GSE67163
KDM5 Lysine Demethylases Are Involved in Maintenance of 3'UTR Length [RNA-Seq]
We discovered that the Saccharomyces cerevisiae lysine demethylase, Jhd2 (also known as KDM5), recruits 3'UTR processing machinery and promotes alteration of 3'UTR length in a demethylase-dependent manner. Interaction of Jhd2 with both chromatin and RNA suggests that Jhd2 affects selection of polyadenylation sites through a transcription-coupled mechanism. Overall design: For 3'READs analysis, wild-type yeast or yeast with JHD2 deleted were grown to mid-log phase and RNA was extracted as mentioned above.  cDNA libraries enriched for 3'UTRs were prepared as previously published and as noted in Figure 4a [28].  Samples were then subjected to RNA sequencing on the Illumina Hiseq 2000 using 50bp single end reads.  Data was analyzed as previously [Hoque M] with the following modifications:   The adapter sequences were trimmed off the single-ended reads.  The remaining reads longer than 15 bp were mapped to yeast genome SacCer3 using bowtie2 [Langmead B 2012] with the setting '-5 4 --local', trimming off 4bp from 5' and allowing soft clipping (S) at both ends.  The alignments were then filtered by 1) MAPQ>=10, 2) mismatches <= 5%, 3) unaligned 5' Ts >=2 to get the PASS reads. The last aligned positions of the PASS reads were grouped to pA clusters with a clustering size of 24 bp.  Each pA cluster was assigned to one of the genes defined by SGD [Cherry JM].  The 3' end of a gene was extended 1000bp from the end of CDS until it overlaps with another gene in the same direction.  The pA Clusters were then filtered by contains 1) >= 3 PASS reads 2) >= 5% of all PASS reads mapped to the gene.