Unveiling the mechanisms of gene regulation at the single cell level in Trichoderma reesei

• Grant number: 16/03763-3
• Support Opportunities: Scholarships in Brazil - Master
• Start date: August 01, 2016
• End date: January 31, 2019
• Field of knowledge: Biological Sciences - Microbiology - Biology and Physiology of Microorganisms
• Principal Investigator: Rafael Silva Rocha
• Grantee: Luísa Czamanski Nora
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated research grant: 12/22921-8 - Synthetic biology approaches for deciphering the logic of signal integration in complex bacterial promoters, AP.JP
• Associated scholarship(s): 18/02227-6 - Construction of engineered yeast strains for achieving better yields of biomass degradation, BE.EP.MS

Abstract

Cellulolytic enzymes produced by filamentous fungi have been the focus of several recent studies due to their ability to degrade agricultural waste and subsequent potential for biotechnological use in biorefinaries. Trichoderma reesei is the leading model of study for the production of these enzymes and it has shown promising results in the search for optimized organisms that generate higher yields. Numerous techniques have already been used to show that this fungus has a complicated molecular regulation network that controls the expression of those proteins depending on the substrate; however, an analysis in a single cell level has never been made to unveil what lies behind that mechanism. Among the cellulase complex, the enzymes Cel6A and Cel7A stand out for their role in degrading cellulose. Therefore, this project intends to clone promoters of both genes in recombinant vectors containing the fluorescent reporter proteins mCherry and GFP, to insert them into wild-type QM9414 and mutant xyr1 strains of T. reesei grown in three different carbon sources, and to analyze their expression cell by cell using fluorescence microscopy. Furthermore, a potential new cis-regulatory element identified in the promoter of cel6a gene will be studied through the construction of mutant promoters. Thus, it will be possible not only to prove the functionality of cellulase promoters of T. reesei, but also to analyze how is the molecular response of this organism when degrading complex carbon sources and to compare the expression of two genes at the same time in a single cell level. (AU)

Scientific publications

(6)

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

NORA, LUISA CZAMANSKI; WESTMANN, CAUA ANTUNES; GUAZZARONI, MARIA-EUGENIA; SIDDAIAH, CHANDRANAYAKA; GUPTA, VIJAI KUMAR; SILVA-ROCHA, RAFAEL. Recent advances in plasmid-based tools for establishing novel microbial chassis. BIOTECHNOLOGY ADVANCES, v. 37, n. 8, DEC 2019. (16/03763-3, 15/04309-1, 16/05472-6, 12/22921-8)

NORA, LUISA CZAMANSKI; WESTMANN, CAUA ANTUNES; MARTINS-SANTANA, LEONARDO; ALVES, LUANA DE FATIMA; OLIVEIRA MONTEIRO, LUMMY MARIA; GUAZZARONI, MARIA-EUGENIA; SILVA-ROCHA, RAFAEL. The art of vector engineering: towards the construction of next-generation genetic tools. MICROBIAL BIOTECHNOLOGY, v. 12, n. 1, p. 23-pg., 2019-01-01. (16/05472-6, 16/06323-4, 17/17924-1, 16/19179-9, 16/03763-3, 12/22921-8, 15/04309-1)

NORA, LUISA CZAMANSKI; WESTMANN, CAUA ANTUNES; MARTINS-SANTANA, LEONARDO; ALVES, LUANA DE FATIMA; OLIVEIRA MONTEIRO, LUMMY MARIA; GUAZZARONI, MARIA-EUGENIA; SILVA-ROCHA, RAFAEL. The art of vector engineering: towards the construction of next-generation genetic tools. MICROBIAL BIOTECHNOLOGY, v. 12, n. 1, p. 125-147, JAN 2019. (16/19179-9, 17/17924-1, 16/06323-4, 16/05472-6, 16/03763-3, 15/04309-1, 12/22921-8)

CAMPOS ANTONIETO, AMANDA CRISTINA; VIEIRA NOGUEIRA, KAROLINE MARIA; DE PAULA, RENATO GRACIANO; NORA, LUFSA CZAMANSKI; ANZOLINI CASSIANO, MURILO HENRIQUE; GUAZZARONI, MARIA-EUGENIA; ALMEIDA, FAUSTO; DA SILVA, THIAGO APARECIDO; RIES, LAURE NICOLAS ANNICK; DE ASSIS, LEANDRO JOSE; et al. A Novel Cys2His2 Zinc Finger Homolog of AZF1 Modulates Holocellulase Expression in Trichoderma reesei. MSYSTEMS, v. 4, n. 4, JUL-AUG 2019. (16/23233-9, 16/03322-7, 15/04309-1, 17/04217-5, 16/03763-3, 18/03766-8, 15/09553-8, 12/22921-8)

LUÍSA CZAMANSKI NORA; RELBER AGUIAR GONÇALES; LEONARDO MARTINS-SANTANA; BEATRIZ HENRIQUES FERREIRA; FERNANDO RODRIGUES; RAFAEL SILVA-ROCHA. Synthetic and minimalist vectors for Agrobacterium tumefaciens-mediated transformation of fungi. GENETICS AND MOLECULAR BIOLOGY, v. 42, n. 2, p. 395-398, 2019-06-13. (16/03763-3, 16/01946-3, 14/22561-7, 12/22921-8)

MARTINS-SANTANA, LEONARDO; NORA, LUISA C.; SANCHES-MEDEIROS, ANANDA; LOVATE, GABRIEL L.; CASSIANO, MURILO H. A.; SILVA-ROCHA, RAFAEL. Systems and Synthetic Biology Approaches to Engineer Fungi for Fine Chemical Production. FRONTIERS IN BIOENGINEERING AND BIOTECHNOLOGY, v. 6, OCT 3 2018. (15/22386-3, 17/17924-1, 16/11093-8, 17/04217-5, 16/03763-3, 12/22921-8)

Academic Publications

(References retrieved automatically from State of São Paulo Research Institutions)

NORA, Luísa Czamanski. Development of synthetic biology tools applied to fungi of medical and industrial importance. 2019. Master's Dissertation - Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC) Ribeirão Preto.