Noncanonical Gβ Gib2 Is a Scaffolding Protein Promoting cAMP Signaling through Functions of Ras1 and Cac1 Proteins in Cryptococcus neoformans

Wang, Yanli; Shen, Guanghui; Gong, Jinjun; Shen, Danyu; Whittington, Amy; Jiang, Qing; Treloar, Joshua; Boisvert, Scott; Cai, Yang; Wang, Ping

doi:10.1074/jbc.m113.537183

Cited by 19 publications

(75 citation statements)

References 60 publications

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“…Among the groups, ribosomal subunits, ribosomal biogenesis components, intracellular trafficking, and signaling components are included in the top of the eight groups. Together with our previous findings, this new round of findings further established crucial function of Gib2 in growth and pathogenicity [9]. Significantly, these findings also suggest that Gib2 represents an important therapeutic target.…”

Section: Discussionsupporting

confidence: 85%

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Comparative proteomic analysis of Gib2 validating its adaptor function in Cryptococcus neoformans

et al. 2017

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Cryptococcus neoformans causes often-fatal fungal meningoencephalitis in immunocompromised individuals. While the exact disease mechanisms remain elusive, signal transduction pathways mediated by key elements such as G-protein α subunit Gpa1, small GTPase Ras1, and atypical Gβ-like/RACK1 protein Gib2 are known to play important roles in C. neoformans virulence. Gib2 is important for normal growth, differentiation, and pathogenicity, and it also positively regulates cAMP levels in conjunction with Gpa1. Interestingly, Gib2 displays a scaffold protein property by interacting with a wide variety of cellular proteins. To explore Gib2 global regulatory functions, we performed two-dimensional differential gel electrophoresis (DIGE) analysis and found that GIB2 disruption results in an increased expression of 304 protein spots (43.4%) and a decreased expression of 396 protein spots (56.6%). Analysis of 96 proteins whose expression changes were deemed significant (≥ +/- 1.5- fold) revealed that 75 proteins belong to at least 12 functional protein groups. Among them, eight groups have the statistical stringency of p ≤ 0.05, and four groups, including Hsp70/71 heat shock protein homologs and ribosomal proteins, survived the Bonferroni correction. This finding is consistent with earlier established roles for the human Gβ-like/RACK1 and the budding yeast Saccharomyces cerevisiae Asc1. It suggests that Gib2 could also be part of the complex affecting ribosomal biogenesis and protein translation in C. neoformans. Since eukaryotic Hsp70/71 proteins are involved in the facilitation of nascent protein folding, processing, and protection of cells against stress, we also propose that Gib2-regulated stress responses are linked to fungal virulence. Collectively, our study supports a conserved role of Gβ-like/RACK/Gib2 proteins in the essential cellular process of ribosomal biogenesis and protein translation. Our study also highlights a multifaceted regulatory role of Gib2 in the growth and pathogenicity of C. neoformans.

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Section: Discussionsupporting

confidence: 85%

“…We also found that Gib2 interacts with approximately 50 proteins, including the protein kinase C homolog Pkc1, the endocytic adaptor protein Cin1, and several ribosomal subunits [9,10]. To further understand the global regulatory role of Gib2, we performed DIGE on protein extracts from the gib2 mutant and the wild-type strains of C .…”

Section: Resultsmentioning

confidence: 99%

Comparative proteomic analysis of Gib2 validating its adaptor function in Cryptococcus neoformans

et al. 2017

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“…Also, a canonical G␤ protein capable of binding to Gpa1 was not found, but the noncanonical G␤ protein Gib2 proved to form a heterotrimeric complex with Gpa1 and Gpg1 or Gpg2 (38). In addition, Gib2 functions as a scaffolding protein that positively regulates cAMP levels through novel functions involving the Ras1 and Cac1 proteins (40). The current identification of the Ric8 protein provides another mechanistic explanation of how Gpa1 functions without a classic G␤ subunit.…”

Section: Discussionmentioning

confidence: 96%

“…Curiously, a classic G protein ␤ subunit capable of coupling with Gpa1 was not found, leading to propositions that Gpa1 may function as a monomeric G protein whose activation depends on GEF or that certain proteins may function as a bona fide G␤. We have since characterized a Gpa1 binding protein (Gib2) as a noncanonical G␤ protein forming a heterotrimeric G protein complex with Gpa1 (38,40). To identify additional proteins capable of binding and potentially activating Gpa1, we used the Y2H screen to examine candidate proteins that included a Ric8 homolog and three members of the Rho family GTPases (Cdc42, Cdc420, and Ran1) (unpublished observations).…”

Section: Resultsmentioning

confidence: 99%

A Ric8/Synembryn Homolog Promotes Gpa1 and Gpa2 Activation To Respectively Regulate Cyclic AMP and Pheromone Signaling in Cryptococcus neoformans

2014

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The G protein ␣ subunits Gpa1, Gpa2, and Gpa3 mediate signal transduction and are important in the growth and virulence of Cryptococcus neoformans. To understand how Gpa1 functions without a conventional G␤ subunit, we characterized a resistance to inhibitors of cholinesterase 8 (Ric8) homolog from C. neoformans, which shares amino acid sequence homology with other Ric8 proteins that exhibit guanine nucleotide exchange factor (GEF) activity toward G␣. We found that the ric8 mutant was reduced in capsule size and melanin formation, which could be suppressed by cyclic AMP (cAMP) supplementation or by introducing the activated GPA1 Q284L allele. Consistent with the fact that Ric8 participates in cAMP signaling to regulate virulence, the ric8 mutant was attenuated in virulence toward mice. Interestingly, disruption of RIC8 also resulted in opposing effects on pheromone signaling, as the ric8 mutant showed reduced mating but an enhanced ability to induce the pheromone response in the mating partner. To identify Ric8 functional mechanisms, we examined the interactions between Ric8 and the three G␣ proteins. Ric8 interacted with Gpa1 and Gpa2, but not Gpa3. The presence of Gpa1 Q284L negatively affected its interaction with Ric8, whereas the activated Gpa2 Q203L allele abolished the interaction. Collectively, these findings suggest that Ric8 functions as a GEF to facilitate the activation of Gpa1-cAMP signaling and to promote Gpa2, affecting mating efficiency. Our study highlights the distinct and conserved characteristics associated with G protein signaling and contributes to our overall understanding of how G protein ␣ subunits function with or without a canonical G␤ partner in C. neoformans.

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“…Like the conventional Gβ sub-unit (Gpb1), Gib2 interacts with Gα (Gpa1) and Gγ (Gpg1 and Gpg2) to form a complex (Fig. 1) (Palmer et al ., 2006; Wang et al ., 2014; Ero et al ., 2015). Gpa1 is a key player in the cAMP/PKA signaling pathway which, as mentioned above, contributes to diverse phenotypes such as virulence, capsule formation, melanization, mating, titan cell formation, and secretion of specific enzymes such as protease and urease (Alspaugh et al ., 1997; D’Souza et al ., 2001; Alspaugh et al ., 2002; Kronstad et al ., 2011a).…”

Section: Association Of Rack1-like Proteins With the Camp/pka Pathwaymentioning

confidence: 99%

The cAMP/protein kinase A signaling pathway in pathogenic basidiomycete fungi: Connections with iron homeostasis

2015

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A number of pathogenic species of basidiomycete fungi are either life-threatening pathogens of humans or major economic pests for crop production. Sensing the host is a key aspect of pathogen proliferation during disease, and signal transduction pathways are critically important for detecting environmental conditions and facilitating adaptation. This review focuses on the contributions of the cAMP/protein kinase A (PKA) signaling pathway in Cryptococcus neoformans, a species that causes meningitis in humans, and Ustilago maydis, a model phytopathogen that causes a smut disease on maize. Environmental sensing by the cAMP/PKA pathway regulates the production of key virulence traits in C. neoformans including the polysaccharide capsule and melanin. For U. maydis, the pathway controls the dimorphic transition from budding growth to the filamentous cell type required for proliferation in plant tissue. We discuss recent advances in identifying new components of the cAMP/PKA pathway in these pathogens and highlight an emerging theme that pathway signaling influences iron acquisition.

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Noncanonical Gβ Gib2 Is a Scaffolding Protein Promoting cAMP Signaling through Functions of Ras1 and Cac1 Proteins in Cryptococcus neoformans

Cited by 19 publications

References 60 publications

Comparative proteomic analysis of Gib2 validating its adaptor function in Cryptococcus neoformans

Comparative proteomic analysis of Gib2 validating its adaptor function in Cryptococcus neoformans

A Ric8/Synembryn Homolog Promotes Gpa1 and Gpa2 Activation To Respectively Regulate Cyclic AMP and Pheromone Signaling in Cryptococcus neoformans

The cAMP/protein kinase A signaling pathway in pathogenic basidiomycete fungi: Connections with iron homeostasis

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