Scholarship 19/18565-0 - Toxicologia, Biologia computacional - BV FAPESP
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Evaluation by quantitative in situ hybridization of the transcript levels of Natterin-like in CRISPR/Cas-9 depleted zebrafish

Grant number: 19/18565-0
Support Opportunities:Scholarships in Brazil - Scientific Initiation
Start date until: November 01, 2019
End date until: September 30, 2020
Field of knowledge:Biological Sciences - Immunology - Applied Immunology
Principal Investigator:Carla Lima da Silva
Grantee:Milena Marcolino de Souza
Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
Associated research grant:13/07467-1 - CeTICS - Center of Toxins, Immune-Response and Cell Signaling, AP.CEPID

Abstract

The Nattering family of proteins found in Thalassophryne nattereri fish venom has five orthologs (1 - 4 and -P) with N-terminal (DM9) and C-terminal (aerolysin) domains. Nattering-like protein sequences can be found in different immune defense cells of various venomous or non-venomous fish species. Moreover, recent studies point to the participation of Nattering-like proteins in immune defense against pathogens. Aerolysin-like proteins, known as pore-forming proteins (PFTs), are present in approximately 30% of all proteins/toxins in pathogenic bacteria and are capable of activating the NRLP3 inflammasome. Bioinformatic analysis of zebrafish specimens reveals some aerolysin isoforms that share 60% identity with the catfish and lamprey nattering-like protein sequences, which are considered pathogen defense molecules. Zebrafish has proven to be an excellent model organism for genetic manipulation because of its transparency, temporal separation of immune responses, easy and low costs of maintenance and manipulation. Its system enables high-resolution quantitative imaging and zebrafish possesses 70% homology to human genes. Our group has been testing the hypothesis that nattering acts as defensive molecules in host fish using the most efficient loss-of-function CRISPR - Cas9 system for nattering-like gene silencing. The objective of this research project is to validate the efficiency of CRISPR-Cas9 depletion of the nattering-like gene by mRNA evaluation in silenced zebrafish larvae using the in situ hybridization technique. (AU

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