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Master's Dissertation
DOI
10.11606/D.17.2019.tde-20112018-114017
Document
Author
Full name
Cláudia Batista Carraro
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
Ribeirão Preto, 2018
Supervisor
Committee
Silva, Roberto do Nascimento (President)
Braga, Gilberto Ubida Leite
Freitas, Fernanda Zanolli
Gomes, Marcelo Damario
Title in Portuguese
Identificação de alvos de fosforilação de MAPK em Trichoderma reesei através de fosfoproteômica durante a produção de celulases
Keywords in Portuguese
Celulases
Fosfoproteômica
MAPK
Trichoderma reesei
Vias de sinalização
Abstract in Portuguese
O fungo filamentoso Trichoderma reesei é uma espécie de grande importância biotecnológica no que tange a degradação de biomassa lignocelulósica para a produção de bioetanol em larga escala. Seu sistema de enzimas celulolíticas é muito eficiente, apesar de ser possuir poucas celulases, sugerindo que o controle dessas enzimas vai além da regulação transcricional. Assim, neste trabalho nós obtivemos o perfil fosfoproteômico de T. reesei cultivado em glicose e bagaço de cana-de-açúcar a partir de análise fosfoproteômica LC-MS/MS por spectral counting. A comparação entre os perfis de fosfoproteínas obtidos das linhagens parental QM6a de T. reesei e dos mutantes knockout para TMK1 e TMK2 permitiu a demonstração de que essas MAPK agem de maneira interconectada com outras vias de transdução de sinal na célula, especialmente a via de TOR e de AMPc-PKA, para regulação da produção de celulases. Além disso, também demonstramos a regulação da resposta a estresse celular por TMK2, e o papel da fosforilação no controle direto de enzimas CAZy. Nossos resultados mostram que a fosforilação desempenha papel importante na regulação dessas enzimas e de outras funções celulares no fungo após a transcrição de seus respectivos genes. O agrupamento desses dados permite melhor entendimento da via de sinalização mediada pelas MAPK TMK1 e TMK2 de T. reesei, e como as modificações pós-traducionais promovidas por elas afetam no sensing de nutrientes celulares e, por consequência, na produção de enzimas celulolíticas, de forma direta ou indireta.
Title in English
Identification of phosphorylation targets for Trichoderma reesei MAPKs through phosphoproteomics during the production of cellulases
Keywords in English
Cellulases
MAPK
Phosphoproteomics
Signaling pathways
Trichoderma reesei
Abstract in English
The filamentous fungus Trichoderma reesei has great biotechnological importance in regards to the lignocellulosic biomass degradation for large-scale production of bioethanol. Its cellulolytic system is very efficient, despite being composed by only a few cellulases, which suggests that the control of these enzymes goes beyond their transcriptional regulation. Thus, in this study, we performed a spectral counting LC-MS/MS analysis and achieved the phosphoproteomic profile of T. reesei grown either in glucose or sugarcane bagasse as sole carbon source. The comparison between the phosphoproteins profiles obtained from the parental strain QM6a and the knockout mutants for TMK1 and TMK2 allowed us to demonstrate that these MAPK act in an interconnected manner with other signaling transduction pathways, especially the TOR and cAMP-PKA pathways, in order to regulate the cellulases production. Furthermore, we were also able to determine the regulation of cellular stress response by TMK2, and the role of phosphorylation in the direct control of CAZymes. Our results show that phosphorylation plays an important role on the control of these enzymes and other cellular functions in T. reesei after the transcription of their respective genes. Taken together, this data allows better comprehension of the signaling pathways mediated by TMK1 and TMK2 in T. reesei, and how the post-translational modification promoted by these MAPK might affect the nutrient sensing and, therefore, the production of the cellulolytic enzymes, either directly or indirectly.
 
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Publishing Date
2019-02-11
 
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