Natural Antisense Transcript-Mediated Regulation of HOXA10-AS in Oral Squamous Cell Carcinoma

IF 2.7 3区医学 Q1 DENTISTRY, ORAL SURGERY & MEDICINE

Journal of Oral Pathology & Medicine Pub Date : 2025-03-04 DOI:10.1111/jop.13613

Kanaka Sai Ram Padam, Satyajit Dey Pereira, Naveena A. N. Kumar, Raghu Radhakrishnan

{"title":"Natural Antisense Transcript-Mediated Regulation of HOXA10-AS in Oral Squamous Cell Carcinoma","authors":"Kanaka Sai Ram Padam, Satyajit Dey Pereira, Naveena A. N. Kumar, Raghu Radhakrishnan","doi":"10.1111/jop.13613","DOIUrl":null,"url":null,"abstract":"<div>\n \n \n <section>\n \n <h3> Background</h3>\n \n <p>The oncogenic role of <i>HOXA10</i>-<i>AS</i> and <i>HOXA10</i> in cancer has been well documented. However, the epigenetic role of <i>HOXA10</i> and the natural antisense-mediated regulation of <i>HOXA10-AS</i> in oral squamous cell carcinoma progression is not understood.</p>\n </section>\n \n <section>\n \n <h3> Methods</h3>\n \n <p>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. <i>HOXA10-AS</i> and <i>HOXA10</i> expression were analyzed using RT-qPCR. Methyl-capture sequencing was performed using lymph node-negative (<i>n</i> = 6) and lymph node-positive (<i>n</i> = 5) matched cases. The promoter activity of <i>HOXA10</i> was determined using a luciferase assay. ChIP-qPCR was performed to determine histone mark localization in the distal promoter region of <i>HOXA10</i>. 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 <i>HOXA10-AS</i> gene, followed by subsequent cell proliferation, cell cycle, and cell migration assays and validation of cancer signaling pathways through western blotting.</p>\n </section>\n \n <section>\n \n <h3> Results</h3>\n \n <p>\n <i>HOXA10-AS</i> and its antisense target <i>HOXA10</i> were significantly overexpressed in the lymph node-positive samples. The transcriptionally active distal promoter of <i>HOXA10</i> 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 <i>HOXA10</i>-<i>AS</i>. Targeted specific knockdown of <i>HOXA10</i>-<i>AS</i> exons significantly reduced the expression of <i>HOXA10</i> and deregulated its downstream targets, contributing to decreased cell cycle progression and epithelial-to-mesenchymal transition.</p>\n </section>\n \n <section>\n \n <h3> Conclusion</h3>\n \n <p>\n <i>HOXA10</i>-<i>AS</i> regulates the expression of <i>HOXA10</i> through a natural antisense-mediated mechanism and is epigenetically regulated by constitutively unmethylated marks in the distally enhancing promoter of <i>HOXA10</i>.</p>\n </section>\n </div>","PeriodicalId":16588,"journal":{"name":"Journal of Oral Pathology & Medicine","volume":"54 4","pages":"217-231"},"PeriodicalIF":2.7000,"publicationDate":"2025-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jop.13613","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Oral Pathology & Medicine","FirstCategoryId":"3","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1111/jop.13613","RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"DENTISTRY, ORAL SURGERY & MEDICINE","Score":null,"Total":0}

引用次数: 0

Abstract

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.

Abstract Image

查看原文本刊更多论文

求助全文

约1分钟内获得全文求助全文

来源期刊

Journal of Oral Pathology & Medicine 医学-病理学

CiteScore

5.90

自引率

6.10%

发文量

121

审稿时长

4-8 weeks

期刊介绍： The aim of the Journal of Oral Pathology & Medicine is to publish manuscripts of high scientific quality representing original clinical, diagnostic or experimental work in oral pathology and oral medicine. Papers advancing the science or practice of these disciplines will be welcomed, especially those which bring new knowledge and observations from the application of techniques within the spheres of light and electron microscopy, tissue and organ culture, immunology, histochemistry and immunocytochemistry, microbiology, genetics and biochemistry.