Related Experiment Video
Updated: May 24, 2025

HOX Loci Focused CRISPR/sgRNA Library Screening Identifying Critical CTCF Boundaries
Published on: March 31, 2019
Natural Antisense Transcript-Mediated Regulation of HOXA10-AS in Oral Squamous Cell Carcinoma
Kanaka Sai Ram Padam1, Satyajit Dey Pereira1, Naveena A N Kumar2
1Department of Cell and Molecular Biology, Manipal School of Life Sciences, Manipal Academy of Higher Education, Manipal, Karnataka, India.
Background:
The oncogenic role of HOXA10-AS and HOXA10 in cancer has been well documented. However, the epigenetic role of HOXA10 and the natural antisense-mediated regulation of HOXA10-AS in oral squamous cell carcinoma progression is not understood.
Methods:
A total of 35 oral squamous cell carcinoma specimens and 35 adjacent normal clinical specimens were collected and categorized on the basis of their lymph node status. HOXA10-AS and HOXA10 expression were analyzed using RT-qPCR. Methyl-capture sequencing was performed using lymph node-negative (n = 6) and lymph node-positive (n = 5) matched cases. The promoter activity of HOXA10 was determined using a luciferase assay. ChIP-qPCR was performed to determine histone mark localization in the distal promoter region of HOXA10. A protein-protein interaction network of genome-wide antisense targets was constructed using StringDB, and functional enrichment was performed using the R package ClusterProfiler. Transient siRNA-mediated transfection was performed to target specific exons of the HOXA10-AS gene, followed by subsequent cell proliferation, cell cycle, and cell migration assays and validation of cancer signaling pathways through western blotting.
Results:
HOXA10-AS and its antisense target HOXA10 were significantly overexpressed in the lymph node-positive samples. The transcriptionally active distal promoter of HOXA10 consists of a constitutively unmethylated CpG island region (CUR). H3K4me3, H3K27ac, and H3K27me3 histone mark deposition at the adjacent methylated loci of the distal promoter suggest the nature of euchromatin-driven regulation. Genome-wide mapping revealed 11 potential targets of HOXA10-AS. Targeted specific knockdown of HOXA10-AS exons significantly reduced the expression of HOXA10 and deregulated its downstream targets, contributing to decreased cell cycle progression and epithelial-to-mesenchymal transition.
Conclusion:
HOXA10-AS regulates the expression of HOXA10 through a natural antisense-mediated mechanism and is epigenetically regulated by constitutively unmethylated marks in the distally enhancing promoter of HOXA10.
More Related Videos
Related Concept Videos
lncRNA - Long Non-coding RNAs
Pleiotropy
MicroRNAs
siRNA - Small Interfering RNAs
In the cytoplasm, siRNA is processed from a double-stranded RNA, which comes from either endogenous DNA transcription or exogenous sources like a virus. This double-stranded RNA is then cleaved by the...
Experimental RNAi
Loss of Tumor Suppressor Gene Functions
When the tumor suppressor genes develop mutations or are lost, cells start growing out of control, leading to cancer. However, a single functional copy of the tumor suppressor gene is enough for the cells to maintain their normal functions and cell...

