Polyadenylation site-induced decay of upstream transcripts enforces promoter directionality

Active human promoters produce promoter-upstream transcripts (PROMPTs). Why these RNAs are coupled to decay, whereas their neighboring promoter-downstream mRNAs are not, is unknown. Here high-throughput sequencing demonstrates that PROMPTs generally initiate in the antisense direction closely upstream of the transcription start sites (TSs) of their associated genes. PROM PT TSs share features with mRNA-producing TSs, including stalled RNA polymerase II (RNAPII) and the production of small TS-associated RNAs. Notably, motif analyses around PROM PT 3' ends reveal polyadenylation (pA)-like signals. Mutagenesis studies demonstrate that PROM PT pA signals are functional but linked to RNA degradation. Moreover, pA signals are under-represented in promoter-downstream versus promoter-upstream regions, thus allowing for more efficient RNAPII progress in the sense direction from gene promoters. We conclude that asymmetric sequence distribution around human gene promoters serves to provide a directional RNA output from an otherwise bidirectional transcription process.