Sex chromosome systems in Neotropical Primates: What have we learnt so far from cytogenetics and genomics?

Neotropical Primates (Platyrrhini) show great diversity in their life histories, ecology, behaviour and genetics. This diversity extends to their chromosome complements, both to autosomes and to sex chromosomes. In this contribution, we will review what is currently known about sex chromosomes in this group, both from cytogenetic and from genomic evidence. The X and Y chromosomes in Neotropical Primates, also known as New World Monkeys, have striking structural differences compared with Old World Monkeys when Catarrhini sex chromosomes are considered. The XY bivalent displays a different meiotic behaviour in prophase I, and their Y chromosome shows extensive genomic differences. Even though the most widespread sex chromosome system is the XX/XY and thus considered the ancestral one for Platyrrhini, modifications of this sexual system are observed within this group. Multiple sex chromosome systems originated from Y-autosome translocations were described in several genera (Aotus, Callimico and Alouatta). In the howler monkeys, genus Alouatta, an independent origin of the sexual systems in South American and Mesoamerican species was postulated. All the above-mentioned evidence suggests that the Y chromosome of Platyrrhini has a different evolutionary history compared with the Catarrhini Y. There is still much to understand regarding their sex chromosome systems.

IgG subclasses in New World Monkeys: an issue for debate?

Immunoglobulin G (IgG) is an essential antibody in adaptive immunity; a differential expansion of the gene encoding the Fc region (IGHG) of this antibody has been observed in mammals. Like humans, animal biomedical models, such as mice and macaques, have four functional genes encoding 4 IgG subclasses; however, the data for New World monkeys (NWM) seems contentious. Some publications argue for the existence of a single-copy gene for IgG Fc; however, a recent paper has suggested the presence of IgG subclasses in some NWM species. Here, we evaluated the genetic distances and phylogenetic relationships in NWM to assess the presence of IgG subclasses using the sequences of IGHG genes from 13 NWM species recovered from genomic data and lab PCR and cloning-based procedures available in GenBank. The results show that several sequences do not cluster into the expected taxon, probably due to cross-contamination during laboratory procedures, and consequently, they appear to be wrongly assigned. Additionally, several sequences reported as subclasses were shown to be 100% identical in the CH domains. The data presented here suggests that there is not enough evidence to establish the presence of IgG subclasses in NWM.

Endocranial asymmetry in New World monkeys: a comparative phylogenetic analysis of morphometric data.

Brain lateralization is a widespread phenomenon although its expression across primates is still controversial due to the reduced number of species analyzed and the disparity of methods used. To gain insight into the diversification of neuroanatomical asymmetries in non-human primates we analyze the endocasts, as a proxy of external brain morphology, of a large sample of New World monkeys and test the effect of brain size, home range and group sizes in the pattern and magnitude of shape asymmetry. Digital endocasts from 26 species were obtained from MicroCT scans and a set of 3D coordinates was digitized on endocast surfaces. Results indicate that Ateles, Brachyteles, Callicebus and Cacajao tend to have a rightward frontal and a leftward occipital lobe asymmetry, whereas Aotus, Callitrichinae and Cebinae have either the opposite pattern or no directional asymmetry. Such differences in the pattern of asymmetry were associated with group and home range sizes. Conversely, its magnitude was significantly associated with brain size, with larger-brained species showing higher inter-hemispheric differences. These findings support the hypothesis that reduction in inter-hemispheric connectivity in larger brains favors the lateralization and increases the structural asymmetries, whereas the patterns of shape asymmetry might be driven by socio-ecological differences among species.

A New World Monkey Resembles Human in Bitter Taste Receptor Evolution and Function via a Single Parallel Amino Acid Substitution.

Bitter taste receptors serve as a vital component in the defense system against toxin intake by animals, and the family of genes encoding these receptors has been demonstrated, usually by family size variance, to correlate with dietary preference. However, few systematic studies of specific Tas2R to unveil their functional evolution have been conducted. Here, we surveyed Tas2R16 across all major clades of primates and reported a rare case of a convergent change to increase sensitivity to ß-glucopyranosides in human and a New World monkey, the white-faced saki. Combining analyses at multiple levels, we demonstrate that a parallel amino acid substitution (K172N) shared by these two species is responsible for this functional convergence of Tas2R16. Considering the specialized feeding preference of the white-faced saki, the K172N change likely played an important adaptive role in its early evolution to avoid potentially toxic cyanogenic glycosides, as suggested for the human TAS2R16 gene.

Molecular characterisation of parvorder Platyrrhini IgG sub-classes.

Non-human primates (NHP) are essential in modern biomedical research; New World monkeys (NWM) are mainly used as an experimental model regarding human malaria as they provide useful information about the parasite's biology and an induced immune response. It is known that a vaccine candidate's efficacy is mediated by a protection-inducing antibody response (IgG). Not enough information is available concerning IgG subclasses' molecular characteristics regarding NHP from parvorder Platyrrhini. Understanding the nature of the humoral immune response and characterising the IgG subclasses' profile will provide valuable information about the immunomodulator mechanisms of vaccines evaluated using an NHP animal model. This article has characterised IgG subclasses in NWM (i.e. genera Aotus, Cebus, Ateles and Alouatta) based on the amplification, cloning and sequencing of the immunoglobulin heavy constant gamma (IGHG) gene's CH1 to CH3 regions. The resulting sequences enabled elucidating IGHG gene organisation; two IgG variants were found in the Aotus and Ateles monkey group and three IgG variants in the Cebus and Alouatta group. The sequences were highly conserved in Platyrrhini and had a similar structure to that reported for monkeys from parvorder Catarrhini. Such information will help in developing tools for a detailed characterisation of the humoral immune response in an NWM experimental animal model.

Evolution of the guanylate binding protein (GBP) genes: Emergence of GBP7 genes in primates and further acquisition of a unique GBP3 gene in simians.

Guanylate binding proteins (GBPs) are major players in the host immunity, providing defense against bacterial and viral invaders. Multigene families may suffer different processes of evolution. Gene families related to the immune system usually follow the birth-and-death evolution process, where duplicated genes can be deleted, gain new functions or become non-functional. We analyzed publicly available primate GBP sequences and their genomic organization and observed that GBP7 genes appear to have emerged from a duplication of GBP4 and seem to be only present in primates. Furthermore, GBP3 genes are only present in Simiiformes and probably originated from GBP1 genes. Finally, a duplication event occurred in the GBP6 in Tarsiiformes and became functional which might also explain the duplication of GBP6 in New World monkeys and Cercopithecidae. Taken together, this study provides new knowledge on the evolution of GBPs in primates and suggests that a revision of the GBPs nomenclature is necessary.

Nomenclature report 2019: major histocompatibility complex genes and alleles of Great and Small Ape and Old and New World monkey species.

The major histocompatibility complex (MHC) is central to the innate and adaptive immune responses of jawed vertebrates. Characteristic of the MHC are high gene density, gene copy number variation, and allelic polymorphism. Because apes and monkeys are the closest living relatives of humans, the MHCs of these non-human primates (NHP) are studied in depth in the context of evolution, biomedicine, and conservation biology. The Immuno Polymorphism Database (IPD)-MHC NHP Database (IPD-MHC NHP), which curates MHC data of great and small apes, as well as Old and New World monkeys, has been upgraded. The curators of the database are responsible for providing official designations for newly discovered alleles. This nomenclature report updates the 2012 report, and summarizes important nomenclature issues and relevant novel features of the IPD-MHC NHP Database.

The CENP-B box, a nucleotide motif involved in centromere formation, has multiple origins in New World monkeys.

Centromere protein B (CENP-B), a protein participating in centromere formation, binds to centromere satellite DNA by recognizing a 17-bp motif called the CENP-B box. This motif is found in hominids (humans and great apes) at an identical location in repeat units of their centromere satellite DNA. We have recently reported that the CENP-B box exists at diverse locations in three New World monkey species (marmoset, squirrel monkey and tamarin). However, the evolutionary origin of the CENP-B box in these species was not determined. It could have been present in a common ancestor, or emerged multiple times in different lineages. Here we present results of a phylogenetic analysis of centromere satellite DNA that support the multiple emergence hypothesis. Repeat units almost invariably formed monophyletic groups in each species and the CENP-B box location was unique for each species. The CENP-B box is not essential for the immediate survival of its host organism. On the other hand, it is known to be required for de novo centromere assembly. Our results suggest that the CENP-B box confers a long-term selective advantage. For example, it may play a pivotal role when a centromere is accidentally lost or impaired.

Igh locus structure and evolution in Platyrrhines: new insights from a genomic perspective.

Non-human primates have been used as animal models because of their phylogenetic closeness to humans. However, the genetic differences between humans and non-human primates must be considered to select the appropriate animal models. Recently, New World monkeys (Platyrrhines) have generated a higher interest in biomedical research, especially in assessing vaccine safety and immunogenicity. Given the continued and renewed interest in Platyrrhines as biomedical models, it is a necessary to have a better and more complete understanding of their immune system and its implications for research. Immunoglobulins (Ig) are the main proteins that mediate humoral immunity. These proteins have evolved as part of an adaptive immune response system derived from ancient vertebrates. There are at least four Ig classes in Prosimians, whereas five have been reported in Catarrhines. Information on the structure and evolution of the loci containing immunoglobulin heavy chain constant genes (Igh) in Platyrrhines, however, is limited. Here, Igh loci were characterized in 10 Platyrrhines using the available whole genome sequences. Human and Macaca Igh loci were also assessed to compare them with their Platyrrhines counterparts. Differences in Igh locus structure were observed between Platyrrhines and Catarrhines. Noteworthy changes occur in the γ gene, which encodes a key Ig involved in organism defense that would favor protection after vaccination. The remarkable differences between the immunoglobulin proteins of Platyrrhines and Catarrhines warrant a cautionary message to biomedical researchers.

Cooperative evolution of two different TEs results in lineage-specific novel transcripts in the BLOC1S2 gene.

BACKGROUND: The BLOC1S2 gene encodes the multifunctional protein BLOS2, a shared subunit of two lysosomal trafficking complexes: i) biogenesis of lysosome-related organelles complex-1 and i) BLOC-1-related complex. In our previous study, we identified an intriguing unreported transcript of the BLOC1S2 gene that has a novel exon derived from two transposable elements (TEs), MIR and AluSp. To investigate the evolutionary footprint and molecular mechanism of action of this transcript, we performed PCR and RT-PCR experiments and sequencing analyses using genomic DNA and RNA samples from humans and various non-human primates. RESULTS: The results showed that the MIR element had integrated into the genome of our common ancestor, specifically in the BLOC1S2 gene region, before the radiation of all primate lineages and that the AluSp element had integrated into the genome of our common ancestor, fortunately in the middle of the MIR sequences, after the divergence of Old World monkeys and New World monkeys. The combined MIR and AluSp sequences provide a 3' splice site (AG) and 5' splice site (GT), respectively, and generate the Old World monkey-specific transcripts. Moreover, branch point sequences for the intron removal process are provided by the MIR and AluSp combination. CONCLUSIONS: We show for the first time that sequential integration into the same location and sequence divergence events of two different TEs generated lineage-specific transcripts through sequence collaboration during primate evolution.

Conservation of cell-intrinsic immune responses in diverse nonhuman primate species.

Differences in immune responses across species can contribute to the varying permissivity of species to the same viral pathogen. Understanding how our closest evolutionary relatives, nonhuman primates (NHPs), confront pathogens and how these responses have evolved over time could shed light on host range barriers, especially for zoonotic infections. Here, we analyzed cell-intrinsic immunity of primary cells from the broadest panel of NHP species interrogated to date, including humans, great apes, and Old and New World monkeys. Our analysis of their transcriptomes after poly(I:C) transfection revealed conservation in the functional consequences of their response. In mapping reads to either the human or the species-specific genomes, we observed that with the current state of NHP annotations, the percent of reads assigned to a genetic feature was largely similar regardless of the method. Together, these data provide a baseline for the cell-intrinsic responses elicited by a potent immune stimulus across multiple NHP donors, including endangered species, and serve as a resource for refining and furthering the existing annotations of NHP genomes.

AVPR1b variation and the emergence of adaptive phenotypes in Platyrrhini primates.

Platyrrhini (New World monkeys, NWm) are a group of primates characterized by behavioral and reproductive traits that are otherwise uncommon among primates, including social monogamy, direct paternal care, and twin births. As a consequence, the study of Platyrrhine primates is an invaluable tool for the discovery of the genetic repertoire underlying these taxon-specific traits. Recently, high conservation of vasopressin (AVP) sequence, in contrast with high variability of oxytocin (OXT), has been described in NWm. AVP and OXT functions are possible due to interaction with their receptors: AVPR1a, AVPR1b, AVPR2, and OXTR; and the variability in this system is associated with the traits mentioned above. Understanding the variability in the receptors is thus fundamental to understand the function and evolution of the system as a whole. Here we describe the variability of AVPR1b coding region in 20 NWm species, which is well-known to influence behavioral traits such as aggression, anxiety, and stress control in placental mammals. Our results indicate that 4% of AVPR1b sites may be under positive selection and a significant number of sites under relaxed selective constraint. Considering the known role of AVPR1b, we suggest that some of the changes described here for the Platyrrhini may be a part of the genetic repertoire connected with the complex network of neuroendocrine mechanisms of AVP-OXT system in the modulation of the HPA axis. Thus, these changes may have promoted the emergence of social behaviors such as direct paternal care in socially monogamous species that are also characterized by small body size and twin births.

Multiple factors behind early diversification of skull morphology in the continental radiation of New World monkeys.

Understanding the origin of diversity is a fundamental problem in evolutionary biology. The null expectation for the evolutionary diversification is that all changes in biological diversity are the result of random processes. Adaptive radiations depart from this expectation as ecological factors and natural selection are supposed to play a central role in driving exceptional diversification. However, it is not well understood how large-scale continental radiations, given their characteristics, fit to these opposing theoretical models. Here, we used phylogenetic comparative methods and geometric morphometrics to study the evolutionary process of cranial diversification in the continental radiation of New World monkeys. Particularly, we tested several alternative evolutionary scenarios for morphological evolution in the clade. Results indicated that despite the platyrrhine radiation being old and geographically widespread, the formative patterns arising from the initial stages of diversification probably associated with an adaptive radiation can still be recognized today. We also show that no single explored factor (e.g., ecological or allometric) can be invoked as a complete explanation for the observed phenotypic diversity patterns in the clade and, moreover, that different cranial regions exhibit particular macroevolutionary patterns. Together, our results highlight the evident complexity behind large-scale evolutionary radiations.

A RAD-sequencing approach to genome-wide marker discovery, genotyping, and phylogenetic inference in a diverse radiation of primates.

Until recently, most phylogenetic and population genetics studies of nonhuman primates have relied on mitochondrial DNA and/or a small number of nuclear DNA markers, which can limit our understanding of primate evolutionary and population history. Here, we describe a cost-effective reduced representation method (ddRAD-seq) for identifying and genotyping large numbers of SNP loci for taxa from across the New World monkeys, a diverse radiation of primates that shared a common ancestor ~20-26 mya. We also estimate, for the first time, the phylogenetic relationships among 15 of the 22 currently-recognized genera of New World monkeys using ddRAD-seq SNP data using both maximum likelihood and quartet-based coalescent methods. Our phylogenetic analyses robustly reconstructed three monophyletic clades corresponding to the three families of extant platyrrhines (Atelidae, Pitheciidae and Cebidae), with Pitheciidae as basal within the radiation. At the genus level, our results conformed well with previous phylogenetic studies and provide additional information relevant to the problematic position of the owl monkey (Aotus) within the family Cebidae, suggesting a need for further exploration of incomplete lineage sorting and other explanations for phylogenetic discordance, including introgression. Our study additionally provides one of the first applications of next-generation sequencing methods to the inference of phylogenetic history across an old, diverse radiation of mammals and highlights the broad promise and utility of ddRAD-seq data for molecular primatology.

Serotonin, behavior, and natural selection in New World monkeys.

Traits that undergo massive natural selection pressure, with multiple events of positive selection, are hard to find. Social behaviour, in social animals, is crucial for survival, and genetic networks involved in behaviour, such as those of serotonin (5-HT) and other neurotransmitters, must be the target of natural selection. Here, we used molecular analyses to search for signals of positive selection in the 5-HT system and found such signals in the M3-M4 intracellular domain of the 5-HT3A serotonin receptor subunit (HTR3A) in primates. We detected four amino acid sites with signs of putatively positive selection (398, 403, 432 and 416); the first three showed indications of being selected in New World monkeys (NWM, Platyrrhini), specifically in the Callitrichinae branch. Additionally, we searched for associations of these amino acid variants with social behavioural traits (i.e. sex-biased dispersal, dominance and social monogamy) using classical and Bayesian methods, and found statistically significant associations for unbiased sex dispersal (398L and 416S), unbiased sex dominance (416S) and social monogamy (416S), as well as significant positive correlation between female dispersal and 403G. Furthermore, we found putatively functional protein motifs determined by three selected sites, of which we highlight a ligand motif to GSK3 in the 416S variant, appearing only in Platyrrhini. 5-HT, 5-HT3A receptor and GSK3 are part of a network that participates in neurodevelopment and regulates behaviour, among other functions. We suggest that these genetic variations, together with those found in other neurotransmitter systems, must contribute to adaptive behaviours and consequently to fitness in NWMs.

Contrasting patterns of RUNX2 repeat variations are associated with palate shape in phyllostomid bats and New World primates.

Establishing the genetic basis that underlies craniofacial variability in natural populations is one of the main topics of evolutionary and developmental studies. One of the genes associated with mammal craniofacial variability is RUNX2, and in the present study we investigated the association between craniofacial length and width and RUNX2 across New World bats (Phyllostomidae) and primates (Catarrhini and Platyrrhini). Our results showed contrasting patterns of association between the glutamate/alanine ratios (Q/A ratio) and palate shape in these highly diverse groups. In phyllostomid bats, we found an association between shorter/broader faces and increase of the Q/A ratio. In New World monkeys (NWM) there was a positive correlation of increasing Q/A ratios to more elongated faces. Our findings reinforced the role of the Q/A ratio as a flexible genetic mechanism that would rapidly change the time of skull ossification throughout development. However, we propose a scenario in which the influence of this genetic adjustment system is indirect. The Q/A ratio would not lead to a specific phenotype, but throughout the history of a lineage, would act along with evolutionary constraints, as well as other genes, as a facilitator for adaptive morphological changes.

Nonhuman primate models of human viral infections.

Humans have a close phylogenetic relationship with nonhuman primates (NHPs) and share many physiological parallels, such as highly similar immune systems, with them. Importantly, NHPs can be infected with many human or related simian viruses. In many cases, viruses replicate in the same cell types as in humans, and infections are often associated with the same pathologies. In addition, many reagents that are used to study the human immune response cross-react with NHP molecules. As such, NHPs are often used as models to study viral vaccine efficacy and antiviral therapeutic safety and efficacy and to understand aspects of viral pathogenesis. With several emerging viral infections becoming epidemic, NHPs are proving to be a very beneficial benchmark for investigating human viral infections.

Oxytocin structure and function in New World monkeys: from pharmacology to behavior.

Oxytocin (OT) is a hypothalamic nonapeptide that mediates a host of physiological and behavioral processes including reproductive physiology and social attachments. While the OT sequence structure is highly conserved among mammals, New World monkeys (NWMs) represent an unusual "hot spot" in OT structure variability among mammals. At least 6 distinct OT ligand variants among NWMs exist, yet it is currently unclear whether these evolved structural changes result in meaningful functional consequences. NWMs offer a new area to explore how these modifications to OT and its canonical G-protein coupled OT receptor (OTR) may mediate specific cellular, physiological and behavioral outcomes. In this review, we highlight relationships between OT ligand and OTR structural variability, specifically examining coevolution between OT ligands, OTRs, and physiological and behavioral phenotypes across NWMs. We consider whether these evolved modifications to the OT structure alter pharmacological profiles at human and marmoset OTRs, including changes to receptor binding, intracellular signaling and receptor internalization. Finally, we evaluate whether exogenous manipulation using OT variants in marmoset monkeys differentially enhance or impair behavioral processes involved in social relationships between pairmates, opposite-sex strangers, and parents and their offspring. Overall, it appears that changes to OT ligands in NWMs result in important changes ranging from cellular signaling to broad measures of social behavior.

An eutherian intronic sequence gave rise to a major satellite DNA in Platyrrhini.

Satellite DNAs (satDNAs) are major components of eukaryote genomes. However, because of their quick divergence, the evolutionary origin of a given satDNA family can rarely be determined. Herein we took advantage of available primate sequenced genomes to determine the origin of the CapA satDNA (approx. 1500 bp long monomers), first described in the tufted capuchin monkey Sapajus apella We show that CapA is an abundant satDNA in Platyrrhini, whereas in the genomes of most eutherian mammals, including humans, this sequence is present only as a single copy located within a large intron of the NOS1AP (nitric oxide synthase 1 adaptor protein) gene. Our data suggest that this intronic CapA-like sequence gave rise to the CapA satDNA and we discuss possible mechanisms implicated in this event. This is the first report to our knowledge of a single copy intronic sequence giving origin to a satDNA that reaches up to 100 000 copies in some genomes.

Ancestral resurrection of anthropoid estrogen receptor ß demonstrates functional consequences of positive selection.

Anthropoid primates arose during the Eocene approximately 55 million years ago (mya), and extant anthropoids share a most recent common ancestor ∼40mya. Paleontology has been very successful at describing the morphological phenotypes of extinct anthropoids. Less well understood is the molecular biology of these extinct species as well as the phenotypic consequences of evolutionary variation in their genomes. Here we resurrect the most recent common ancestral anthropoid estrogen receptor ß gene (ESR2) and demonstrate that the function of this ancestral estrogen receptor has been maintained during human descent but was altered during early New World monkey (NWM) evolution by becoming a more potent transcriptional activator. We tested hypotheses of adaptive evolution in the protein coding sequences of ESR2, and determined that ESR2 evolved via episodic positive selection on the NWM stem lineage. We separately co-transfected ESR2 constructs for human, NWM, and the anthropoid ancestor along with reporter gene vectors and performed hormone binding dose response experiments that measure transactivation activity. We found the transactivation potentials of the ancestral and human sequences to be significantly lower (p<0.0001 in each comparison) than that of the NWM when treated with estradiol, the most prevalent estrogen. We conclude the difference in fold activation is due to positive selection in the NWM ERß ligand binding domain. Our study validates inferential methods for detecting adaptive evolution that predict functional consequences of nucleotide substitutions and points a way toward examining the functional consequences of positive Darwinian selection.