Non-coding antisense transcription detected by conventional and single-stranded cDNA microarray

Johan Vallon-Christersson,Åke Borg,Patrik Medstrand,Carlos Rovira,Anders Kvist,Johan Staaf

doi:10.1186/1471-2164-8-295

Abstract

BackgroundRecent studies revealed that many mammalian protein-coding genes also transcribe their complementary strands. This phenomenon raises questions regarding the validity of data obtained from double-stranded cDNA microarrays since hybridization to both strands may occur. Here, we wanted to analyze experimentally the incidence of antisense transcription in human cells and to estimate their influence on protein coding expression patterns obtained by double-stranded microarrays. Therefore, we profiled transcription of sense and antisense independently by using strand-specific cDNA microarrays.ResultsUp to 88% of expressed protein coding loci displayed concurrent expression from the complementary strand. Antisense transcription is cell specific and showed a strong tendency to be positively correlated to the expression of the sense counterparts. Even if their expression is wide-spread, detected antisense signals seem to have a limited distorting effect on sense profiles obtained with double-stranded probes.ConclusionAntisense transcription in humans can be far more common than previously estimated. However, it has limited influence on expression profiles obtained with conventional cDNA probes. This can be explained by a biological phenomena and a bias of the technique: a) a co-ordinate sense and antisense expression variation and b) a bias for sense-hybridization to occur with more efficiency, presumably due to variable exonic overlap between antisense transcripts.

Highlights

Recent studies revealed that many mammalian protein-coding genes transcribe their complementary strands
We found that the most reliable method for processing double stranded cDNAs into single stranded capture probes was the sequential application of both approaches
In spite of stringent signal-to-noise ratio criteria, our strand-specific cDNA microarrays detected a number of antisense transcripts that exceeds by far the number of previously annotated antisense genes in humans

Summary

Introduction

Recent studies revealed that many mammalian protein-coding genes transcribe their complementary strands This phenomenon raises questions regarding the validity of data obtained from double-stranded cDNA microarrays since hybridization to both strands may occur. NATs (cis and trans) may be higher yet, since nuclear nonpolyadenylated transcripts are underrepresented in transcript sequence databases This fact may have important implications for researchers, because of their potential biological function but they may turn out to be influential on the interpretation of large experimental data sets. The cDNA microarray technique has been used in genomewide expression studies to address basic questions about gene function and in the pursuit of a more precise molecular classification of tumors In this case, the ability to monitor the expression of thousands of genes simultaneously has allowed the identification of disease-specific subsets of genes useful to improve diagnosis and disease management [11]. We reasoned that antisense transcripts are either not efficiently detected by conventional cDNA capture probes or that important information must be hidden behind this paradox

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