Transcription of the mammalian genome is pervasive, but productive transcription outside of protein-coding genes is limited by unknown mechanisms1. In particular, although RNA polymerase II (RNAPII) initiates divergently from most active gene promoters, productive elongation occurs primarily in the sense-coding direction2,3,4. Here we show in mouse embryonic stem cells that asymmetric sequence determinants flanking gene transcription start sites control promoter directionality by regulating promoter-proximal cleavage and polyadenylation. We find that upstream antisense RNAs are cleaved and polyadenylated at poly(A) sites (PASs) shortly after initiation. De novo motif analysis shows PAS signals and U1 small nuclear ribonucleoprotein (snRNP) recognition sites to be the most depleted and enriched sequences, respectively, in the sense direction relative to the upstream antisense direction. These U1 snRNP sites and PAS sites are progressively gained and lost, respectively, at the 5′ end of coding genes during vertebrate evolution. Functional disruption of U1 snRNP activity results in a dramatic increase in promoter-proximal cleavage events in the sense direction with slight increases in the antisense direction. These data suggest that a U1–PAS axis characterized by low U1 snRNP recognition and a high density of PASs in the upstream antisense region reinforces promoter directionality by promoting early termination in upstream antisense regions, whereas proximal sense PAS signals are suppressed by U1 snRNP. We propose that the U1–PAS axis limits pervasive transcription throughout the genome.
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The authors acknowledge the service to the MIT community of the late Sean Collier. We would like to thank N. Spies for generously sharing his optimized 3′-end sequencing protocol, C. Lin for providing computational assistance, M. Lindstrom for assistance on constructing Supplementary Figure 11, and S. Chen, A. Chiu, M. Jangi, Q. Liu, J. Wilusz and J. Zamudio for reading of the manuscript. We also thank the Core Facility in the Swanson Biotechnology Center at the David H. Koch Institute for Integrative Cancer Research at MIT for their assistance with high-throughput sequencing. This work was supported by United States Public Health Service grants RO1-GM34277 and R01-CA133404 from the National Institutes of Health (P.A.S.), partially by Cancer Center Support (core) grant P30-CA14051 from the National Cancer Institute, and by a Public Health Service research grant (GM-085319) from the National Institute of General Medical Sciences (C.B.B.). X.W. is a Howard Hughes Medical Institute International Student Research fellow.
The authors declare no competing financial interests.
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Almada, A., Wu, X., Kriz, A. et al. Promoter directionality is controlled by U1 snRNP and polyadenylation signals. Nature 499, 360–363 (2013). https://doi.org/10.1038/nature12349
Screening thousands of transcribed coding and non-coding regions reveals sequence determinants of RNA polymerase II elongation potential
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