Genome-Wide Prediction of Transcription Start Sites in Conifers.

The identification of promoters is an essential step in the genome annotation process, providing a framework for gene regulatory networks and their role in transcription regulation. Despite considerable advances in the high-throughput determination of transcription start sites (TSSs) and transcription factor binding sites (TFBSs), experimental methods are still time-consuming and expensive. Instead, several computational approaches have been developed to provide fast and reliable means for predicting the location of TSSs and regulatory motifs on a genome-wide scale. Numerous studies have been carried out on the regulatory elements of mammalian genomes, but plant promoters, especially in gymnosperms, have been left out of the limelight and, therefore, have been poorly investigated. The aim of this study was to enhance and expand the existing genome annotations using computational approaches for genome-wide prediction of TSSs in the four conifer species: loblolly pine, white spruce, Norway spruce, and Siberian larch. Our pipeline will be useful for TSS predictions in other genomes, especially for draft assemblies, where reliable TSS predictions are not usually available. We also explored some of the features of the nucleotide composition of the predicted promoters and compared the GC properties of conifer genes with model monocot and dicot plants. Here, we demonstrate that even incomplete genome assemblies and partial annotations can be a reliable starting point for TSS annotation. The results of the TSS prediction in four conifer species have been deposited in the Persephone genome browser, which allows smooth visualization and is optimized for large data sets. This work provides the initial basis for future experimental validation and the study of the regulatory regions to understand gene regulation in gymnosperms.

Insights into corn genes derived from large-scale cDNA sequencing.

We present a large portion of the transcriptome of Zea mays, including ESTs representing 484,032 cDNA clones from 53 libraries and 36,565 fully sequenced cDNA clones, out of which 31,552 clones are non-redundant. These and other previously sequenced transcripts have been aligned with available genome sequences and have provided new insights into the characteristics of gene structures and promoters within this major crop species. We found that although the average number of introns per gene is about the same in corn and Arabidopsis, corn genes have more alternatively spliced isoforms. Examination of the nucleotide composition of coding regions reveals that corn genes, as well as genes of other Poaceae (Grass family), can be divided into two classes according to the GC content at the third position in the amino acid encoding codons. Many of the transcripts that have lower GC content at the third position have dicot homologs but the high GC content transcripts tend to be more specific to the grasses. The high GC content class is also enriched with intronless genes. Together this suggests that an identifiable class of genes in plants is associated with the Poaceae divergence. Furthermore, because many of these genes appear to be derived from ancestral genes that do not contain introns, this evolutionary divergence may be the result of horizontal gene transfer from species not only with different codon usage but possibly that did not have introns, perhaps outside of the plant kingdom. By comparing the cDNAs described herein with the non-redundant set of corn mRNAs in GenBank, we estimate that there are about 50,000 different protein coding genes in Zea. All of the sequence data from this study have been submitted to DDBJ/GenBank/EMBL under accession numbers EU940701-EU977132 (FLI cDNA) and FK944382-FL482108 (EST).

Features of Arabidopsis genes and genome discovered using full-length cDNAs.

Arabidopsis is currently the reference genome for higher plants. A new, more detailed statistical analysis of Arabidopsis gene structure is presented including intron and exon lengths, intergenic distances, features of promoters, and variant 5'-ends of mRNAs transcribed from the same transcription unit. We also provide a statistical characterization of Arabidopsis transcripts in terms of their size, UTR lengths, 3'-end cleavage sites, splicing variants, and coding potential. These analyses were facilitated by scrutiny of our collection of sequenced full-length cDNAs and much larger collection of 5'-ESTs, together with another set of full-length cDNAs from Salk/Stanford/Plant Gene Expression Center/RIKEN. Examples of alternative splicing are observed for transcripts from 7% of the genes and many of these genes display multiple spliced isoforms. Most splicing variants lie in non-coding regions of the transcripts. Non-canonical splice sites constitute less than 1% of all splice sites. Genes with fewer than four introns display reduced average mRNA levels. Putative alternative transcription start sites were observed in 30% of highly expressed genes and in more than 50% of the genes with low expression. Transcription start sites correlate remarkably well with a CG skew peak in the DNA sequences. The intergenic distances vary considerably, those where genes are transcribed towards one another being significantly shorter. New transcripts, missing in the current TIGR genome annotation and ESTs that are non-coding, including those antisense to known genes, are derived and cataloged in the Supplementary Material. They identify 148 new loci in the Arabidopsis genome. The conclusions drawn provide a better understanding of the Arabidopsis genome and how the gene transcripts are processed. The results also allow better predictions to be made for, as yet, poorly defined genes and provide a reference for comparisons with other plant genomes whose complete sequences are currently being determined. Some comparisons with rice are included in this paper.