁的高c-di-GMP株的群体运动来探索这一知识鸿沟。两个二鸟苷酸环化酶ScrJ和ScrL，每个都包含四肽重复偶联的GGDEF结构域，被证明有助于蜂拥而至的基因抑制。两种蛋白质都需要完整的催化基序来调节。在lafV中映射的另一个抑制器，在鞭毛外侧操纵子中的最后一个基因。LafV含有简并的磷酸二酯酶（EAL）域，可抑制表面传感调节子中多个基因的转录；其抑制活动需要主要的蜂群调节剂LafK。标志性EAL基序的突变对LafV的抑制活性影响很小，这表明LafV属于EAL型蛋白的亚类，该蛋白具有调节性，但不具有酶促性。与这些活动及其对c-di-GMP，scrJ和scrL的预期影响一致，但对lafV没有影响，这些突变体影响了c-di-GMP响应性生物膜报道基因cpsA :: lacZ的转录。我们的结果扩大了副溶血性弧菌的知识 GGDEF / EAL库及其在此表面定殖调控网络中的作用。