Pervasive adaptive evolution in primate seminal proteins

Abstract

Seminal fluid proteins show striking effects on reproduction, involving manipulation of female behavior and physiology, mechanisms of sperm competition, and pathogen defense. Strong adaptive pressures are expected for such manifestations of sexual selection and host defense, but the extent of positive selection in seminal fluid proteins from divergent taxa is unknown. We identified adaptive evolution in primate seminal proteins using genomic resources in a tissue-specific study. We found extensive signatures of positive selection when comparing 161 human seminal fluid proteins and 2,858 prostate-expressed genes to those in chimpanzee. Seven of eight outstanding genes yielded statistically significant evidence of positive selection when analyzed in divergent primates. Functional clues were gained through divergent analysis, including several cases of species-specific loss of function in copulatory plug genes, and statistically significant spatial clustering of positively selected sites near the active site of kallikrein 2. This study reveals previously unidentified positive selection in seven primate seminal proteins, and when considered with findings in Drosophila, indicates that extensive positive selection is found in seminal fluid across divergent taxonomic groups.

Figure 1. Plots of d _N Versus d _S for Primate and Drosophila Seminal Fluid Genes

(A) Genes encoding seminal fluid proteins identified by mass spectrometry in human versus chimpanzee. (B) Drosophila simulans male-specific accessory gland genes versus D. melanogaster [2]. The diagonal represents neutral evolution, a d _N/d _S ratio of one. Most genes are subject to purifying selection and fall below the diagonal, while several genes fall above or near the line suggesting positive selection. Comparison of the two plots shows elevated d _N/d _S ratios in seminal fluid genes of both taxonomic groups.

Figure 2. Variable Selective Pressure is Seen Between Lineages for Semen Coagulum Protein TGM4

This primate phylogeny shows selective pressure on TGM4 with estimated d _N/d _S ratios indicated on branches. Ratios greater than one are suggestive of either relaxed constraint or positive selection. Ratios are only shown for long branches, those with at least eight substitutions. A null model with a uniform d _N/d _S ratio across all lineages is rejected in favor of these estimates (p = 0.003). Branch lengths are estimated from TGM4 coding sequences. NWM, New World monkeys; OWM, Old World monkeys.

Figure 3. Positive Selection at Sites Involved in Substrate Binding in KLK2

(A) Several amino acid sites predicted to be under positive selection (red) are near the protease active site (yellow). Three selected sites are found in known structural components of kallikrein proteins (light blue residues): Gly191 is part of the S1 substrate binding pocket, and His89 and Gln90 are part of the kallikrein loop [30]. Selected sites are labeled with the human residue on this threaded model. (B) Positively selected sites are significantly clustered on the surface of KLK2. The observed mean pairwise distance between predicted positively selected sites is significantly lower than random sets of surface sites (p = 0.0043). This spatial clustering suggests that positive selection acted during KLK2 evolution to alter substrate binding.

Figure 4. Positively Selected Sites on MSMB are Spread across the Protein Surface.

According to sites models of codon evolution, 42% of MSMB residues experienced adaptive pressure to alter their amino acids. Those predicted with high support are shown in red on this threaded structural model of human MSMB. Blue and purple residues demarcate two structural domains of the protein [62]. The amino acid sites show no clustering and are almost significantly dispersed throughout the protein (p = 0.066). This pattern is quite different from that shown by KLK2 (Figure 3). Although MSMB is one of the most abundant human seminal proteins, its function remains unknown.