Pascal Gagneux

Human uptake and incorporation of an immunogenic nonhuman dietary sialic acid

Humans are genetically unable to produce the sialic acid N-glycolylneuraminic acid (Neu5Gc), because of a mutation that occurred after our last common ancestor with great apes. Although Neu5Gc is presumed absent from normal humans, small amounts have been claimed to exist in human tumors and fetal meconium. We have generated an antibody with high specificity and avidity for Neu5Gc. Fetal tissues, normal adult tissues, and breast carcinomas from humans showed reactivity to this antibody, primarily within secretory epithelia and blood vessels. The presence of small amounts of Neu5Gc was confirmed by MS. Absent any known alternate pathway for its synthesis, we reasoned that these small amounts of Neu5Gc might originate from exogenous sources. Indeed, human cells fed with Neu5Gc incorporated it into endogenous glycoproteins. When normal human volunteers ingested Neu5Gc, a portion was absorbed and eliminated in urine, and small quantities were incorporated into newly synthesized glycoproteins. Neu5Gc has never been reported in plants or microbes to our knowledge. We found that Neu5Gc is rare in poultry and fish, common in milk products, and enriched in red meats. Furthermore, normal humans have variable amounts of circulating IgA, IgM, and IgG antibodies against Neu5Gc, with the highest levels comparable to those of the previously known anti-α-galactose xenoreactive antibodies. This finding represents an instance wherein humans absorb and metabolically incorporate a nonhuman dietary component enriched in foods of mammalian origin, even while generating xenoreactive, and potentially autoreactive, antibodies against the same molecule. Potential implications for human diseases are briefly discussed.

Great ape genetic diversity and population history

Most great ape genetic variation remains uncharacterized; however, its study is critical for understanding population history, recombination, selection and susceptibility to disease. Here we sequence to high coverage a total of 79 wild- and captive-born individuals representing all six great ape species and seven subspecies and report 88.8 million single nucleotide polymorphisms. Our analysis provides support for genetically distinct populations within each species, signals of gene flow, and the split of common chimpanzees into two distinct groups: Nigeria–Cameroon/western and central/eastern populations. We find extensive inbreeding in almost all wild populations, with eastern gorillas being the most extreme. Inferred effective population sizes have varied radically over time in different lineages and this appears to have a profound effect on the genetic diversity at, or close to, genes in almost all species. We discover and assign 1,982 loss-of-function variants throughout the human and great ape lineages, determining that the rate of gene loss has not been different in the human branch compared to other internal branches in the great ape phylogeny. This comprehensive catalogue of great ape genome diversity provides a framework for understanding evolution and a resource for more effective management of wild and captive great ape populations.

Microsatellite scoring errors associated with noninvasive genotyping based on nuclear DNA amplified from shed hair

In the context of a study of wild chimpanzees, Pan troglodytes verus, we found that genotypes based on single PCR amplifications of microsatellite loci from single shed hair have a high error rate. We quantified error rates using the comparable results of 791 single shed hair PCR amplifications of 11 microsatellite loci of 18 known individuals. The most frequent error was the amplification of only one of the two alleles present at a heterozygous locus. This phenomenon, called allelic dropout, produced false homozygotes in 31% of single‐hair amplifications. There was no difference in the probability of preferential amplification between longer and shorter alleles. The probability of scoring false homozygotes can be reduced to below 0.05 by three separate amplifications from single hairs of the same individual or by pooling hair samples from the same individual. In this study an additional 5.6% of the amplifications gave wrong genotypes because of contamination, labelling and loading errors, and possibly amplification artefacts. In contrast, amplifications from plucked hair taken from four dead individuals gave consistent results (error rate < 0.01%, n= 120). Allelic dropout becomes a problem when the DNA concentration falls below 0.05 ng/10 μL in the template as it can with shed hair, and extracts from faeces and masticated plant matter.

Multifarious roles of sialic acids in immunity

Sialic acids are a diverse family of monosaccharides widely expressed on all cell surfaces of vertebrates and so‐called “higher” invertebrates, and on certain bacteria that interact with vertebrates. This overview surveys examples of biological roles of sialic acids in immunity, with emphasis on an evolutionary perspective. Given the breadth of the subject, the treatment of individual topics is brief. Subjects discussed include biophysical effects regulation of factor H; modulation of leukocyte trafficking via selectins; Siglecs in immune cell activation; sialic acids as ligands for microbes; impact of microbial and endogenous sialidases on immune cell responses; pathogen molecular mimicry of host sialic acids; Siglec recognition of sialylated pathogens; bacteriophage recognition of microbial sialic acids; polysialic acid modulation of immune cells; sialic acids as pathogen decoys or biological masks; modulation of immunity by sialic acid O‐acetylation; sialic acids as antigens and xeno‐autoantigens; antisialoglycan antibodies in reproductive incompatibility; and sialic‐acid–based blood groups.

Loss of Siglec expression on T lymphocytes during human evolution

We report here that human T cells give much stronger proliferative responses to specific activation via the T cell receptor (TCR) than those from chimpanzees, our closest evolutionary relatives. Nonspecific activation using phytohemagglutinin was robust in chimpanzee T cells, indicating that the much lower response to TCR simulation is not due to any intrinsic inability to respond to an activating stimulus. CD33-related Siglecs are inhibitory signaling molecules expressed on most immune cells and are thought to down-regulate cellular activation pathways via cytosolic immunoreceptor tyrosine-based inhibitory motifs. Among human immune cells, T lymphocytes are a striking exception, expressing little to none of these molecules. In stark contrast, we find that T lymphocytes from chimpanzees as well as the other closely related “great apes” (bonobos, gorillas, and orangutans) express several CD33-related Siglecs on their surfaces. Thus, human-specific loss of T cell Siglec expression occurred after our last common ancestor with great apes, potentially resulting in an evolutionary difference with regard to inhibitory signaling. We confirmed this by studying Siglec-5, which is prominently expressed on chimpanzee lymphocytes, including CD4 T cells. Ab-mediated clearance of Siglec-5 from chimpanzee T cells enhanced TCR-mediated activation. Conversely, primary human T cells and Jurkat cells transfected with Siglec-5 become less responsive; i.e., they behave more like chimpanzee T cells. This human-specific loss of T cell Siglec expression associated with T cell hyperactivity may help explain the strikingly disparate prevalence and severity of T cell-mediated diseases such as AIDS and chronic active hepatitis between humans and chimpanzees.

Evidence for an ancient selective sweep in the MHC class I gene repertoire of chimpanzees

MHC class I molecules play an essential role in the immune defense against intracellular infections. The hallmark of the MHC is its extensive degree of polymorphism at the population level. However, the present comparison of MHC class I gene intron variation revealed that chimpanzees have experienced a severe repertoire reduction at the orthologues of the HLA-A, -B, and -C loci. The loss of variability predates the (sub)speciation of chimpanzees and did not effect other known gene systems. Therefore the selective sweep in the MHC class I gene may have resulted from a widespread viral infection. Based on the present results and the fact that chimpanzees have a natural resistance to the development of AIDS, we hypothesize that the selective sweep was caused by the chimpanzee-derived simian immunodeficiency virus (SIVcpz), the closest relative of HIV-1, or a closely related retrovirus. Hence, the contemporary chimpanzee populations represent the offspring of AIDS-resistant animals, the survivors of a HIV-like pandemic that took place in the distant past.

Alu-mediated inactivation of the human CMP- N-acetylneuraminic acid hydroxylase gene

Inactivation of the CMP-N-acetylneuraminic acid hydroxylase gene has provided an example of human-specific genomic mutation that results in a widespread biochemical difference between human and nonhuman primates. We have found that, although a region containing a 92-bp exon and an AluSq element in the hydroxylase gene is intact in all nonhuman primates examined, the same region in the human genome is replaced by an AluY element that was disseminated at least one million years ago. We propose a mechanistic model for this Alu-mediated replacement event, which deleted the 92-bp exon and thus inactivated the human hydroxylase gene. It is suggested that Alu elements have played potentially important roles in genotypic and phenotypic evolution in the hominid lineage.

Evolution of genetic and genomic features unique to the human lineage

Given the unprecedented tools that are now available for rapidly comparing genomes, the identification and study of genetic and genomic changes that are unique to our species have accelerated, and we are entering a golden age of human evolutionary genomics. Here we provide an overview of these efforts, highlighting important recent discoveries, examples of the different types of human-specific genomic and genetic changes identified, and salient trends, such as the localization of evolutionary adaptive changes to complex loci that are highly enriched for disease associations. Finally, we discuss the remaining challenges, such as the incomplete nature of current genome sequence assemblies and difficulties in linking human-specific genomic changes to human-specific phenotypic traits.

Innovations in host and microbial sialic acid biosynthesis revealed by phylogenomic prediction of nonulosonic acid structure

Sialic acids (Sias) are nonulosonic acid (NulO) sugars prominently displayed on vertebrate cells and occasionally mimicked by bacterial pathogens using homologous biosynthetic pathways. It has been suggested that Sias were an animal innovation and later emerged in pathogens by convergent evolution or horizontal gene transfer. To better illuminate the evolutionary processes underlying the phenomenon of Sia molecular mimicry, we performed phylogenomic analyses of biosynthetic pathways for Sias and related higher sugars derived from 5,7-diamino-3,5,7,9-tetradeoxynon-2-ulosonic acids. Examination of ≈1,000 sequenced microbial genomes indicated that such biosynthetic pathways are far more widely distributed than previously realized. Phylogenetic analysis, validated by targeted biochemistry, was used to predict NulO types (i.e., neuraminic, legionaminic, or pseudaminic acids) expressed by various organisms. This approach uncovered previously unreported occurrences of Sia pathways in pathogenic and symbiotic bacteria and identified at least one instance in which a human archaeal symbiont tentatively reported to express Sias in fact expressed the related pseudaminic acid structure. Evaluation of targeted phylogenies and protein domain organization revealed that the “unique” Sia biosynthetic pathway of animals was instead a much more ancient innovation. Pathway phylogenies suggest that bacterial pathogens may have acquired Sia expression via adaptation of pathways for legionaminic acid biosynthesis, one of at least 3 evolutionary paths for de novo Sia synthesis. Together, these data indicate that some of the long-standing paradigms in Sia biology should be reconsidered in a wider evolutionary context of the extended family of NulO sugars.

Evolutionary glycomics: characterization of milk oligosaccharides in primates.

Free oligosaccharides are abundant components of mammalian milk and have primary roles as prebiotic compounds, in immune defense, and in brain development. A mass spectrometry-based technique is applied to profile milk oligosaccharides from apes (chimpanzee, gorilla, and siamang), new world monkeys (golden lion tamarin and common marmoset), and an old world monkey (rhesus). The purpose of this study is to evaluate the patterns of primate milk oligosaccharide composition from a phylogenetic perspective to assess the extent to which the compositions of HMOs derives from ancestral primate patterns as opposed to more recent evolutionary events. Milk oligosaccharides were quantitated by nanoflow liquid chromatography on chip-based devices. The relative abundances of fucosylated and sialylated milk oligosaccharides in primates were also determined. For a systematic and comprehensive study of evolutionary patterns of milk oligosaccharides, cluster analysis of primate milk was performed using the chromatographic profile. In general, the oligosaccharides in primate milk, including humans, are more complex and exhibit greater diversity compared to the ones in nonprimate milk. A detailed comparison of the oligosaccharides across evolution revealed nonsequential developmental pattern, that is, that primate milk oligosaccharides do not necessarily cluster according to the primate phylogeny. This report represents the first comprehensive and quantitative effort to profile and elucidate the structures of free milk oligosaccharides so that they can be related to glycan function in different primates.