Motivation: Genes with identical patterns of occurrence across the phyla tend to function together in the same protein complexes or participate in the same biochemical pathway. However, the requirement that the profiles be identical (i) severely restricts the number of functional links that can be established by such phylogenetic profiling; (ii) limits detection to very strong functional links, failing to capture relations between genes that are not in the same pathway, but nevertheless subserve a common function and (iii) misses relations between analogous genes. Here we present and apply a method for relaxing the restriction, based on the probability that a given arbitrary degree of similarity between two profiles would occur by chance, with no biological pressure. Function is then inferred at any desired level of confidence.
Results: We derive an expression for the probability distribution of a given number of chance co-occurrences of a pair of non-homologous orthologs across a set of genomes. The method is applied to 2905 clusters of orthologous genes (COGs) from 44 fully sequenced microbial genomes representing all three domains of life. Among the results are the following. (1) Of the 51 000 annotated intrapathway gene pairs, 8935 are linked at a level of significance of 0.01. This is over 30-fold greater than the 271 intrapathway pairs obtained at the same confidence level when identical profiles are used. (2) Of the 540 000 interpathway genes pairs, some 65 000 are linked at the 0.01 level of significance, some 12 standard deviations beyond the number expected by chance at this confidence level. We speculate that many of these links involve nearest-neighbor path, and discuss some examples. (3) The difference in the percentage of linked interpathway and intrapathway genes is highly significant, consistent with the intuitive expectation that genes in the same pathway are generally under greater selective pressure than those that are not. (4) The method appears to recover well metabolic networks. This is illustrated by the TCA cycle which is recovered as a highly connected, weighted edge network of 30 of its 31 COGs. (5) The fraction of pairs having a common pathway is a symmetric function of the Hamming distance between their profiles. This finding, that the functional correlation between profiles with near maximum Hamming distance is as large as between profiles with near zero Hamming distance, and as statistically significant, is plausibly explained if the former group represents analogous genes.