The identification of untranslated regions, introns, and coding regions within an organism remains challenging. We developed a quantitative sequencing-based method called RNA-Seq for mapping transcribed regions, in which complementary DNA fragments are subjected to high-throughput sequencing and mapped to the genome. We applied RNA-Seq to generate a high-resolution transcriptome map of the yeast genome and demonstrated that most (74.5%) of the nonrepetitive sequence of the yeast genome is transcribed. We confirmed many known and predicted introns and demonstrated that others are not actively used. Alternative initiation codons and upstream open reading frames also were identified for many yeast genes. We also found unexpected 3'-end heterogeneity and the presence of many overlapping genes. These results indicate that the yeast transcriptome is more complex than previously appreciated.

DNA sequence has long been recognized as an important contributor to nucleosome positioning, which has the potential to regulate access to genes. The extent to which the nucleosomal architecture at promoters is delineated by the underlying sequence is now being worked out. Here we use comparative genomics to report a genome-wide map of nucleosome positioning sequences (NPSs) located in the vicinity of all Saccharomyces cerevisiae genes. We find that the underlying DNA sequence provides a very good predictor of nucleosome locations that have been experimentally mapped to a small fraction of the genome. Notably, distinct classes of genes possess characteristic arrangements of NPSs that may be important for their regulation. In particular, genes that have a relatively compact NPS arrangement over the promoter region tend to have a TATA box buried in an NPS and tend to be highly regulated by chromatin modifying and remodeling factors.

Although histones can form nucleosomes on virtually any genomic sequence, DNA sequences show considerable variability in their binding affinity. We have used DNA sequences of Saccharomyces cerevisiae whose nucleosome binding affinities have been experimentally determined (Yuan et al. 2005) to train a support vector machine to identify the nucleosome formation potential of any given sequence of DNA. The DNA sequences whose nucleosome formation potential are most accurately predicted are those that contain strong nucleosome forming or inhibiting signals and are found within nucleosome length stretches of genomic DNA with continuous nucleosome formation or inhibition signals. We have accurately predicted the experimentally determined nucleosome positions across a well-characterized promoter region of S. cerevisiae and identified strong periodicity within 199 center-aligned mononucleosomes studied recently (Segal et al. 2006) despite there being no periodicity information used to train the support vector machine. Our analysis suggests that only a subset of nucleosomes are likely to be positioned by intrinsic sequence signals. This observation is consistent with the available experimental data and is inconsistent with the proposal of a nucleosome positioning code. Finally, we show that intrinsic nucleosome positioning signals are both more inhibitory and more variable in promoter regions than in open reading frames in S. cerevisiae.