Impact of c-di-GMP on the Extracellular Proteome of Rhizobium etli
Por:
Lorite MJ, Casas-Román A, Girard L, Encarnación S, Díaz-Garrido N, Badia-Palacin J, Baldoma L, Pérez-Mendoza D and Sanjuán J
Publicada:
1 ene 2023
Ahead of Print:
26 dic 2022
Resumen:
Simple Summary The second messenger cyclic diguanylate (c-di-GMP, cdG)
is a bacterial lifestyle-switch molecule, well known for its role in
biofilm formation. Extracellular biofilm matrix components include
diverse biopolymers such as polysaccharides, nucleic acids, proteins and
lipids. The production and/or secretion of many these matrix biopolymers
can be directly or indirectly regulated by cdG. We studied the
extracellular proteome of a Rhizobium etli strain expressing
artificially high levels of intracellular cdG. We found that, in
addition to promoting the secretion of various extracellular proteins
likely involved in adhesion and biofilm formation, high cdG levels also
promote the export of cytoplasmic proteins (ECP) to the cell exterior.
Intriguingly, most these cytoplasmic proteins have been previously
described as moonlighting or multifunctional proteins in other
organisms, often found extracellularly or at the bacterial cell surface.
We obtained evidence that this ECP may involve an active process that
would be enhanced by cdG. For a typical cytoplasmic protein,
glyceraldehyde 3-phosphate dehydrogenase (Gap), we also observed that
cdG increases the number of extracellular Gap proteoforms that can be
separated by two-dimensional gel electrophoresis. The results suggest
that cdG promotes the active exportation of cytoplasmic proteins through
yet unknown mechanisms involving the post-translational modification of
proteins. Extracellular matrix components of bacterial biofilms include
biopolymers such as polysaccharides, nucleic acids and proteins. Similar
to polysaccharides, the secretion of adhesins and other matrix proteins
can be regulated by the second messenger cyclic diguanylate (cdG). We
have performed quantitative proteomics to determine the extracellular
protein contents of a Rhizobium etli strain expressing high cdG
intracellular levels. cdG promoted the exportation of proteins that
likely participate in adhesion and biofilm formation: the rhizobial
adhesion protein RapA and two previously undescribed likely adhesins,
along with flagellins. Unexpectedly, cdG also promoted the selective
exportation of cytoplasmic proteins. Nearly 50% of these cytoplasmic
proteins have been previously described as moonlighting or candidate
moonlighting proteins in other organisms, often found extracellularly.
Western blot assays confirmed cdG-promoted export of two of these
cytoplasmic proteins, the translation elongation factor (EF-Tu) and
glyceraldehyde 3-phosphate dehydrogenase (Gap). Transmission Electron
Microscopy immunolabeling located the Gap protein in the cytoplasm but
was also associated with cell membranes and extracellularly, indicative
of an active process of exportation that would be enhanced by cdG. We
also obtained evidence that cdG increases the number of extracellular
Gap proteoforms, suggesting a link between cdG, the post-translational
modification and the export of cytoplasmic proteins.
Filiaciones:
Lorite MJ:
Department of Soil and Plant Microbiology, Estación Experimental del Zaidín, CSIC, 18008 Granada, Spain
Casas-Román A:
Department of Soil and Plant Microbiology, Estación Experimental del Zaidín, CSIC, 18008 Granada, Spain
Girard L:
Centro de Ciencias Genómicas (CCG), Universidad Nacional Autónoma de México (UNAM), Cuernavaca 62210, Morelos, Mexico
Encarnación S:
Centro de Ciencias Genómicas (CCG), Universidad Nacional Autónoma de México (UNAM), Cuernavaca 62210, Morelos, Mexico
Díaz-Garrido N:
Secció de Bioquímica i Biologia Molecular, Departament de Bioquímica i Fisiologia, Facultat de Farmàcia i Ciències de l'Alimentació, Universitat de Barcelona, 08028 Barcelona, Spain
Badia-Palacin J:
Secció de Bioquímica i Biologia Molecular, Departament de Bioquímica i Fisiologia, Facultat de Farmàcia i Ciències de l'Alimentació, Universitat de Barcelona, 08028 Barcelona, Spain
Institut de Biomedicina de la Universitat de Barcelona (IBUB), Institut de Recerca Sant Joan de Déu (IRSJD), 08028 Barcelona, Spain
Baldoma L:
Secció de Bioquímica i Biologia Molecular, Departament de Bioquímica i Fisiologia, Facultat de Farmàcia i Ciències de l'Alimentació, Universitat de Barcelona, 08028 Barcelona, Spain
Pérez-Mendoza D:
Department of Soil and Plant Microbiology, Estación Experimental del Zaidín, CSIC, 18008 Granada, Spain
Sanjuán J:
Department of Soil and Plant Microbiology, Estación Experimental del Zaidín, CSIC, 18008 Granada, Spain
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