Functional analysis of microRNAs on cellular dedifferentiation during tissue regeneration in the zebrafish model

• Grant number: 18/05484-0
• Support Opportunities: Regular Research Grants
• Field of knowledge: Biological Sciences - Genetics - Animal Genetics
• Principal Investigator: Danillo Pinhal
• Grantee: Danillo Pinhal
• Host Institution: Instituto de Biociências (IBB). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil
• Associated researchers: César Martins ; Flávia Karina Delella ; Luiz Augusto Bovolenta ; Ney Lemke
• Associated scholarship(s): 19/26289-3 - Functional analysis of microRNAs on cellular dedifferentiation during tissue regeneration in the zebrafish model, BP.TT; 18/14348-2 - Functional analysis of microRNAs on cellular dedifferentiation during tissue regeneration in the zebrafish model, BP.TT

Abstract

Regeneration of lost or damaged body parts is an intriguing biological phenomenon. During regeneration, cell populations adjacent to the lesion dedifferentiate, reorganize and then differentiate, resulting in a structure functionally and morphologically identical to the lost one. The study of teleost caudal fin regeneration, especially in zebrafish (Danio rerio), has been providing support for understanding the biological mechanisms controlling tissue regeneration in vertebrates. In this process, mature osteoblasts dedifferentiate in osteoprogenitor cells, repopulating the bony parts lost after fin injury or amputation. At the molecular level, this event induces the expression of gene signaling networks involving endogenous small non-coding RNAs, called microRNAs, and transcription factors. In this context, sp7, a mature osteoblast marker, exhibit high expression levels adjacent to the lesion, decreasing progressively as they move distally to the amputation site, suggesting that this transcription factor plays an important role in osteoblast dedifferentiation in zebrafish. Also, several microRNAs have been associated with regeneration in zebrafish, although their exact functional role in cell dedifferentiation has not been elucidated. Thus, it is relevant to identify the group of differentially expressed microRNAs and their targets, as well as microRNAs regulating sp7, in osteoblast dedifferentiation during regeneration. To accomplish this, we propose the association of diverse experimental techniques, including the isolation of dedifferentiating cells using cell sorting, the analysis of expression profiles of microRNAs and messenger RNAs (mRNA) using next-generation sequencing (RNA-seq), followed by validation of microRNA / target interactions, as well as superexpression and inhibition experiments in zebrafish embryos in vivo. Still, we propose the investigation of the evolutive conservation level of the observed regulatory interactions through superexpression and inhibition experiments in mice cells, in vitro. The activity of microRNAs in the modulation of their targets will be evaluated by the quantification of transcripts (quantitative PCR) and proteins (Western blotting). We hope to contribute to a better understanding of cell dedifferentiation process, as well as to enlarge the list of microRNA targets experimentally validated. The obtained knowledge about the regulation of this cellular mechanism, essential to the regeneration process, can be applied in future regenerative medicine technologies, making possible the manipulation of regulatory pathways to control alterations in cell differentiation states at lesion sites. (AU)

Scientific publications

(4)

(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)

RIBEIRO, AMANDA OLIVEIRA; DE OLIVEIRA, ARTHUR CASULLI; COSTA, JULIANA MARA; NACHTIGALL, PEDRO GABRIEL; HERKENHOFF, MARCOS EDGAR; CAMPOS, VINICIUS FARIAS; DELELLA, FLAVIA KARINA; PINHAL, DANILLO. MicroRNA roles in regeneration: Multiple lessons from zebrafish. DEVELOPMENTAL DYNAMICS, SEP 2021. (14/03062-0, 13/06864-7, 18/14348-2, 18/05484-0, 19/15494-5)

BLODORN, EDUARDO B.; DOMINGUES, WILLIAM B.; NUNES, LEANDRO S.; KOMNINOU, ELIZA R.; PINHAL, DANILLO; CAMPOS, VINICIUS F.. MicroRNA roles and their potential use as selection tool to cold tolerance of domesticated teleostean species: A systematic review. Aquaculture, v. 540, JUL 15 2021. (18/05484-0)

OLIVEIRA, UR C.; BOVOLENTA, LUIZ A.; ALVES, LUCAS; FIGUEIREDO, LUCAS; RIBEIRO, AMANDA O.; CAMPOS, VINICIUS E.; LEMKE, NEY; PINHAL, DANILLO. Understanding the Modus Operandi of MicroRNA Regulatory Clusters. CELLS, v. 8, n. 9, SEP 2019. (18/05484-0, 18/26409-6, 12/13450-1, 12/15589-7, 15/19176-7, 17/17510-2)

NUNES, LEANDRO SILVA; DOMINGUES, WILLIAM BORGES; KREMER, FREDERICO SCHMITT; PINHAL, DANILLO; CAMPOS, VINICIUS FARIAS. econstruction of regulatory network predicts transcription factors driving the dynamics of zebrafish heart regeneratio. Gene, v. 819, APR 20 2022. (18/05484-0)