Species specific exome probes reveal new insights in positively selected genes in nonhuman primates

Nonhuman primates (NHP) are important biomedical animal models for the study of human disease. Of these, the most widely used models in biomedical research currently are from the genus Macaca. However, evolutionary genetic divergence between human and NHP species makes human-based probes inefficient for the capture of genomic regions of NHP for sequencing and study. Here we introduce a new method to resequence the exome of NHP species by a designed capture approach specifically targeted to the NHP, and demonstrate its superior performance on four NHP species or subspecies. Detailed investigation on biomedically relevant genes demonstrated superior capture by the new approach. We identified 28 genes that appeared to be pseudogenized and inactivated in macaque. Finally, we identified 187 genes showing strong evidence for positive selection across all branches of the primate phylogeny including many novel findings

Two to Tango: Co-evolution of Hominid Natural Killer Cell Receptors and MHC

Natural killer (NK) cells have diverse roles in hominid immunity and reproduction. Modulating these functions are the interactions between major histocompatibility complex (MHC) class I molecules that are ligands for two NK cell surface receptor types. Diverse killer cell immunoglobulin-like receptors (KIR) bind specific motifs encoded within the polymorphic MHC class I cell surface glycoproteins, while, in more conserved interactions, CD94:NKG2A receptors recognize MHC-E with bound peptides derived from MHC class I leader sequences. The hominid lineage presents a choreographed co-evolution of KIR with their MHC class I ligands. MHC-A, -B, and -C are present in all great apes with species-specific haplotypic variation in gene content. The Bw4 epitope recognized by lineage II KIR is restricted to MHC-B but also present on some gorilla and human MHC-A. Common to great apes, but rare in humans, are MHC-B possessing a C1 epitope recognized by lineage III KIR. MHC-C arose from duplication of MHC-B and is fixed in all great apes except orangutan, where it exists on approximately 50% of haplotypes and all allotypes are C1-bearing. Recent study showed that gorillas possess yet another intermediate MHC organization compared to humans. Like orangutans, but unlike the Pan-Homo species, duplication of MHC-B occurred. However, MHC-C is fixed, and the MHC-C C2 epitope (absent in orangutans) emerges. The evolution of MHC-C drove expansion of its cognate lineage III KIR. Recently, position −21 of the MHC-B leader sequence has been shown to be critical in determining NK cell educational outcome. In humans, methionine (−21M) results in CD94:NKG2A-focused education whereas threonine (−21T) produces KIR-focused education. This is another dynamic position among hominids. Orangutans have exclusively −21M, consistent with their intermediate stage in lineage III KIR-focused evolution. Gorillas have both −21M and −21T, like humans, but they are unequally encoded by their duplicated B genes. Chimpanzees have near-fixed −21T, indicative of KIR-focused NK education. Harmonious with this observation, chimpanzee KIR exhibit strong binding and, compared to humans, smaller differences between binding levels of activating and inhibitory KIR. Consistent between these MHC-NK cell receptor systems over the course of hominid evolution is the evolution of polymorphism favoring the more novel and dynamic KIR system

Estudio comparativo del splicing alternativo del gen NCR3 en diferentes especies de mamíferos y sus posibles implicaciones funcionales

Tesis doctoral inédita. Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 13-12-2013Alternative splicing (AS) is a major source of transcriptome and proteome diversity in higher eukaryotes allowing the generation of several structurally and functionally distinct mRNAs and protein isoforms from a single gene. Differential AS variants expression has been implicated in tissue and cellular differentiation, which in an evolutionary context could account for the phenotypic divergence of mammals that share a common repertoire of genes. On the other hand, the miss-regulation of AS is one of the mayor sources of human disease. However, there are not many detailed comparative analyses showing the alternative splice forms generated from particular genes among different organisms. The work presented here is a deep study of the splice variants expressed by the “Natural Cytotoxicity Receptor 3” (NCR3) gene and its comparative analysis among 13 mammals. NCR3 is a member of the NCR family, which represents the major NK cells triggering receptors. It has been involved in the recognition and killing of tumoral and infected cells, and in the maturation of dendritic cells. By using a combination of nested RT-PCR and RNA-seq analysis, it was possible to detect the expression of several NCR3 transcripts in all the analysed mammalian species, except in Mus musculus, where NCR3 is a pseudogen. It was observed an increase in the number of coding variants in primates, mainly by the internal splicing of exon 2 and the presence of exons 4II and 4III, which are only expressed in higher primates (Hominoidea). In contrast, the diversity of non-coding variants is similar among all analysed species, although there are some conserved ones, which could have a potential regulatory role. The nine human splice variants, six coding and three non-coding, were only detected at high expression levels in immune-related tissues, being the coding A, B y C the most abundant ones. The predicted human protein isoforms are potential transmembrane type I receptors, which extracellular domains are predicted to be Immunoglobulin type V (IgV) or type C (IgC). The interaction of these potential NCR3 extracellular domains with the ligand B7-H6 was analysed using flow citometry assays. Firstly, they were over-expressed in insect cells finding that defective glycosylation avoids binding. In a second approach, it was used a mammalian system detecting only specific binding of IgV domain to B7H6, indicating that B7H6 is not a ligand for IgC containing isoforms. In conclusion, all the presented data suggest a potential role of AS in the NCR3 functions and immune system regulation and evolution