Dados do Resumo

Título

Role of SIVA1 in Breast Cancer Development: Insights from In Vitro Models

Introdução

Despite directed efforts to enhance breast cancer treatments, the disease continues to progress. One of the characteristic features of cancers, in general, is the alteration in the processes of cell growth, division, and death, processes that depend on the coordinated and integrated activity of multiple genes and proteins. There is an anticipation of a more profound exploration of those genes with oncogenic potential, including SIVA1, known for its involvement in apoptosis, migration, cytoskeletal dynamics, and epithelial-mesenchymal transition.

Objetivo

This study aims to describe correlations between SIVA1 expression and patient prognosis, along with pathways associated with its expression through bioinformatics analyses and its characterization in in vitro breast cancer models.

Métodos

Prognostic data were obtained from the cBioPortal platform, gene expression studies utilized IBM SPSS 20, and results were processed with GraphPad Prism 8. The Timer 2.0 platform was employed for analyzing SIVA1 differential expression among cancers. Molecular interactions were investigated using the GSEA program, and gene expression correlations were verified using Galaxy Analysis and ShinyGO 0.80. Spearman’s correlation was conducted with Prism 8 software. Siva content in total extracts from the cell lines MCF7, MDA-MB-231, Hs578T, T47D, HCC1937, HCC70, BT549, MCF10A, and MCF12A was evaluated by Western Blot, using the SuperSignal™ West Dura Extended Duration Substrate and G-box system. Simultaneously, RNA extraction was performed, with the relative gene expression for each cell line quantified by 32 QuantStudio 3 Real-Time PCR System and Power SybrGreen PCR Master Mix reagent. MDA-MB-231 and Hs578T cell lines were prepared for immunofluorescence, involving fixation with 4% paraformaldehyde, blocking and permeabilization with 3% BSA and 0.5% Triton solution, and labeling with Alexa Fluor® 467 and anti-Siva antibody (1:50, 1:200, and 1:500).

Resultados

SIVA1 expression was significantly higher in breast cancer patients compared to healthy donors (p<0.001), showing no correlations with overall, progression-free, and specific survival or disease-free status. Cellular mechanisms related to SIVA1 activity included protein secretion, spliceosome and ribosome activity, base excision repair, activation of ROS pathways, and the main rRNA processing pathway. Protein flow alterations were induced through their involvement in lipoprotein metabolism, lipidation, and ubiquitin binding. Direct interaction with genes associated with apoptosis, the p53 pathway, and other cascades hyperactive in cancers were identified. Experimental research confirmed a higher protein content of Siva in triple-negative (basal-like) cell lines, as supported by q-PCR results. Two of these triple-negative cell lines (MDA-MB-231 and Hs578T) were selected for further study, and immunofluorescence labeling revealed a perinuclear localization of the protein, displaying a punctate pattern, strengthening its involvement in both nuclear and cytoplasmic processes, as detailed in the bioinformatics analyses.

Conclusões

Our findings suggest that decreased SIVA1 expression may be linked to events commonly observed in tumor cells, such as attenuated repair mechanisms and decreased apoptotic rate.

Financiador do resumo

N.S.P. and B.O.A. received fellowships from FAPESP (grants #2023/11752-5 and #2022/11038-8). This study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, Brazil (CAPES), Finance Code 001.

Palavras Chave

SIVA1 gene; Breast cancer; Cell signaling pathways