Multi-species microarrays reveal the effect of sequence divergence on gene expression profiles

Abstract

Interspecies comparisons of gene expression levels will increase our understanding of the evolution of transcriptional mechanisms and help to identify targets of natural selection. This approach holds particular promise for apes, as many human-specific adaptations are thought to result from differences in gene expression rather than in coding sequence. To date, however, all studies directly comparing interspecies gene expression have been performed on single-species arrays, so that it has been impossible to distinguish differential hybridization due to sequence mismatches from underlying expression differences. To evaluate the severity of this potential problem, we constructed a new multiprimate cDNA array using probes from human, chimpanzee, orangutan, and rhesus. We find a large effect of sequence divergence on hybridization signal, even in the closest pair of species, human and chimpanzee. By comparing single-species array analyses with results from multispecies arrays, we examine how estimates of differential gene expression are affected by sequence divergence. Our results indicate that naive use of single-species arrays in direct interspecies comparisons can yield spurious results.

Expression profiling in primates reveals a rapid evolution of human transcription factors

“…All probes on the array were adjusted by the fitted lowess curve (see Supplementary Methods). We concentrated on the 907 genes on the array for which successful polymerase chain reaction (PCR) products were obtained from all species 14 . The expression log ratios for each gene were analysed using the linear mixed model:…”

“…To make comparisons between more distantly related primate species, we generated a multi-species cDNA array that allows comparison of gene expression between species without the confounding effects of sequence divergence 14 . This cDNA array contains probes for 1,056 orthologous genes from four species (see Supplementary Methods) 14 .We used this array to compare gene expression profiles in the livers of humans, chimpanzees (Pan troglodytes), orangutans (Pongo pygmaeus) and rhesus macaques (Macaca mulatta), the phylogeny of which represents approximately 70 million years (Myr) of evolution. By assigning expression changes in the liver to particular lineages, we were able to identify the first set of genes for which regulation seems to be under lineage-specific selection pressures.…”

“…To make comparisons between more distantly related primate species, we generated a multi-species cDNA array that allows comparison of gene expression between species without the confounding effects of sequence divergence 14 . This cDNA array contains probes for 1,056 orthologous genes from four species (see Supplementary Methods) 14 .…”

Genomic signatures of diet-related shifts during human origins

“…Cases where a lineage-specific shift in gene expression levels is seen in conjunction with low variance within the species are suggestive of directional selection driving that change [68,86]. There are clear changes in gene expression in humans, compared with non-human primates, that are centred in metabolic pathways in a number of different tissues [67 -71], and some of these may be changes owing to directional selection.…”

Deep homology in the age of next-generation sequencing

“…Variation in hybridization efficiencies, due to species-inherent sequence differences in the targeted parts of mRNAs, and other technical issues made direct interpretations difficult. A number of normalization and analytics procedures helped to bypass these problems [36,37]. However, most of these shortcomings can now easily be avoided, thanks to the development of high-throughput next-generation sequencing (NGS) techniques [38].…”