A large-scale full-length cDNA analysis to explore the budding yeast transcriptome

Abstract

We performed a large-scale cDNA analysis to explore the transcriptome of the budding yeast Saccharomyces cerevisiae. We sequenced two cDNA libraries, one from the cells exponentially growing in a minimal medium and the other from meiotic cells. Both libraries were generated by using a vector-capping method that allows the accurate mapping of transcription start sites (TSSs). Consequently, we identified 11,575 TSSs associated with 3,638 annotated genomic features, including 3,599 ORFs, to suggest that most yeast genes have two or more TSSs. In addition, we identified 45 previously undescribed introns, including those affecting current ORF annotations and those spliced alternatively. Furthermore, the analysis revealed 667 transcription units in the intergenic regions and transcripts derived from antisense strands of 367 known features. We also found that 348 ORFs carry TSSs in their 3-halves to generate sense transcripts starting from inside the ORFs. These results indicate that the budding yeast transcriptome is considerably more complex than previously thought, and it shares many recently revealed characteristics with the transcriptomes of mammals and other higher eukaryotes. Thus, the genome-wide active transcription that generates novel classes of transcripts appears to be an intrinsic feature of the eukaryotic cells. The budding yeast will serve as a versatile model for the studies on these aspects of transcriptome, and the full-length cDNA clones can function as an invaluable resource in such studies.alternative splicing ͉ antisense transcript ͉ noncoding RNA ͉ transcription start site

Interactions between Mei4, Rec114, and other proteins required for meiotic DNA double-strand break formation in Saccharomyces cerevisiae

“…5c), and the expected sequence at the splice junction was confirmed by sequencing of the RT-PCR product (data not shown). Splicing of the REC102 message was also demonstrated by large-scale microarray and RT-PCR analyses in recent independent studies (Miura et al 2006;Juneau et al 2007). Interestingly, splicing was relatively inefficient in that the intron had been removed in only ∼70% of the amplified message from RNA samples isolated 6 hr after transfer to sporulation medium (Fig.…”

Regulated post-transcriptional RNA cleavage diversifies the eukaryotic transcriptome

“…Indeed, it seems likely that post-transcriptional cleavage is an ancient mechanism that occurs (in at least some form) throughout both prokaryotic and eukaryotic lineages. In support of this, it was recently reported that the 59 termini of ;7% of full-length cDNAs initiate from within the coding sequence of Saccharomyces cerevisiae genes (Miura et al 2006), raising the possibility that post-transcriptional cleavage is an ancient mechanism of RNA processing that is conserved throughout the eukaryotic lineage. Although we were unable to identify any compelling instances of full-length cDNA within this subset that mapped proximally across EEJs, this was perhaps not surprising given the relative scarcity of splice junctions in yeast.…”

Stable and dynamic nucleosome states during a meiotic developmental process

“…Tag counts located between 64 kb of 6576 annotated TSSs were collected in 1-bp bins. The TSS/TES annotation was compiled from existing TSS/TES resources (David et al 2006;Miura et al 2006;Nagalakshmi et al 2008;Xu et al 2009). Each H3 data set was scaled to set total counts to be equal.…”