Identification of structural and regulatory cell-shape determinants in Haloferax volcanii

Schiller, Heather; Hong, Yirui; Kouassi, Joshua; Rados, Theopi; Kwak, Jasmin; DiLucido, Anthony; Safer, Daniel; Marchfelder, Anita; Pfeiffer, Friedhelm; Bisson, Alexandre; Schulze, Stefan; Pohlschroder, Mechthild

doi:10.1038/s41467-024-45196-0

Cited by 14 publications

(13 citation statements)

References 79 publications

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“…Interestingly, ArlA1 abundance is not as drastically different in the growth phase comparisons of JK3, suggesting that CirA is involved in the regulation of ArlA1. While differences in protein abundance of ArlI or ArlJ were not observed in wild type or JK3 25 , the abundance of archaella biosynthetic components could be below the detection level of quantitative proteomics experiment, consistent with the absence of any ArlI or ArlJ peptides identified in the Archaeal Proteome Project (ArcPP) database 34 .…”

Section: Discussionsupporting

confidence: 60%

“…5 ), we predicted that the loss of CirA affects the abundance of archaellins and the archaella biosynthetic components. Consistent with this hypothesis, the proteomics studies conducted by Schiller et al determined that archaellin ArlA1 occurs at a higher abundance in early-log rod-shaped cells when compared to late-log disk-shaped cells 25 . While no differential abundance was observed for the biosynthetic components of the archaella, to date, proteomics has failed to identify ArlI or ArlJ peptides under any conditions tested 34 , suggesting low abundance of these biosynthetic components.…”

Section: Resultsmentioning

confidence: 73%

“…It is transcribed in the opposite direction as the arl genes. The genic locations of the residues corresponding to the mutations found in the Δ pilA [ 1-6 ] suppressor mutants as well as hypermotile mutants from a previous study 25 are indicated, along with the conserved Walker A and Walker B motifs.…”

Section: Resultsmentioning

confidence: 99%

“…determined that archaellin ArlA1 occurs at a higher abundance in early-log rod-shaped cells when compared to late-log disk-shaped cells 25 . While no differential abundance was observed for the biosynthetic components of the archaella, to date, proteomics has failed to identify ArlI or ArlJ peptides under any conditions tested 34 , suggesting low abundance of these biosynthetic components.…”

Section: Cira Regulates Motility By Reducing Transcription Of Arl Genesmentioning

confidence: 96%

“…In our previous study, two mutants isolated from a hypermotility screen were rod-only in the conditions tested. Those mutants had premature stop codons in the gene cirA which encodes Circadian regulator CirA 25 . Similar to ArlH, CirA is also a KaiC-like protein, and cirA is located within the arl gene cluster 22 .…”

Section: Introductionmentioning

confidence: 99%

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Involvement of ArlI, ArlJ, and CirA in Archaeal Type-IV Pilin-Mediated Motility Regulation

Chatterjee,

Garcia,

Cote

et al. 2024

Preprint

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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 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 EMS 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. Overexpression of these 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 overexpression in wild-type cells led to decreased motility. Moreover, qRT-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.

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Section: Discussionsupporting

confidence: 60%

Section: Resultsmentioning

confidence: 73%

Section: Resultsmentioning

confidence: 99%

Section: Cira Regulates Motility By Reducing Transcription Of Arl Genesmentioning

confidence: 96%

Section: Introductionmentioning

confidence: 99%

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Involvement of ArlI, ArlJ, and CirA in Archaeal Type-IV Pilin-Mediated Motility Regulation

Chatterjee,

Garcia,

Cote

et al. 2024

Preprint

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Halofilins as emerging bactofilin families of archaeal cell shape plasticity orchestrators

Curtis,

Escudeiro,

Mallon

et al. 2024

Proc. Natl. Acad. Sci. U.S.A.

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Bactofilins are rigid, nonpolar bacterial cytoskeletal filaments that link cellular processes to specific curvatures of the cytoplasmic membrane. Although homologs of bactofilins have been identified in archaea and eukaryotes, functional studies have remained confined to bacterial systems. Here, we characterize representatives of two families of archaeal bactofilins from the pleomorphic archaeon Haloferax volcanii , halofilin A (HalA) and halofilin B (HalB). HalA and HalB polymerize in vitro, assembling into straight bundles. HalA polymers are highly dynamic and accumulate at positive membrane curvatures in vivo, whereas HalB forms more static foci that localize in areas of local negative curvatures on the outer cell surface. Gene deletions and live-cell imaging show that halofilins are critical in maintaining morphological integrity during shape transition from disk (sessile) to rod (motile). Morphological defects in Δ halA result in accumulation of highly positive curvatures in rods but not in disks. Conversely, disk-shaped cells are exclusively affected by halB deletion, resulting in flatter cells. Furthermore, while Δ halA and Δ halB cells imprecisely determine the future division plane, defects arise predominantly during the disk-to-rod shape remodeling. The deletion of halA in the haloarchaeon Halobacterium salinarum , whose cells are consistently rod-shaped, impacted morphogenesis but not cell division. Increased levels of halofilins enforced drastic deformations in cells devoid of the S-layer, suggesting that HalB polymers are more stable at defective S-layer lattice regions. Our results suggest that halofilins might play a significant mechanical scaffolding role in addition to possibly directing envelope synthesis.

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Halofilins as Emerging Bactofilin Families of Archaeal Cell Shape Plasticity Orchestrators

Curtis,

Escudeiro,

Mallon

et al. 2024

Preprint

Self Cite

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Bactofilins are rigid, non-polar bacterial cytoskeletal filaments that link cellular processes to specific curvatures of the cytoplasmic membrane. Although homologs of bactofilins have been identified in archaea and eukaryotes, functional studies have remained confined to bacterial systems. Here, we characterized representatives of two families of archaeal bactofilins from the pleomorphic archaeon Haloferax volcanii, halofilin A (HalA) and halofilin B (HalB). Unlike bacterial bactofilins, HalA polymerizes into polar filaments in vivo at positive membrane curvature, whereas HalB forms more static foci and accumulates in areas of local negative curvatures on the outer cell surface. Combining gene deletions, super-resolution, and single-cell microscopy showed that halofilins are critical in maintaining H. volcanii cell integrity during shape transition from disk (sessile) to rod (motile). Morphological defects in ΔhalA primarily affected rod-shaped cells from accumulating highly positive curvatures. Conversely, disk-shaped cells were exclusively affected by halB deletion, showing a decrease in positive and negative curvatures, resulting in flatter cells. Furthermore, while ΔhalA and ΔhalB cells displayed lower cell division placement precision, morphological defects arose predominantly during the disk-to-rod shape remodeling. We propose halofilins provide mechanical scaffolding, dynamically coupling the cytoplasmic membrane and the S-layer. We speculate that HalA filaments support rods under low S-layer lipidation (flexible, fast membrane diffusion). In contrast, HalB connects the S-layer to negative curvatures in disks under high lipidation levels (rigid, slow membrane diffusion).

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Identification of structural and regulatory cell-shape determinants in Haloferax volcanii

Cited by 14 publications

References 79 publications

Involvement of ArlI, ArlJ, and CirA in Archaeal Type-IV Pilin-Mediated Motility Regulation

Involvement of ArlI, ArlJ, and CirA in Archaeal Type-IV Pilin-Mediated Motility Regulation

Halofilins as emerging bactofilin families of archaeal cell shape plasticity orchestrators

Halofilins as Emerging Bactofilin Families of Archaeal Cell Shape Plasticity Orchestrators

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