Project description:We report the identification of potential novel RNA targets for box C/D snoRNAs in budding yeast, using the approach of UV crosslinking and sequencing of hybrids (CLASH).
Project description:We report the identification of potential novel RNA targets for box C/D snoRNAs in human HEK293 cells, using the approaches of UV crosslinking and sequencing of hybrids (CLASH), and formaldehyde crosslinking and sequencing of hybrids (FLASH).
Project description:Eukaryotic RNA polymerase II (RNAPII) transcribes mRNA genes and non-protein coding RNAs (ncRNAs) including small nuclear and nucleolar RNAs (sn/snoRNAs). In metazoans, RNAPII transcription of sn/snoRNAs is facilitated by a number of specialized complexes, but no such complexes have been discovered in yeast. It has been proposed that yeast sn/snoRNA promoters use the same factors as mRNA promoters, but the extent to which regulators of mRNA genes function at yeast sn/snoRNA genes is unclear. Here, we investigated a potential role for the Mediator complex, essential for mRNA gene transcription, in the transcription of sn/snoRNA genes. We found that the Mediator maps to most sn/snoRNA gene regulatory regions and that rapid depletion of the essential structural subunit Med14 strongly reduces RNAPII and TFIIB occupancy as well as nascent transcription of sn/snoRNA genes. Deletion of Med3 and Med15, subunits of the activator-interacting Mediator tail module, does not affect Mediator recruitment to or RNAPII and TFIIB occupancy of sn/snoRNA genes. Our analyses suggest that Mediator promotes PIC formation and transcription at sn/snoRNA genes, expanding the role of this critical regulator beyond its known functions in mRNA gene transcription and demonstrating further mechanistic similarity between the transcription of mRNA and sn/snoRNA genes.