Identification of novel endogenous antisense transcripts by DNA microarray analysis targeting complementary strand of...

Area of Science:

• Genomics
• Molecular Biology
• Gene Expression Regulation

Background:

• Natural antisense transcripts (NATs) are widespread in mammals, transcribed from the opposite strand of protein-coding genes.
• Their regulatory mechanisms on gene expression are largely unknown, and functionally characterized NATs are limited.
• Previous identification methods missed many potential NATs, especially those lacking extensive cDNA evidence.

Purpose of the Study:

• To develop and apply a DNA microarray-based screening method to identify novel NATs in mouse tissues and mammary tumors.
• To investigate the expression patterns and conservation of NATs, including those not found in public cDNA databases.
• To identify NATs with altered expression in mammary cancer for further functional studies.

Main Methods:

• Designed DNA microarray probes targeting the complementary strand of annotated genes, including those with only human cDNA evidence.
• Screened adult mouse normal tissues and mammary tumors for NAT expression.
• Analyzed expression of selected NATs (e.g., Acaa1b, Aard) in vivo.
• Compared sense and antisense gene expression between cancerous and non-pathological mammary tissues.

Main Results:

• Identified prominent expression signals for 66.1% of 635 target genes not previously identified in mouse cDNA.
• Observed tissue-specific expression in 58 of these genes, indicating dynamic transcription.
• Found that antisense expression of 95 out of 404 well-annotated genes was significantly altered in mammary tumor tissue.
• Detected changes in sense gene expression for 19 of these 95 genes, highlighting coordinated regulation.

Conclusions:

• The developed microarray platform successfully identified novel NATs missed by conventional methods.
• Differentially expressed NATs in tumors represent promising candidates for investigating gene function and regulatory roles.
• This platform is versatile and applicable to any gene and sample type for NAT discovery.