Nascent vs. Steady-State Sequencing Reveals Extensive but Unstable Antisense Transcription in Yeast

Abstract: While only around 1% of our entire human genome consists of protein-coding regions, nearly all of it is shown to be active through transcription. A lot of this transcription occurs on the opposite strand of the annotated gene (the so-called "antisense" strand). The exact biological function of this form of transcription has yet to be fully understood. To characterize the extent, distribution, and stability of antisense transcription, both nascent (using NET-seq) and steady-state (using RNA-seq) transcription data for 2649 actively transcribed yeast genes were analyzed. Antisense transcription has been detected in 98.8% of all actively transcribed genes by means of NET-seq, compared to 77% by RNA-seq steady-state measurement; however, the level of this transcription is generally lower than that of the corresponding sense-transcribed RNA. Additionally, the ratio of sense-to-antisense reads measured by RNA-seq is much higher than the corresponding ratio measured by NET-seq; this suggests that, following transcription, antisense transcripts are degraded at a faster rate than sense transcripts. This was further confirmed by means of stability studies; median RNA-seq/NET-seq ratios show that antisense transcripts have a much lower median ratio than sense-transcribed RNA (-2.85 and -0.47 on a log2 scale, respectively), indicating a rapid rate of turnover for antisense RNA molecules. In summary, these findings provide evidence that not only is antisense transcription widely prevalent, but it is subject to strict regulation, primarily through the process of post-transcriptional degradation, which ultimately serves to limit accumulation and therefore expression of antisense transcripts.