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C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus

RasA is a major regulator of fungal morphogenesis and virulence in Aspergillus fumigatus. The proper localization of RasA to the plasma membrane is essential for the formation of invasive hyphae during infection. In yeast, the localization of Ras2p to the plasma membrane is orchestrated by several p...

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Autores principales: Al Abdallah, Qusai, Martin-Vicente, Adela, Souza, Ana Camila Oliveira, Ge, Wenbo, Fortwendel, Jarrod R.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5879109/
https://www.ncbi.nlm.nih.gov/pubmed/29632525
http://dx.doi.org/10.3389/fmicb.2018.00562
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author Al Abdallah, Qusai
Martin-Vicente, Adela
Souza, Ana Camila Oliveira
Ge, Wenbo
Fortwendel, Jarrod R.
author_facet Al Abdallah, Qusai
Martin-Vicente, Adela
Souza, Ana Camila Oliveira
Ge, Wenbo
Fortwendel, Jarrod R.
author_sort Al Abdallah, Qusai
collection PubMed
description RasA is a major regulator of fungal morphogenesis and virulence in Aspergillus fumigatus. The proper localization of RasA to the plasma membrane is essential for the formation of invasive hyphae during infection. In yeast, the localization of Ras2p to the plasma membrane is orchestrated by several post-translational modifications (PTM) at the C-terminal CAAX box that are thought to occur in sequential order. These PTMs include: (1) CAAX motif farnesylation by the farnesyltransferase complex composed of Ram1p and Ram2p; (2) proteolysis of the -AAX residues by Rce1p or Ste24p; (3) methylation of the remaining prenylated cysteine residue by Ste14p, and; (4) palmitoylation at a single conserved cysteine residue mediated by the Erf2p/Erf4p palmitoyltransferase. We previously reported that homologs of each RasA PTM enzyme are conserved in A. fumigatus. Additionally, we delineated a major role for protein farnesylation in A. fumigatus growth and virulence. In this work, we characterize the post-prenylation processing enzymes of RasA in A. fumigatus. The genes encoding the RasA post-prenylation enzymes were first deleted and examined for their roles in growth and regulation of RasA. Only when strains lacked cppB, the A. fumigatus homologue of yeast RCE1, there was a significant reduction in fungal growth and conidial germination. In addition, cppB-deletion mutants displayed hypersensitivity to the cell wall-perturbing agents Calcofluor White and Congo Red and the cell wall biosynthesis inhibitor Caspofungin. In contrast to the previously published data in yeast, the deletion of post-prenylation modifying enzymes did not alter the plasma membrane localization or activation of RasA. To delineate the molecular mechanisms underlying these differences, we investigated the interplay between dual-palmitoylation of the RasA hypervariable region and CAAX proteolysis for stabilization of RasA at the plasma membrane. Our data indicate that, in the absence of proper CAAX proteolysis, RasA accumulation at the plasma membrane is stabilized by dual palmitoyl groups on the dual cysteine residues. Therefore, we conclude CAAX proteolysis and dual-palmitoylation of the hypervariable region is important for maintaining a stable attachment association of RasA with the plasma membrane to support optimal fungal growth and development.
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spelling pubmed-58791092018-04-09 C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus Al Abdallah, Qusai Martin-Vicente, Adela Souza, Ana Camila Oliveira Ge, Wenbo Fortwendel, Jarrod R. Front Microbiol Microbiology RasA is a major regulator of fungal morphogenesis and virulence in Aspergillus fumigatus. The proper localization of RasA to the plasma membrane is essential for the formation of invasive hyphae during infection. In yeast, the localization of Ras2p to the plasma membrane is orchestrated by several post-translational modifications (PTM) at the C-terminal CAAX box that are thought to occur in sequential order. These PTMs include: (1) CAAX motif farnesylation by the farnesyltransferase complex composed of Ram1p and Ram2p; (2) proteolysis of the -AAX residues by Rce1p or Ste24p; (3) methylation of the remaining prenylated cysteine residue by Ste14p, and; (4) palmitoylation at a single conserved cysteine residue mediated by the Erf2p/Erf4p palmitoyltransferase. We previously reported that homologs of each RasA PTM enzyme are conserved in A. fumigatus. Additionally, we delineated a major role for protein farnesylation in A. fumigatus growth and virulence. In this work, we characterize the post-prenylation processing enzymes of RasA in A. fumigatus. The genes encoding the RasA post-prenylation enzymes were first deleted and examined for their roles in growth and regulation of RasA. Only when strains lacked cppB, the A. fumigatus homologue of yeast RCE1, there was a significant reduction in fungal growth and conidial germination. In addition, cppB-deletion mutants displayed hypersensitivity to the cell wall-perturbing agents Calcofluor White and Congo Red and the cell wall biosynthesis inhibitor Caspofungin. In contrast to the previously published data in yeast, the deletion of post-prenylation modifying enzymes did not alter the plasma membrane localization or activation of RasA. To delineate the molecular mechanisms underlying these differences, we investigated the interplay between dual-palmitoylation of the RasA hypervariable region and CAAX proteolysis for stabilization of RasA at the plasma membrane. Our data indicate that, in the absence of proper CAAX proteolysis, RasA accumulation at the plasma membrane is stabilized by dual palmitoyl groups on the dual cysteine residues. Therefore, we conclude CAAX proteolysis and dual-palmitoylation of the hypervariable region is important for maintaining a stable attachment association of RasA with the plasma membrane to support optimal fungal growth and development. Frontiers Media S.A. 2018-03-26 /pmc/articles/PMC5879109/ /pubmed/29632525 http://dx.doi.org/10.3389/fmicb.2018.00562 Text en Copyright © 2018 Al Abdallah, Martin-Vicente, Souza, Ge and Fortwendel. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Al Abdallah, Qusai
Martin-Vicente, Adela
Souza, Ana Camila Oliveira
Ge, Wenbo
Fortwendel, Jarrod R.
C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus
title C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus
title_full C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus
title_fullStr C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus
title_full_unstemmed C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus
title_short C-terminus Proteolysis and Palmitoylation Cooperate for Optimal Plasma Membrane Localization of RasA in Aspergillus fumigatus
title_sort c-terminus proteolysis and palmitoylation cooperate for optimal plasma membrane localization of rasa in aspergillus fumigatus
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5879109/
https://www.ncbi.nlm.nih.gov/pubmed/29632525
http://dx.doi.org/10.3389/fmicb.2018.00562
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