Analysis of the transcription factor regulation TCF21 and its role in cell migration and invasion in tumors.

Author(s)

Jean Lucas Kremer

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI)
• Defense date: 2020-12-14

Examining board members

Claudimara Ferini Pacicco Lotfi; Mariana Brait Rodrigues de Oliveira; Carlos Alberto Scrideli; Bryan Eric Strauss

• Advisor: Claudimara Ferini Pacicco Lotfi

Abstract

Adenomas of the adrenal cortex (ACA) are common tumors while carcinomas (ACC) are rare with limited prognosis and frequent recurrences and/or metastases. In Brazil, there is interest in the study of these tumors, because its incidence is higher than the rest of the world, mainly in children. The transcription factor TCF21 is epigenetically regulated through the transcription of a non-coding RNA (TARID), responsible for the demethylation of the promoter of TCF21 gene and activation of the its transcription. TCF21 is involved in the development of different organs, including the adrenal gland, and is important for development, as TCF21 knockout mice do not survive. TCF21 is less expressed in several tumors, and epigenetically silenced due to methylation of its promoter.TCF21 inactivation is related to an increase capacity for cellular invasion and metastasis, through a mechanism that involves the expression of genes associated with the control of cellular motility. Microarray results showed that TCF21 is less expressed in ACC than ACA. However, the methylation condition of TCF21, the effect of its expression or silencing on adrenocortical tumor cells, as well as its effect on cell migration and invasion, and the mechanisms involved, are unknown in these tumors. Our results showed that the TCF21 promoter is more methylated in ACC cell lines than in an ACA cell culture. This condition was correlated with the lower expression of TCF21 in ACC cells, when compared with cultures of hyperplasia cells and adenoma cells of the adrenal cortex. The use of the demethylating agent 5-aza-20-deoxycytidine in ACC cell lines suggests that TCF21 is epigenetic silencing by methylation of its promoter. The increase in the endogenous expression of TCF21 through the CRISPR/dCas9/TCF21 system or its overexpression with the plasmid pCMVMycPOD1, reduced the migration and cell invasion in the ACC cell lines. TCF21 positively modulated the expression of genes involved with anti-invasive function such as the tumor metastasis suppressor gene (KISS1), metalloproteinase 8 (MMP8) and metalloproteinase 1 inhibitor (TIMP1). On the other hand, genes with invasive function, genes MMP2, MMP9 and MMP14, were inhibited in the presence of TCF21. In adenoma cells, siRNA-TCF21 use for the silencing of TCF21 induced the invasive capacity of these cells through the opposite mechanism. In hepatocarcinoma cells, data from our group showed opposite effects of TCF21 compared ACC cells. In hepatocarcinoma cells, data from our group showed opposite effects of TCF21 compared to ACC cells. In fact, in hepatocarcinoma cells, TCF21 increased migration and invasion through the expression of invasive genes and inhibition of the antiinvasive genes studied. Therefore, in ACC cells, TCF21 is regulated by promoter methylation, and the effect is observed on cell motility. The mechanism by which TCF21 modifies cell motility suggests the involvement of metalloproteinases and their regulators. On the other hand, the results obtained in hepatocarcinoma cells, showed that the definition of TCF21 as a tumor suppressor is conditioned to cellular context analyzed. (AU)

• FAPESP's process: 16/17285-6 - Study of epigenetic regulation of transcription factor POD-1/TCF21in adrenocortical tumor cell cultures and its role in the cellular migration and invasion
• Grantee: Jean Lucas Kremer
• Support Opportunities: Scholarships in Brazil - Doctorate