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Published on: March 12, 2021
RNA-seq and ChIP-seq Identification of Unique and Overlapping Targets of GLI Transcription Factors in Melanoma Cell
Matea Kurtović1, Nikolina Piteša1, Nenad Bartoniček2,3
1Division of Molecular Medicine, Ruđer Bošković Institute, 10 000 Zagreb, Croatia.
Background:
Despite significant progress in therapy, melanoma still has a rising incidence worldwide, and novel treatment strategies are needed. Recently, researchers have recognized the involvement of the Hedgehog-GLI (HH-GLI) signaling pathway in melanoma and its consistent crosstalk with the MAPK pathway. In order to further investigate the link between the two pathways and to find new target genes that could be considered for combination therapy, we set out to find transcriptional targets of all three GLI proteins in melanoma.
Methods:
We performed RNA sequencing on three melanoma cell lines (CHL-1, A375, and MEL224) with overexpressed GLI1, GLI2, and GLI3 and combined them with the results of ChIP-sequencing on endogenous GLI1, GLI2, and GLI3 proteins. After combining these results, 21 targets were selected for validation by qPCR.
Results:
RNA-seq revealed a total of 808 differentially expressed genes (DEGs) for GLI1, 941 DEGs for GLI2, and 58 DEGs for GLI3. ChIP-seq identified 527 genes that contained GLI1 binding sites in their promoters, 1103 for GLI2 and 553 for GLI3. A total of 15 of these targets were validated in the tested cell lines, 6 of which were detected by both RNA-seq and ChIP-seq.
Conclusions:
Our study provides insight into the unique and overlapping transcriptional output of the GLI proteins in melanoma. We suggest that our findings could provide new potential targets to consider while designing melanoma-targeted therapy.
Insights
Researchers identified new target genes in melanoma by studying the Hedgehog-GLI (HH-GLI) pathway. These findings offer potential targets for combination therapies to combat rising melanoma incidence.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Melanoma incidence is increasing globally, necessitating novel therapeutic strategies.
- The Hedgehog-GLI (HH-GLI) signaling pathway is implicated in melanoma and interacts with the MAPK pathway.
Purpose of the Study:
- To investigate the crosstalk between HH-GLI and MAPK pathways in melanoma.
- To identify novel transcriptional targets of GLI proteins for potential combination therapy.
Main Methods:
- RNA sequencing was performed on melanoma cell lines with overexpressed GLI1, GLI2, and GLI3.
- ChIP-sequencing was used to identify endogenous GLI protein binding sites.
- Quantitative PCR (qPCR) was used to validate selected target genes.
Main Results:
- RNA-seq identified hundreds of differentially expressed genes for GLI1 and GLI2, and fewer for GLI3.
- ChIP-seq revealed numerous genes with GLI binding sites in their promoters.
- Fifteen potential target genes were validated, with six identified by both RNA-seq and ChIP-seq.
Conclusions:
- The study elucidates the distinct and shared transcriptional targets of GLI proteins in melanoma.
- Identified targets represent potential candidates for developing new melanoma-targeted therapies.
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