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Mucor rouxii Rho1 protein; characterization and possible role in polarized growth

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Abstract

We have previously shown that protein kinase A of the medically important zygomycete Mucor rouxii participates in fungal morphology through cytoskeletal organization. As a first step towards finding the link between protein kinase A and cytoskeletal organization we here demonstrate the cloning of the Rho1 gene and the characterization of its protein product. The RHO1 protein primary sequence shows 70–85% identity with fungal RHO1 or mammalian RhoA. Two protein kinase A phosphorylation sequences in adequate context are predicted, Ser73 and Ser135. The peptide IRRNSQKFV, containing Ser135 proved to be a good substrate for M. rouxii protein kinase A catalytic subunit. The over-expressed Rho1 fully complements a Saccharomyces cerevisiae null mutant. The endogenous protein was identified by western blot against a developed antibody and by ADP-ribosylation. Localization in germlings was visualized by immunofluorescence; the protein was localized in patches in the mother cell surface and excluded from the germ tube. Measurement of Rho1 expression during germination indicates that Rho1, at both the mRNA and protein levels, correlates with differentiation and not with growth. Rho1 has been shown to be the regulatory protein of the β-1,3-glucan synthase complex in fungi in which β-1,3-glucans are major components of the cell wall. Even though glucans have not been detected in zygomycetes, caspofungin, an echinochandin known to be an inhibitor of β-1,3-glucan synthase complex, is shown here to have a negative effect on growth and to produce an alteration on morphology when added to M. rouxii growth culture medium. This result has an important impact on the possible participation of β-1,3-glucans on the regulation of morphology of zygomycetes.

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Abbreviations

Mr:

Mucor rouxii

PKA:

protein kinase A

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Acknowledgements

This study was supported by grants from the University of Buenos Aires, grant X-270, from CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas) grant PIP 5239 and from ANPCYT (Agencia Nacional de Promoción Científica y Tecnológica) grant PICT 01-08793.

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  1. Silvia Argimón

    Present address: Aberdeen Fungal Group, University of Aberdeen, Aberdeen, AB25 2ZD, UK

Authors and Affiliations

  1. Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Ciudad Universitaria, Pabellón 2, Piso 4, 1428, Buenos Aires, Argentina

    Silvia Argimón, Fiorella Galello, Elba Pereyra, Silvia Rossi & Silvia Moreno

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  1. Silvia Argimón
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  2. Fiorella Galello
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Correspondence to Silvia Moreno.

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Argimón, S., Galello, F., Pereyra, E. et al. Mucor rouxii Rho1 protein; characterization and possible role in polarized growth. Antonie van Leeuwenhoek 91, 237–251 (2007). https://doi.org/10.1007/s10482-006-9113-7

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