O projeto genoma de Penicillium echinulatum 2HH e S1M29: a genômica viabilizando a descoberta de conhecimento

Abstract

Cellulolytic enzymes produced by fungi are used in several industrial processes, such as the production of fabrics, paper, food and biofuels. Penicillium echinulatum 2HH is an ascomycete isolated from the digestive tract of coleoptera larvae in 1979, also known for its enzymatic cocktails. To improve the saccharification yield of cellulosic biomass for commercial exploitation, one strategy is the design of hypersecretory strains of enzymes. However, the molecular knowledge about the lignocellulolytic system of this fungus is quite limited. In 2013, both genomes of S1M29 mutant and 2HH wild-type of P. echinulatum were sequenced. The purpose of this thesis included the assembly, annotation and deposit of both draft genomes in public databases, enabling knowledge discovery from genomic data. The knowledge discovery covered a series of domains: i) molecular identification to reposition the species; ii) characterization of accumulated mutations in S1M29 mutant; iii) evolutionary analyzes based on general and specific markers; iv) characterization of target genes: carbohydrate-active enzymes (CAZymes), sugar transporters (STs) and transcription factors (TFs); v) construction of the gene regulatory network (GRN); and vi) identification of target genes to obtain commercial strains. The molecular identification of 2HH wild-type strain and its repositioning in the Oxalica series stand out for the primordiality for comparative studies with other microorganisms. The deposited draft genomes of 2HH wild-type and S1M29 mutant at GenBank makes it possible to expand the molecular understanding of this fungus. Analysis of accumulated mutations in S1M29 mutant highlighted a wide range of mutations, highlighting BGL2 enzyme and FlbA transcription factor that probably contains the main mutations involved in hyperproduction of cellulases. We also identified that the albino phenotype of S1M29 mutant resulted from a mutation in ALB1 enzyme, which belongs to the DHN-melanin biosynthesis pathway. Our results related to evolutionary analyzes hypothesize a potential long-term mutualistic symbiosis between P. echinulatum 2HH and Anobium punctatum, whose environment-specific interactions could explain the difference in gene composition in relation to Penicillium oxalicum 114-2. Furthermore, the CAZyome characterization of P. echinulatum 2HH demonstrates that the genes of the cellulolytic system are predominantly orthologous to P. oxalicum 114-2, including a monooxygenase of AA16 family, described for the first time in both fungi. Furthermore, the sugar transportome characterization has demonstrated the diversity and specificity of STs from P. echinulatum 2HH, including eight families with specificity for different groups of sugars. Finally, the TFome characterization and the GRNs of P. echinulatum 2HH and P. oxalicum 114-2 comprise regulatory interactions that cover several biological processes, exploring diverse regulatory modules, such as CpcA, FF-7, COL-26, AmyR, ClrB, CreA and XlnR. This thesis contributes strongly in building a framework for molecular understanding of P. echinulatum 2HH, revealing characteristics related to the cellulolytic enzymes production, sugar uptake, melanin production, cell wall composition and regulation of gene expression of this fungus. Knowledge of regulatory pathways, coupled with the characterization of CAZymes and STs provide tools to design commercial strains, enabling the use of P. echinulatum 2HH for large-scale 2G ethanol production. Finally, we highlight P. echinulatum 2HH as an important biotechnological ally for lignocellulosic biofuels production, helping in the global energy transition [resumo fornecido pelo autor].