pANT: a method for the pairwise assessment of nonfunctionalization times of processed pseudogenes.

We present a method for pairwise Assessment of Nonfunctionalization Times (pANT) in processed pseudogenes. Contrary to existing methods for estimating nonfunctionalization times, pANT utilizes previously calculated probabilities of nucleotide substitution as explicit rate measurements, rather than assume that the substitution rates are the same for all nucleotides. Thus, the method allows a more accurate computation of the time that has elapsed since the nonfunctionalization of a pseudogene. Whereas existing methods require the sequence of an orthologous functional gene, which is not always at hand, pANT only uses the pairwise alignment of the gene/pseudogene pair, thus expanding the range of problems that can be tackled. To estimate evolutionary times in nonfunctional sequences, pANT measures the differences in the pairwise alignment of a gene and its paralogous processed pseudogene, using only the first and second codon positions. It assumes that, because of functional constraints, these positions in the sequence of the functional homolog have not changed since the time of nonfunctionalization of the pseudogene. Hence, the sequence of the gene may be used as the ancestor of the pseudogene. We show that the method's reliance on a detailed substitution matrix, which is derived separately for each species, makes it more accurate than existing methods. We applied pANT to the case of the unitary alpha-1,3-galactosyltransferase human pseudogene and found that our estimate of the nonfunctionalization time was in agreement with that obtained by taxonomic and paleontological considerations pertaining to the divergence between platyrrhines (New World monkeys) and cattarhines (Old World monkeys).