Xanthomonas citri MinC Oscillates from Pole to Pole to Ensure Proper Cell Division and Shape

Abstract

Xanthomonas citri (Xac) is the causal agent of citrus canker, a disease that affects citrus crops and causes economic impact worldwide. To further characterize cell division in this plant pathogen, we investigated the role of the protein MinC in cell division, chromosome segregation, and peptidoglycan incorporation by deleting the gene minC using allele exchange. Xac with minC deleted exhibited the classic Δmin phenotype observed in other bacteria deleted for min components: minicells and short filamentation. In addition we noticed the formation of branches, which is similar to what was previously described for Escherichia coli deleted for either min or for several low molecular weight penicillin-binding proteins (PBPs). The branching phenotype was medium dependent and probably linked to gluconeogenic growth. We complemented the minC gene by integrating gfp-minC into the amy locus. Xac complemented strains displayed a wild-type phenotype. In addition, GFP-MinC oscillated from pole to pole, similar to MinCD oscillations observed in E. coli and more recently in Synechococcus elongatus. Further investigation of the branching phenotype revealed that in branching cells nucleoid organization, divisome formation and peptidoglycan incorporation were disrupted.

mCherry fusions enable the subcellular localization of periplasmic and cytoplasmic proteins in Xanthomonas sp.

“…This result demonstrates that the integration of the plasmid pMAJIIc into the amy locus did not affect the ability of X. citri to cause citrus canker. A similar result was found by Lorenzoni et al [22] with the plasmid pGCD21, which was used as backbone for pMAJIIc.…”

“…The PCR-amplified fragment was cleaned up by phenol/chloroform extraction and ethanol-precipitation [20] and then digested with XhoI and NotI (New England Biolabs). The digested fragment was also cleaned up by phenol/chloroform extraction and ethanol-precipitation and then ligated (T4 DNA ligase) into pGCD21 plasmid [22] cleaved with XhoI and NotI (removes gfpmut1 sequence), resulting in plasmid pMAJIc (GenBank MT119764). pGCD21 plasmid harbors a fragment of X. citri amy gene (bases 106-912), which allows the integration into X. citri chromosome by disrupting amy [22].…”

“…The digested fragment was also cleaned up by phenol/chloroform extraction and ethanol-precipitation and then ligated (T4 DNA ligase) into pGCD21 plasmid [22] cleaved with XhoI and NotI (removes gfpmut1 sequence), resulting in plasmid pMAJIc (GenBank MT119764). pGCD21 plasmid harbors a fragment of X. citri amy gene (bases 106-912), which allows the integration into X. citri chromosome by disrupting amy [22]. A complementary pair of oligonucleotides containing the restriction sites NdeI, KpnI and SalI were designed (primers pMAJ1_PL_F and pMAJ1_PL_R- Table 2) and subjected to hybridization, and the resulting double-stranded DNA (owning a 5' NheI and a 5' XhoI overhang cohesive ends) was ligated into pMAJIc upstream of the mCherry sequence (within the NheI/XhoI sites), giving rise to pMAJIIc plasmid (GenBank MT119765), which has a polylinker region containing the unique restriction sites NheI, NdeI, KpnI, SalI and XhoI (Fig 1).…”

The Min Oscillator Defines Sites of Asymmetric Cell Division in Cyanobacteria during Stress Recovery

“…Oscillatory Min systems are often found in rod-shaped bacterial species such as E. coli (de Boer et al, 1989), V. parahaemolyticus (Muraleedharan et al, 2018), Xanthomonas citri (Lorenzoni et al, 2017), and S. elongatus, all of which can undergo filamentation in response to stress or other changes in growth conditions. Other bacteria that undergo different morphological changes in response to stress, such as Bacillus subtilis, may also possess a Min system that is nonetheless non-dynamic (Jamroskovic et al, 2012).…”

A cell length‐dependent transition in MinD‐dynamics promotes a switch in division‐site placement and preservation of proliferating elongated Vibrio parahaemolyticus swarmer cells

“…The placement of the Z-ring has been shown to be regulated by proteins belonging to the ParA-like family of Walker-type ATPases in many organisms such as Escherichia coli (de Boer et al, 1989;Raskin and de Boer, 1999), Bacillus subtilis (Marston et al, 1998), Caulobacter crescentus (Thanbichler and Shapiro, 2006;Kiekebusch et al, 2012), Corynebacterium glutamicum (Donovan et al, 2010), Myxococcus xanthus (Treuner-Lange et al, 2013;Schumacher et al, 2017), Xanthomonas citri (Lorenzoni et al, 2017) and Vibrio cholerae (Galli et al, 2016). One such example of a ParA-like protein is MinD, a part of the Min-system.…”