Genetic involvement of a cAMP-dependent protein kinase in a G protein signaling pathway regulating morphological and chemical transitions in Aspergillus nidulans.

In the filamentous fungus Aspergillus nidulans, a heterotrimeric G protein alpha-subunit and an RGS domain protein, encoded by fadA and flbA, respectively, regulate production of the carcinogenic metabolite sterigmatocystin (ST) and asexual spores (i.e., conidia). We investigated the genetic involvement of the cAMP-dependent protein kinase catalytic subunit (PkaA), a potential downstream target of FadA activity, in ST production and conidiation. Relative to wild type, sporulation was decreased in the pkaA overexpression strain but was not totally absent, as occurs in DeltaflbA or fadA(G42R) (fadA-dominant active) strains. Deletion of pkaA resulted in a hyper-conidiating strain with limited radial growth. This phenotype was epistatic to mutation in flbA or fadA; the double mutants DeltapkaA; DeltaflbA and DeltapkaA; fadA(G42R) recovered sporulation and their radial growth was severely restricted. PkaA overexpression also negatively regulated AflR, the ST biosynthesis-specific transcription factor, both transcriptionally and post-transcriptionally. Deletion of pkaA restored ST production in the DeltaflbA background but not in the fadA(G42R) background. These data provide genetic evidence that the FlbA/FadA signaling pathway regulating ST production and morphological development is partially mediated through PkaA.

[1]  F. Sanger,et al.  DNA sequencing with chain-terminating inhibitors. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[2]  Bee-Na Lee,et al.  Overexpression of fIbA, an early regulator of Aspergillus asexual sporulation, leads to activation of brIA and premature initiation of development , 1994, Molecular microbiology.

[3]  M. Forte,et al.  Drosophila stimulatory G protein alpha subunit activates mammalian adenylyl cyclase but interacts poorly with mammalian receptors: implications for receptor-G protein interaction. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[4]  S. Gold,et al.  cAMP regulates morphogenesis in the fungal pathogen Ustilago maydis. , 1994, Genes & development.

[5]  G. Fink,et al.  The three yeast A kinases have specific signaling functions in pseudohyphal growth. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[6]  C. Anjard,et al.  The catalytic subunit of Dictyostelium cAMP-dependent protein kinase -- role of the N-terminal domain and of the C-terminal residues in catalytic activity and stability. , 1997, European journal of biochemistry.

[7]  L S Robertson,et al.  The yeast A kinases differentially regulate iron uptake and respiratory function. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[8]  W. Timberlake,et al.  brlA is necessary and sufficient to direct conidiophore development in aspergillus nidulans , 1988, Cell.

[9]  T. Hunter,et al.  The eukaryotic protein kinase superfamily: kinase (catalytic) domain structure and classification 1 , 1995, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[10]  Divergent cAMP Signaling Pathways Regulate Growth and Pathogenesis in the Rice Blast Fungus Magnaporthe grisea , 1998, Plant Cell.

[11]  Jae-Hyuk Yu,et al.  The Aspergillus FlbA RGS domain protein antagonizes G protein signaling to block proliferation and allow development. , 1996, The EMBO journal.

[12]  T. C. Nesbitt,et al.  Twenty-five coregulated transcripts define a sterigmatocystin gene cluster in Aspergillus nidulans. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[13]  T. Som,et al.  Developmental decisions in Aspergillus nidulans are modulated by Ras activity , 1994, Molecular and cellular biology.

[14]  M. Bölker,et al.  G proteins in Ustilago maydis: transmission of multiple signals? , 1997, The EMBO journal.

[15]  F. Banuett.  Signalling in the Yeasts: An Informational Cascade with Links to the Filamentous Fungi , 1998, Microbiology and Molecular Biology Reviews.

[16]  M. M. Bradford.  A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. , 1976, Analytical biochemistry.

[17]  M. Farquhar,et al.  RGS proteins: more than just GAPs for heterotrimeric G proteins. , 1999 .

[18]  J. Ramírez,et al.  Isolation of a gene encoding a G protein α subunit involved in the regulation of cAMP levels in the yeast Kluyveromyces lactis , 1996, Yeast.

[19]  T. Hunter.  Protein-Tryosine Kinases , 1985 .

[20]  R. Dean,et al.  G Protein α Subunit Genes Control Growth, Development, and Pathogenicity of Magnaporthe grisea , 1997 .

[21]  M. Dickman,et al.  Colletotrichum trifolii mutants disrupted in the catalytic subunit of cAMP-dependent protein kinase are nonpathogenic. , 1999, Molecular plant-microbe interactions : MPMI.

[22]  R. Kahmann,et al.  Crosstalk between cAMP and pheromone signalling pathways in Ustilago maydis , 1998, Molecular and General Genetics MGG.

[23]  G. Choi,et al.  Extensive alteration of fungal gene transcript accumulation and elevation of G-protein-regulated cAMP levels by a virulence-attenuating hypovirus. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[24]  J. D. de Winde,et al.  Novel sensing mechanisms and targets for the cAMP–protein kinase A pathway in the yeast Saccharomyces cerevisiae , 1999, Molecular microbiology.

[25]  R. Dean,et al.  The cAMP-dependent protein kinase catalytic subunit is required for appressorium formation and pathogenesis by the rice blast pathogen Magnaporthe grisea. , 1995, The Plant cell.

[26]  G. Fink,et al.  MAP kinase and cAMP filamentation signaling pathways converge on the unusually large promoter of the yeast FLO11 gene , 1999, The EMBO journal.

[27]  J. Kronstad,et al.  Signaling via cAMP in fungi: interconnections with mitogen-activated protein kinase pathways , 1998, Archives of Microbiology.

[28]  N. Keller,et al.  Aspergillus nidulans mutants defective in stc gene cluster regulation. , 1999, Genetics.

[29]  J. Heitman,et al.  Cryptococcus neoformans mating and virulence are regulated by the G-protein alpha subunit GPA1 and cAMP. , 1997, Genes & development.

[30]  K. Borkovich,et al.  Positive Regulation of Adenylyl Cyclase Activity by a GαiHomolog inNeurospora crassa , 1999 .

[31]  M. Yamamoto,et al.  Characterization of a fission yeast gene, gpa2, that encodes a G alpha subunit involved in the monitoring of nutrition. , 1992, Genes & development.

[32]  J. Thorner,et al.  Model systems for the study of seven-transmembrane-segment receptors. , 1991, Annual review of biochemistry.

[33]  Michael Wigler,et al.  Three different genes in S. cerevisiae encode the catalytic subunits of the cAMP-dependent protein kinase , 1987, Cell.

[34]  Jae-Hyuk Yu,et al.  Aspergillus sporulation and mycotoxin production both require inactivation of the FadA Gα protein‐dependent signaling pathway , 1997, The EMBO journal.

[35]  R. Dean,et al.  The adenylate cyclase gene MAC1 of Magnaporthe grisea controls appressorium formation and other aspects of growth and development. , 1997, The Plant cell.

[36]  J. Kronstad,et al.  Identification of a cAMP-dependent protein kinase catalytic subunit required for virulence and morphogenesis in Ustilago maydis. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[37]  N. Keller,et al.  Sequence‐specific binding by Aspergillus nidulans AflR, a C6 zinc cluster protein regulating mycotoxin biosynthesis , 1998, Molecular microbiology.

[38]  J. Hamer,et al.  The CPKA gene of Magnaporthe grisea is essential for appressorial penetration , 1997 .

[39]  R. Plasterk,et al.  G protein hyperactivation of the Caenorhabditis elegans adenylyl cyclase SGS‐1 induces neuronal degeneration , 1998, The EMBO journal.

[40]  Jae-Hyuk Yu,et al.  The Aspergillus nidulans sfaD gene encodes a G protein β subunit that is required for normal growth and repression of sporulation , 1999, The EMBO journal.