Abstract
BackgroundAccurate annotation of translation initiation sites (TISs) is essential for understanding the translation initiation mechanism. However, the reliability of TIS annotation in widely used databases such as RefSeq is uncertain due to the lack of experimental benchmarks.ResultsBased on a homogeneity assumption that gene translation-related signals are uniformly distributed across a genome, we have established a computational method for a large-scale quantitative assessment of the reliability of TIS annotations for any prokaryotic genome. The method consists of modeling a positional weight matrix (PWM) of aligned sequences around predicted TISs in terms of a linear combination of three elementary PWMs, one for true TIS and the two others for false TISs. The three elementary PWMs are obtained using a reference set with highly reliable TIS predictions. A generalized least square estimator determines the weighting of the true TIS in the observed PWM, from which the accuracy of the prediction is derived. The validity of the method and the extent of the limitation of the assumptions are explicitly addressed by testing on experimentally verified TISs with variable accuracy of the reference sets. The method is applied to estimate the accuracy of TIS annotations that are provided on public databases such as RefSeq and ProTISA and by programs such as EasyGene, GeneMarkS, Glimmer 3 and TiCo. It is shown that RefSeq's TIS prediction is significantly less accurate than two recent predictors, Tico and ProTISA. With convincing proofs, we show two general preferential biases in the RefSeq annotation, i.e. over-annotating the longest open reading frame (LORF) and under-annotating ATG start codon. Finally, we have established a new TIS database, SupTISA, based on the best prediction of all the predictors; SupTISA has achieved an average accuracy of 92% over all 532 complete genomes.ConclusionLarge-scale computational evaluation of TIS annotation has been achieved. A new TIS database much better than RefSeq has been constructed, and it provides a valuable resource for further TIS studies.
Highlights
Accurate annotation of translation initiation sites (TISs) is essential for understanding the translation initiation mechanism
We create a series of partially accurate O with accuracy α from 40% to 90% at a step of 10% by replacing 100(1 α)% of the true TISs by randomly choosing false TISs
A positional weight matrix (PWM) of aligned sequences around TIS summarizes the statistical information of the signal, and is a tool to use for study how much, in a given set of annotation, the true signal has contributed
Summary
Accurate annotation of translation initiation sites (TISs) is essential for understanding the translation initiation mechanism. The sequence upstream to the TIS, the start codon itself and the sequence downstream to the TIS show specific patterns which differ from genome to genome. The sequence at about 20 bps upstream to the TIS in most prokaryotic genes contains primarily purine rich Shine-Dalgarno sequence [1]. Genome-wide computational analysis on leaderless genes revealed A/T rich sequences in a region at about 30 bps further upstream [3]. With the aid of a sequence logo tool, Torarinsson et al [3] and Zhu et al [5] reported the variation of sequence patterns among dozens of archaeal genomes, which shed light on the understanding of the divergence of translation initiation mechanisms in prokaryote
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