Phosphorylation of a novel cytoskeletal protein (RsmP) regulates rod-shaped morphology in Corynebacterium glutamicum.
Corynebacteria grow by wall extension at the cell poles, with DivIVA being an essential protein orchestrating cell elongation and morphogenesis. DivIVA is considered a scaffolding protein able to recruit other proteins and enzymes involved in polar peptidoglycan biosynthesis. Partial depletion of DivIVA induced overexpression of cg3264, a previously uncharacterized gene that encodes a novel coiled coil-rich protein specific for corynebacteria and a few other actinomycetes. By partial depletion and overexpression of Cg3264, we demonstrated that this protein is an essential cytoskeletal element needed for maintenance of the rod-shaped morphology of Corynebacterium glutamicum, and it was therefore renamed RsmP (rod-shaped morphology protein). RsmP forms long polymers in vitro in the absence of any cofactors, thus resembling eukaryotic intermediate filaments. We also investigated whether RsmP could be regulated post-translationally by phosphorylation, like eukaryotic intermediate filaments. RsmP was phosphorylated in vitro by the PknA protein kinase and to a lesser extent by PknL. A mass spectrometric analysis indicated that phosphorylation exclusively occurred on a serine (Ser-6) and two threonine (Thr-168 and Thr-211) residues. We confirmed that mutagenesis to alanine (phosphoablative protein) totally abolished PknA-dependent phosphorylation of RsmP. Interestingly, when the three residues were converted to aspartic acid, the phosphomimetic protein accumulated at the cell poles instead of making filaments along the cell, as observed for the native or phosphoablative RsmP proteins, indicating that phosphorylation of RsmP is necessary for directing cell growth at the cell poles.
Fiuza
Maria
M
Departamento de Biología Molecular, Area de Microbiología, Facultad de Biología, Universidad de León, León 24071, Spain.
Letek
Michal
M
Leiba
Jade
J
Villadangos
Almudena F
AF
Vaquera
José
J
Zanella-Cléon
Isabelle
I
Mateos
Luís M
LM
Molle
Virginie
V
Gil
José A
JA
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D048230:Corynebacterium glutamicum / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure*; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D019143:Evolution, Molecular; D015964:Gene Expression Regulation, Bacterial; D016680:Genome, Bacterial / Q000235:genetics; D046529:Microscopy, Electron, Transmission; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
D001426:Bacterial Proteins; D003598:Cytoskeletal Proteins