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The transcription factor Ste12 mediates the regulatory role of the Tmk1 MAP kinase in mycoparasitism and vegetative hyphal fusion in the filamentous fungus Trichoderma atroviride.

Gruber S, Zeilinger S - PLoS ONE (2014)

Bottom Line: However, the transcription factors acting downstream of Tmk1 are hitherto unknown.Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases.Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

View Article: PubMed Central - PubMed

Affiliation: Research Area Biotechnology and Microbiology, Institute of Chemical Engineering, Vienna University of Technology, Wien, Austria.

ABSTRACT
Mycoparasitic species of the fungal genus Trichoderma are potent antagonists able to combat plant pathogenic fungi by direct parasitism. An essential step in this mycoparasitic fungus-fungus interaction is the detection of the fungal host followed by activation of molecular weapons in the mycoparasite by host-derived signals. The Trichoderma atroviride MAP kinase Tmk1, a homolog of yeast Fus3/Kss1, plays an essential role in regulating the mycoparasitic host attack, aerial hyphae formation and conidiation. However, the transcription factors acting downstream of Tmk1 are hitherto unknown. Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases. Deletion of the ste12 gene in T. atroviride not only resulted in reduced mycoparasitic overgrowth and lysis of host fungi but also led to loss of hyphal avoidance in the colony periphery and a severe reduction in conidial anastomosis tube formation and vegetative hyphal fusion events. The transcription of several orthologues of Neurospora crassa hyphal fusion genes was reduced upon ste12 deletion; however, the Δste12 mutant showed enhanced expression of mycoparasitism-relevant chitinolytic and proteolytic enzymes and of the cell wall integrity MAP kinase Tmk2. Based on the comparative analyses of Δste12 and Δtmk1 mutants, an essential role of the Ste12 transcriptional regulator in mediating outcomes of the Tmk1 MAPK pathway such as regulation of the mycoparasitic activity, hyphal fusion and carbon source-dependent vegetative growth is suggested. Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

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Phylogeny and transcriptional regulation of T. atroviride ste12 (A) Phylogenetic analysis of Ste12-like proteins from various filamentous fungi.Ste12 orthologues identified in the genomes of the three Trichoderma species, T. atroviride (ID 29631), Trichoderma virens (ID 75179), and Trichoderma reesei (ID 36543), and Fusarium oxysporum Ste12 (ACM80357), Magnaporthe oryzae (M. grisea) Mst12 (AF432913), Neurospora crassa PP-1 (EAA28575), Botrytis cinerea Ste12 (ACJ06644.1), Aspergillus nidulans SteA (XP_659894.1), Aspergillus fumigatus SteA (EDP51368.1), Cryptococcus neoformans Ste12 (XP_776009.1), and Saccharomyces cerevisiae Ste12 (CAX80094.1) were aligned using ClustalX and the tree constructed using neighbor-joining algorithm with 1,000 bootstraps. (B) ste12 transcription during growth of T. atroviride alone, in the mycoparasitic interaction with R. solani as host, in self-confrontation, and at contact with plain nylon fibers. Trichoderma mycelia were harvested from the interaction zone before direct contact (BC), at direct contact (C) and after contact (AC) between the two fungi. Products from RT-PCR reactions with primers targeting ste12 and tef1 (loading control) were separated by agarose gel electrophoresis.
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pone-0111636-g001: Phylogeny and transcriptional regulation of T. atroviride ste12 (A) Phylogenetic analysis of Ste12-like proteins from various filamentous fungi.Ste12 orthologues identified in the genomes of the three Trichoderma species, T. atroviride (ID 29631), Trichoderma virens (ID 75179), and Trichoderma reesei (ID 36543), and Fusarium oxysporum Ste12 (ACM80357), Magnaporthe oryzae (M. grisea) Mst12 (AF432913), Neurospora crassa PP-1 (EAA28575), Botrytis cinerea Ste12 (ACJ06644.1), Aspergillus nidulans SteA (XP_659894.1), Aspergillus fumigatus SteA (EDP51368.1), Cryptococcus neoformans Ste12 (XP_776009.1), and Saccharomyces cerevisiae Ste12 (CAX80094.1) were aligned using ClustalX and the tree constructed using neighbor-joining algorithm with 1,000 bootstraps. (B) ste12 transcription during growth of T. atroviride alone, in the mycoparasitic interaction with R. solani as host, in self-confrontation, and at contact with plain nylon fibers. Trichoderma mycelia were harvested from the interaction zone before direct contact (BC), at direct contact (C) and after contact (AC) between the two fungi. Products from RT-PCR reactions with primers targeting ste12 and tef1 (loading control) were separated by agarose gel electrophoresis.

Mentions: The T. atroviride ste12 gene (ID 29631;http://genome.jgi-psf.org/Triat2/Triat2.home.html) consists of an open reading frame of 2142-bp with two C-terminally located introns (60-bp and 63-bp) and is predicted to encode a protein of 672 amino acids. Ste12 displays considerable amino acid identity to the already functionally characterized Ste12 proteins of Fusarium oxysporum (ACM80357; 77%), Magnaporthe oryzae (M. grisea) (AF432913; 74%), Neurospora crassa (EAA28575; 65%), and Aspergillus nidulans (XP_659894; 60%) [12], [36]–[38]. Phylogenetic analysis showed Ste12 in a clade together with the orthologues from Trichoderma virens (ID 75179, http://genome.jgi-psf.org/TriviGv29_8_2/TriviGv29_8_2.home.html) and Trichoderma reesei (ID 36543, http://genome.jgi-psf.org/Trire2/Trire2.home.html), and with F. oxysporum Ste12 and M. oryzae Mst12 (Fig. 1). Similar to Ste12 proteins from other filamentous fungi, T. atroviride Ste12 contains an N-terminally located homeodomain-like (STE) motif (amino acids 54–163) which is presumed to be involved in DNA binding [39] and two distinct C-terminal C2H2 zinc finger domains (amino acids 546–582) which are distinguishing features of Ste12-like proteins of filamentous fungi [15].


The transcription factor Ste12 mediates the regulatory role of the Tmk1 MAP kinase in mycoparasitism and vegetative hyphal fusion in the filamentous fungus Trichoderma atroviride.

Gruber S, Zeilinger S - PLoS ONE (2014)

Phylogeny and transcriptional regulation of T. atroviride ste12 (A) Phylogenetic analysis of Ste12-like proteins from various filamentous fungi.Ste12 orthologues identified in the genomes of the three Trichoderma species, T. atroviride (ID 29631), Trichoderma virens (ID 75179), and Trichoderma reesei (ID 36543), and Fusarium oxysporum Ste12 (ACM80357), Magnaporthe oryzae (M. grisea) Mst12 (AF432913), Neurospora crassa PP-1 (EAA28575), Botrytis cinerea Ste12 (ACJ06644.1), Aspergillus nidulans SteA (XP_659894.1), Aspergillus fumigatus SteA (EDP51368.1), Cryptococcus neoformans Ste12 (XP_776009.1), and Saccharomyces cerevisiae Ste12 (CAX80094.1) were aligned using ClustalX and the tree constructed using neighbor-joining algorithm with 1,000 bootstraps. (B) ste12 transcription during growth of T. atroviride alone, in the mycoparasitic interaction with R. solani as host, in self-confrontation, and at contact with plain nylon fibers. Trichoderma mycelia were harvested from the interaction zone before direct contact (BC), at direct contact (C) and after contact (AC) between the two fungi. Products from RT-PCR reactions with primers targeting ste12 and tef1 (loading control) were separated by agarose gel electrophoresis.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4214791&req=5

pone-0111636-g001: Phylogeny and transcriptional regulation of T. atroviride ste12 (A) Phylogenetic analysis of Ste12-like proteins from various filamentous fungi.Ste12 orthologues identified in the genomes of the three Trichoderma species, T. atroviride (ID 29631), Trichoderma virens (ID 75179), and Trichoderma reesei (ID 36543), and Fusarium oxysporum Ste12 (ACM80357), Magnaporthe oryzae (M. grisea) Mst12 (AF432913), Neurospora crassa PP-1 (EAA28575), Botrytis cinerea Ste12 (ACJ06644.1), Aspergillus nidulans SteA (XP_659894.1), Aspergillus fumigatus SteA (EDP51368.1), Cryptococcus neoformans Ste12 (XP_776009.1), and Saccharomyces cerevisiae Ste12 (CAX80094.1) were aligned using ClustalX and the tree constructed using neighbor-joining algorithm with 1,000 bootstraps. (B) ste12 transcription during growth of T. atroviride alone, in the mycoparasitic interaction with R. solani as host, in self-confrontation, and at contact with plain nylon fibers. Trichoderma mycelia were harvested from the interaction zone before direct contact (BC), at direct contact (C) and after contact (AC) between the two fungi. Products from RT-PCR reactions with primers targeting ste12 and tef1 (loading control) were separated by agarose gel electrophoresis.
Mentions: The T. atroviride ste12 gene (ID 29631;http://genome.jgi-psf.org/Triat2/Triat2.home.html) consists of an open reading frame of 2142-bp with two C-terminally located introns (60-bp and 63-bp) and is predicted to encode a protein of 672 amino acids. Ste12 displays considerable amino acid identity to the already functionally characterized Ste12 proteins of Fusarium oxysporum (ACM80357; 77%), Magnaporthe oryzae (M. grisea) (AF432913; 74%), Neurospora crassa (EAA28575; 65%), and Aspergillus nidulans (XP_659894; 60%) [12], [36]–[38]. Phylogenetic analysis showed Ste12 in a clade together with the orthologues from Trichoderma virens (ID 75179, http://genome.jgi-psf.org/TriviGv29_8_2/TriviGv29_8_2.home.html) and Trichoderma reesei (ID 36543, http://genome.jgi-psf.org/Trire2/Trire2.home.html), and with F. oxysporum Ste12 and M. oryzae Mst12 (Fig. 1). Similar to Ste12 proteins from other filamentous fungi, T. atroviride Ste12 contains an N-terminally located homeodomain-like (STE) motif (amino acids 54–163) which is presumed to be involved in DNA binding [39] and two distinct C-terminal C2H2 zinc finger domains (amino acids 546–582) which are distinguishing features of Ste12-like proteins of filamentous fungi [15].

Bottom Line: However, the transcription factors acting downstream of Tmk1 are hitherto unknown.Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases.Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

View Article: PubMed Central - PubMed

Affiliation: Research Area Biotechnology and Microbiology, Institute of Chemical Engineering, Vienna University of Technology, Wien, Austria.

ABSTRACT
Mycoparasitic species of the fungal genus Trichoderma are potent antagonists able to combat plant pathogenic fungi by direct parasitism. An essential step in this mycoparasitic fungus-fungus interaction is the detection of the fungal host followed by activation of molecular weapons in the mycoparasite by host-derived signals. The Trichoderma atroviride MAP kinase Tmk1, a homolog of yeast Fus3/Kss1, plays an essential role in regulating the mycoparasitic host attack, aerial hyphae formation and conidiation. However, the transcription factors acting downstream of Tmk1 are hitherto unknown. Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases. Deletion of the ste12 gene in T. atroviride not only resulted in reduced mycoparasitic overgrowth and lysis of host fungi but also led to loss of hyphal avoidance in the colony periphery and a severe reduction in conidial anastomosis tube formation and vegetative hyphal fusion events. The transcription of several orthologues of Neurospora crassa hyphal fusion genes was reduced upon ste12 deletion; however, the Δste12 mutant showed enhanced expression of mycoparasitism-relevant chitinolytic and proteolytic enzymes and of the cell wall integrity MAP kinase Tmk2. Based on the comparative analyses of Δste12 and Δtmk1 mutants, an essential role of the Ste12 transcriptional regulator in mediating outcomes of the Tmk1 MAPK pathway such as regulation of the mycoparasitic activity, hyphal fusion and carbon source-dependent vegetative growth is suggested. Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

Show MeSH
Related in: MedlinePlus