Large-scale analysis of microRNA function in cell cycle, pluripotency, self-renewal and differentiation of stem cells using high-content screening

• Grant number: 15/08070-3
• Support Opportunities: Scholarships abroad - Research
• Effective date (Start): December 01, 2015
• Effective date (End): June 30, 2016
• Field of knowledge: Health Sciences - Medicine - Medical Clinics
• Principal Investigator: Rodrigo Alexandre Panepucci
• Grantee: Rodrigo Alexandre Panepucci
• Host Investigator: Marc Bernard Thomas Bickle
• Host Institution: Hemocentro de Ribeirão Preto. Hospital das Clínicas da Faculdade de Medicina de Ribeirão Preto da USP (HCMRP). Secretaria da Saúde (São Paulo - Estado). Ribeirão Preto , SP, Brazil
• Research place: Max Planck Society, Dresden, Germany
• Associated research grant: 13/08135-2 - CTC - Center for Cell-Based Therapy, AP.CEPID

Abstract

Pluripotency and self-renewal are distinctive properties of embryonic stem cells (ESCs). microRNAs (miRs) play important roles in the post-transcriptional control of gene expression, by targeting multiple mRNA, and have been implicated in the maintenance of pluripotent cells characteristics. Many miRs effects are attributed to cell cycle regulation, in line with the intimate link between cell-cycle regulation, self-renewal, pluripotency and differentiation. We will systematically investigate the roles of all currently annotated human miRs (2,588 mature miRs) in these characteristics, using automated High-Content Screening fluorescence microscopy. Images derived from pluripotent cells transfected with microRNA mimics will be analyzed, in order to quantify biological features reflecting, cell cycle status (based on Cyclin B1 staining and Hoechst/DNA content analysis), pluripotency (based on Oct4 staining) and differentiation (cell morphology). Single-cell data will be combined into multiparametric population profiles describing miR induced phenotypes. Profiles will be clustered to identify microRNA groups with common effects. Enrichment functional analysis of predicted miR targets will be carried out to identify biological pathways and processes mediating the phenotypes observed. The primary objective of the present FAPESP-BPE application is to allow the applicant to develop specific image and data analysis pipelines, under the guidance of Dr. Marc Bickle (Max Planck Institute of Molecular Cell Biology and Genetics, Dresden-Germany). For this, images from a pilot focused screen (30 miRs) will be used in the development and optimization of the analysis pipelines. Once established, the pipelines will be adapted to the full library screen, which will be carried by the PhD student Ildercílio Lima (supervised by Dr. Panepucci, FAPESP 2013/27061-0), in collaboration with Dr. Miguel Mano (ICGEB, Trieste-Italy). In view of the complexity of the analytical approaches, this fellowship will bring important immediate and precisely defined benefits for the success of this project (a subproject of the Center for Cell Therapy - CTC, a CEPID FAPESP).

Scientific publications

(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)

PANEPUCCI, RODRIGO ALEXANDRE; DE SOUZA LIMA, ILDERCILIO MOTA. Arrayed functional genetic screenings in pluripotency reprogramming and differentiation. STEM CELL RESEARCH & THERAPY, v. 10, JAN 11 2019. (15/15864-6, 13/08135-2, 15/08070-3, 13/27061-0)

SANGIORGI, BRUNO; DE SOUZA, FELIPE CANTO; DE SOUZA LIMA, ILDERCILIO MOTA; DOS SANTOS SCHIAVINATO, JOSIANE LILIAN; CORVELONI, AMANDA CRISTINA; THOME, CAROLINA HASSIBE; SILVA JR, WILSON ARAUJO; FACA, VITOR MARCEL; COVAS, DIMAS TADEU; ZAGO, MARCO ANTONIO; et al. A High-Content Screening Approach to Identify MicroRNAs Against Head and Neck Cancer Cell Survival and EMT in an Inflammatory Microenvironment. FRONTIERS IN ONCOLOGY, v. 9, NOV 8 2019. (15/08070-3, 13/08135-2)

DE SOUZA LIMA, ILDERCILIO MOTA; DOS SANTOS SCHIAVINATO, JOSIANE LILIAN; PAULINO LEITE, SARAH BLIMA; SASTRE, DANUTA; DE OLIVEIRA BEZERRA, HUDSON LENORMANDO; SANGIORGI, BRUNO; CORVELONI, AMANDA CRISTINA; THOME, CAROLINA HASSIBE; FACA, VITOR MARCEL; COVAS, DIMAS TADEU; et al. High-content screen in human pluripotent cells identifies miRNA-regulated pathways controlling pluripotency and differentiation. STEM CELL RESEARCH & THERAPY, v. 10, JUL 8 2019. (15/08070-3, 15/15864-6, 13/27061-0, 13/08135-2)