许多原核生物利用游泳运动向有利的条件移动并逃避不利的环境。控制细菌鞭毛驱动运动的调节机制已经建立;然而,关于古细菌细胞表面结构推动的游泳运动的调节还知之甚少,Archaella.先前的研究表明,粘附菌毛(PilA1-6)的缺失,IV型菌毛细胞表面结构的亚基,使古细菌Haloferax火山模型无法活动。在这项研究中,我们使用甲磺酸乙酯诱变和运动性测定来鉴定pilaA[1-6]菌株的运动性抑制剂。在确定的八种抑制剂中,其中6个包含古细菌生物合成基因的错义突变,ArlI和ArlJ.在各自的多缺失菌株ΔpilA[1-6]ΔarlI和arlJ突变体构建体中的反式表达中,ΔpilA[1-6]ΔarlJ证实了它们在抑制ΔpilA[1-6]运动性缺陷中的作用。此外,三种抑制因子在cirA中同时存在破坏性错义和无义突变,编码一种拟议的调节蛋白的基因。cirA缺失导致运动过度,而野生型细胞中cirA反式表达导致运动性降低。此外,实时定量PCR分析显示,在野生型细胞中,较高的ARLI表达水平,arlJ,与非运动性对数中期盘状细胞相比,在活动的早期对数期杆状细胞中观察到了古细菌基因arlA1。相反,取决于cirA细胞,在对数早期和中期形成棒,在两个生长期中显示出相似的arl基因表达水平。我们的发现有助于更深入地了解控制古细菌运动性的机制,强调ArlI的参与,ArlJ,和CirA在菌丝介导的运动性调节中的作用。重要古细菌是真核生物的近亲,起着至关重要的生态作用。某些行为,如游泳运动,被认为对古细菌环境适应很重要。Archaella,古细菌的运动性附属物,在进化上与细菌鞭毛不同,驱动古细菌运动性的调节机制在很大程度上是未知的。先前的研究已将IV型菌毛亚基的丢失与古细菌运动性抑制联系起来。这项研究揭示了三种Haloferax火山蛋白参与菌毛蛋白介导的运动调节,在这个未被研究的领域中提供了对运动性调节的更深入的了解,同时也为发现控制古细菌运动性的新机制铺平了道路。了解古细菌细胞过程将有助于阐明古细菌的生态作用以及这些过程的跨域演变。
Many prokaryotes use swimming motility to move toward favorable conditions and escape adverse surroundings. Regulatory mechanisms governing bacterial flagella-driven motility are well-established; however, little is yet known about the regulation underlying swimming motility propelled by the archaeal cell surface structure, the archaella. Previous research showed that the deletion of the adhesion pilins (PilA1-6), subunits of the type IV pili cell surface structure, renders the model archaeon Haloferax volcanii non-motile. In this study, we used ethyl methanesulfonate mutagenesis and a motility assay to identify motile suppressors of the ∆pilA[1-6] strain. Of the eight suppressors identified, six contain missense mutations in archaella biosynthesis genes, arlI and arlJ. In trans expression of arlI and arlJ mutant constructs in the respective multi-deletion strains ∆pilA[1-6]∆arlI and ∆pilA[1-6]∆arlJ confirmed their role in suppressing the ∆pilA[1-6] motility defect. Additionally, three suppressors harbor co-occurring disruptive missense and nonsense mutations in cirA, a gene encoding a proposed regulatory protein. A deletion of cirA resulted in hypermotility, while cirA expression in trans in wild-type cells led to decreased motility. Moreover, quantitative real-time PCR analysis revealed that in wild-type cells, higher expression levels of arlI, arlJ, and the archaellin gene arlA1 were observed in motile early-log phase rod-shaped cells compared to non-motile mid-log phase disk-shaped cells. Conversely, ∆cirA cells, which form rods during both early- and mid-log phases, exhibited similar expression levels of arl genes in both growth phases. Our findings contribute to a deeper understanding of the mechanisms governing archaeal motility, highlighting the involvement of ArlI, ArlJ, and CirA in pilin-mediated motility regulation.IMPORTANCEArchaea are close relatives of eukaryotes and play crucial ecological roles. Certain behaviors, such as swimming motility, are thought to be important for archaeal environmental adaptation. Archaella, the archaeal motility appendages, are evolutionarily distinct from bacterial flagella, and the regulatory mechanisms driving archaeal motility are largely unknown. Previous research has linked the loss of type IV pili subunits to archaeal motility suppression. This study reveals three Haloferax volcanii proteins involved in pilin-mediated motility regulation, offering a deeper understanding of motility regulation in this understudied domain while also paving the way for uncovering novel mechanisms that govern archaeal motility. Understanding archaeal cellular processes will help elucidate the ecological roles of archaea as well as the evolution of these processes across domains.