The goal of the current study was to use the CRISPR/Cas9 method to efficiently knock down Elk1 and examine how it affected the interacting genes in triple-negative breast cancer cells. Elk1 expression analysis, enrichment analysis, and survival analysis were performed on TNBC cells to provide clarification. MDA-MB-231 breast cancer cells were transfected with CRISPR/Cas9 that targeted the Elk1 gene using the Lipofectamine CRISPR MAX reagent. Sanger sequencing and sequence trace decomposition were used to verify DNA target editing. Target gene expression levels were determined using qRT-PCR. Protein expression was examined using immunofluorescence. According to this work, Elk1-edited triple-negative breast cancer cells had decreased Elk1 protein levels and substantial DNA damage at the gene level. Elk1 has important interactors in a number of pathways, according to Reactome and IntACT studies. Mapk3, Sumo1, Nfe2l2, Eag1, Ube2, Tert, Plcg2, and CRK genes were the factors that were up-regulated (P < 0.05). Furthermore, Elk1 interactors’ genes were shown to be engaged in biological processes and cellular components related to signal transduction and cell communication, according to David enrichment analyses. Gene ontology terms such as enzyme binding, phosphotyrosine residue binding, phosphoprotein binding, protein tyrosine kinase binding, protein phosphorylated amino acid binding, DNA-binding transcription factor binding, protein binding, and transcription factor binding, are also cooperatively participated in by the gene list, according to DAVID analyses (P < 0.05). Furthermore, the current study indicated a significant reduction in metastatic potential and invasive features, as well as enhanced caspase 3/7 activity following Elk1 knockdown (P < 0.05). Overall, the current work successfully knocked down Elk1 by concentrating on coding exons of the Elk1 gene. When Elk1 was knocked down, multiple Elk1 interactors genes were highly expressed by triple-negative breast cancer cells. Meanwhile, apoptotic activity was increased, whereas metastatic potential and invasive characteristics were reduced. All things considered, the current study raised uncertainties about the specific carcinogenic role of the transcription factor Elk-1. Consequently, additional research is necessary to completely understand the methods via which Elk1 influences triple-negative breast cancer cells.

CRISPR/Cas9 Mediated Elk1 Gene Editing Enhances Elk1 Interactors’ Genes in Triple Negative Breast Cancer Cells

期刊论文
发表日期：

2026年1月9日

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Shaymaa M. M. Yahya,
Mahmoud T. Abo-Elfadl,
Shereen M. M. Yahya,
Neama I. Ali,
Sohair M. Salem

摘要

The goal of the current study was to use the CRISPR/Cas9 method to efficiently knock down Elk1 and examine how it affected the interacting genes in triple-negative breast cancer cells. Elk1 expression analysis, enrichment analysis, and survival analysis were performed on TNBC cells to provide clarification. MDA-MB-231 breast cancer cells were transfected with CRISPR/Cas9 that targeted the Elk1 gene using the Lipofectamine CRISPR MAX reagent. Sanger sequencing and sequence trace decomposition were used to verify DNA target editing. Target gene expression levels were determined using qRT-PCR. Protein expression was examined using immunofluorescence. According to this work, Elk1-edited triple-negative breast cancer cells had decreased Elk1 protein levels and substantial DNA damage at the gene level. Elk1 has important interactors in a number of pathways, according to Reactome and IntACT studies. Mapk3, Sumo1, Nfe2l2, Eag1, Ube2, Tert, Plcg2, and CRK genes were the factors that were up-regulated (P < 0.05). Furthermore, Elk1 interactors’ genes were shown to be engaged in biological processes and cellular components related to signal transduction and cell communication, according to David enrichment analyses. Gene ontology terms such as enzyme binding, phosphotyrosine residue binding, phosphoprotein binding, protein tyrosine kinase binding, protein phosphorylated amino acid binding, DNA-binding transcription factor binding, protein binding, and transcription factor binding, are also cooperatively participated in by the gene list, according to DAVID analyses (P < 0.05). Furthermore, the current study indicated a significant reduction in metastatic potential and invasive features, as well as enhanced caspase 3/7 activity following Elk1 knockdown (P < 0.05). Overall, the current work successfully knocked down Elk1 by concentrating on coding exons of the Elk1 gene. When Elk1 was knocked down, multiple Elk1 interactors genes were highly expressed by triple-negative breast cancer cells. Meanwhile, apoptotic activity was increased, whereas metastatic potential and invasive characteristics were reduced. All things considered, the current study raised uncertainties about the specific carcinogenic role of the transcription factor Elk-1. Consequently, additional research is necessary to completely understand the methods via which Elk1 influences triple-negative breast cancer cells.