The Cell Cycle–Regulated Genes of Schizosaccharomyces pombe

Many genes are regulated as an innate part of the eukaryotic cell cycle, and a complex transcriptional network helps enable the cyclic behavior of dividing cells. This transcriptional network has been studied in Saccharomyces cerevisiae (budding yeast) and elsewhere. To provide more perspective on these regulatory mechanisms, we have used microarrays to measure gene expression through the cell cycle of Schizosaccharomyces pombe (fission yeast). The 750 genes with the most significant oscillations were identified and analyzed. There were two broad waves of cell cycle transcription, one in early/mid G2 phase, and the other near the G2/M transition. The early/mid G2 wave included many genes involved in ribosome biogenesis, possibly explaining the cell cycle oscillation in protein synthesis in S. pombe. The G2/M wave included at least three distinctly regulated clusters of genes: one large cluster including mitosis, mitotic exit, and cell separation functions, one small cluster dedicated to DNA replication, and another small cluster dedicated to cytokinesis and division. S. pombe cell cycle genes have relatively long, complex promoters containing groups of multiple DNA sequence motifs, often of two, three, or more different kinds. Many of the genes, transcription factors, and regulatory mechanisms are conserved between S. pombe and S. cerevisiae. Finally, we found preliminary evidence for a nearly genome-wide oscillation in gene expression: 2,000 or more genes undergo slight oscillations in expression as a function of the cell cycle, although whether this is adaptive, or incidental to other events in the cell, such as chromatin condensation, we do not know

Sorting with Fixed-Length Reversals

this paper, we study the problem of sorting permutations and circular permutations using as few fixed-length reversals as possible. Our problem is implicit in the popular TOP-SPI

Re-annotation of 12,495 prokaryotic 16S rRNA 3' ends and analysis of Shine-Dalgarno and anti-Shine-Dalgarno sequences.

We examined 20,648 prokaryotic unique taxids with respect to the annotation of the 3' end of the 16S rRNA, which contains the anti-Shine-Dalgarno sequence. We used the sequence of highly conserved helix 45 of the 16S rRNA as a guide. By this criterion, 8,153 annotated 3' ends correctly included the anti-Shine-Dalgarno sequence, but 12,495 were foreshortened or otherwise mis-annotated, missing part or all of the anti-Shine-Dalgarno sequence, which immediately follows helix 45. We re-annotated, giving a total of 20,648 16S rRNA 3' ends. The vast majority indeed contained a consensus anti-Shine-Dalgarno sequence, embedded in a highly conserved 13 base "tail". However, 128 exceptional organisms had either a variant anti-Shine-Dalgarno, or no recognizable anti-Shine-Dalgarno, in their 16S rRNA(s). For organisms both with and without an anti-Shine-Dalgarno, we identified the Shine-Dalgarno motifs actually enriched in front of each organism's open reading frames. This showed to what extent the Shine-Dalgarno motifs correlated with anti-Shine Dalgarno motifs. In general, organisms whose rRNAs lacked a perfect anti-Shine-Dalgarno motif also lacked a recognizable Shine-Dalgarno. For organisms whose 16S rRNAs contained a perfect anti-Shine-Dalgarno motif, a variety of results were obtained. We found one genus, Alteromonas, where several taxids apparently maintain two different types of 16S rRNA genes, with different, but conserved, antiSDs. The fact that some organisms do not seem to have or use Shine-Dalgarno motifs supports the idea that prokaryotes have other robust mechanisms for recognizing start codons for translation

Measurement of average decoding rates of the 61 sense codons in vivo

Abstract Most amino acids can be encoded by several synonymous codons, which are used at unequal frequencies. The significance of unequal codon usage remains unclear. One hypothesis is that frequent codons are translated relatively rapidly. However, there is little direct, in vivo, evidence regarding codon-specific translation rates. In this study, we generate high-coverage data using ribosome profiling in yeast, analyze using a novel algorithm, and deduce events at the A-and P-sites of the ribosome. Different codons are decoded at different rates in the A-site. In general, frequent codons are decoded more quickly than rare codons, and AT-rich codons are decoded more quickly than GC-rich codons. At the P-site, proline is slow in forming peptide bonds. We also apply our algorithm to short footprints from a different conformation of the ribosome and find strong amino acid-specific (not codon-specific) effects that may reflect interactions with the exit tunnel of the ribosome

Prokaryotic coding regions have little if any specific depletion of Shine-Dalgarno motifs.

The Shine-Dalgarno motif occurs in front of prokaryotic start codons, and is complementary to the 3' end of the 16S ribosomal RNA. Hybridization between the Shine-Dalgarno sequence and the anti-Shine-Dalgarno region of the16S rRNA (CCUCCU) directs the ribosome to the start AUG of the mRNA for translation. Shine-Dalgarno-like motifs (AGGAGG in E. coli) are depleted from open reading frames of most prokaryotes. This may be because hybridization of the 16S rRNA at Shine-Dalgarnos inside genes would slow translation or induce internal initiation. However, we analyzed 128 species from diverse phyla where the 16S rRNA gene(s) lack the anti-Shine-Dalgarno sequence, and so the 16S rRNA is incapable of interacting with Shine-Dalgarno-like sequences. Despite this lack of an anti-Shine-Dalgarno, half of these species still displayed depletion of Shine-Dalgarno-like sequences when analyzed by previous methods. Depletion of the same G-rich sequences was seen by these methods even in eukaryotes, which do not use the Shine-Dalgarno mechanism. We suggest previous methods are partly detecting a non-specific depletion of G-rich sequences. Alternative informatics approaches show that most prokaryotes have only slight, if any, specific depletion of Shine-Dalgarno-like sequences from open reading frames. Together with recent evidence that ribosomes do not pause at ORF-internal Shine-Dalgarno motifs, these results suggest the presence of ORF-internal Shine-Dalgarno-like motifs may be inconsequential, perhaps because internal regions of prokaryotic mRNAs may be structurally "shielded" from translation initiation

Oscillation of Ribosome Biogenesis Genes in <i>S. cerevisiae</i>

<p>One cell cycle of elutriation data is shown for 52 <i>S. cerevisiae</i> genes involved in ribosome biogenesis. The genes chosen for analysis were those listed [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030225#pbio-0030225-b42" target="_blank">42</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030225#pbio-0030225-b75" target="_blank">75</a>] as involved in ribosome biogenesis. At the top of <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030225#pbio-0030225-g011" target="_blank">Figure 11</a> are three histone genes <i>(HTA2, HHF1,</i> and <i>HHT10)</i> known to peak in S, and three genes <i>(CLN1, CLN2,</i> and <i>MCD1)</i> known to peak in late G1. The raw data for this figure are taken from Spellman et al. [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030225#pbio-0030225-b06" target="_blank">6</a>]. In their experiment, cells were grown in ethanol medium, and then small G1 cells were isolated by elutriation and re-inoculated into ethanol medium. Samples were taken at intervals from 0 to 390 min, the duration of one cell cycle under these conditions.</p

Cell Cycle–Regulated Genes Ordered by Time of Peak Expression

<div><p>(A) Expression data for the top 750 genes is shown, with genes ordered by time of peak expression. Every row represents a gene; every column represents an array from a time-course experiment. Red signifies up-regulation (i.e., an experiment/control ratio greater than one); green signifies down-regulation (i.e., an experiment/control ratio less than one). Black is a ratio close to one, and grey is missing data. Dynamic range is 16-fold from reddest red to greenest green. The time in hours since the beginning of the time course is shown in black numerals at the top of <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030225#pbio-0030225-g003" target="_blank">Figure 3</a>. The peaks in septation index are marked with purple rectangles at the top and bottom of the figure. Genes from defined clusters are marked on the left by colored lines, according to the cluster color code shown at the bottom of the figure.</p> <p>(B) As (A), but only the 514 genes found in our study but not found by Rustici et al. [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030225#pbio-0030225-b07" target="_blank">7</a>] are shown.</p></div

Distribution of Promoter Motifs

<div><p>A total of 23 genes from the core of the Cdc15 cluster (cdc15), the 18 genes from the Cdc18 cluster (cdc18), the nine genes from the Eng1 cluster, plus one similarly regulated gene (<i>SPBC83.18c</i>; eng1) and 15 randomly chosen genes (Random) had their promoters examined for six sequence motifs using SpikeChart. For each gene, the DNA sequence examined was the 2,000 bp immediately upstream of the Start codon; the Start codon is at the right edge of the figure, and the upstream 2,000 bp extend to the left. The beginning of the next upstream open reading frame is indicated by a triangle; for instance, for the <i>pof3</i> gene (Cdc18 cluster), the next open reading frame begins about 700 bp upstream of the <i>pof3</i> Start codon. For the Cdc18 cluster gene <i>ams2,</i> all 2,000 bp are intergenic.</p> <p>Consensus motifs are as follows: Dark Blue Fkh motif, TGTAAACAAA; Purple Ace2 motif, ACCAGCCT; Green MBF motif, GACGCGTC; Black Dbl10 motif, ACGCGACGCG; Light Blue/Aquamarine New 1v motif, TGACAAC; Yellow New 3v motif, (A/T)ACC(A/T)CG(T/C)(A/T)(C/A)C. </p> <p>Taller spikes indicate a better match to the consensus; the weight matrices, and the rules governing spike height, are given in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030225#st002" target="_blank">Table S2</a>. Dbl10 spikes (black) are obscured by overlapping MBF spikes (green), and so are hard to see. Only tall spikes (i.e., good matches to the consensus) are shown, so an acceptable motif may exist even when no spike is shown,</p></div