Francis Roubinet

A Natural History of FUT2 Polymorphism in Humans

Because pathogens are powerful selective agents, host-cell surface molecules used by pathogens as identification signals can reveal the signature of selection. Most of them are oligosaccharides, synthesized by glycosyltransferases. One known example is balancing selection shaping ABO evolution as a consequence of both, A and B antigens being recognized as receptors by some pathogens, and anti-A and/or anti-B natural antibodies produced by hosts conferring protection against the numerous infectious agents expressing A and B motifs. These antigens can also be found in tissues other than blood if there is activity of another enzyme, FUT2, a fucosyltransferase responsible for ABO biosynthesis in body fluids. Homozygotes for null variants at this locus present the nonsecretor phenotype (se), because they cannot express ABO antigens in secretions. Multiple independent mutations have been shown to be responsible for the nonsecretor phenotype, which is coexisting with the secretor phenotype in most populations. In this study, we have resequenced the coding region of FUT2 in 732 individuals from 39 worldwide human populations. We report a complex pattern of natural selection acting on the gene. Although frequencies of secretor and nonsecretor phenotypes are similar in different populations, the point mutations at the base of the phenotypes are different, with some variants showing a long history of balancing selection among Eurasian and African populations, and one recent variant showing a fast spread in East Asia, likely due to positive selection. Thus, a convergent phenotype composition has been achieved through different mutations with different evolutionary histories.

FOXP3 mRNA levels are decreased in peripheral blood CD4+ lymphocytes from HIV-positive patients.

The impact of HIV infection on regulatory CD4+CD25high (Treg) lymphocyte subpopulations was evaluated by FOXP3 quantitative reverse transcriptase polymerase chain reaction and by flow cytometry. FOXP3 mRNA was quantified in peripheral blood mononuclear cells or purified CD4+ lymphocytes from HIV+ lymphopenic patients. Patients were distributed among clinical stages A, B, and C and received highly active antiretroviral therapy. The frequency of CD4+CD25high lymphocytes, measured by flow cytometry, was decreased in HIV+ patients (n = 38) compared with the group of uninfected subjects (n = 39). FOXP3 mRNA levels were found decreased in HIV+ patients (n = 25) compared with controls (n = 17) when expression of CD3γ or β-actin but not that of TATA box binding protein 1 was used for data normalization. Our results are compatible with a decrease of the Treg lymphocytes during HIV infection. The consequences of a Treg decrease are discussed in the context of immunologic anomalies observed during HIV infection.

Evolutionary dynamics of the human ABO gene

The ABO polymorphism has long been suspected to be under balancing selection. To explore this possibility, we analyzed two datasets: (1) a set of 94 23-Kb sequences in European- and African-Americans produced by the Seattle SNPs project, and (2) a set of 814 2-Kb sequences in O alleles from seven worldwide populations. A phylogenetic analysis of the Seattle sequences showed a complex pattern in which the action of recombination and gene conversion are evident, and in which four main lineages could be individuated. The sequence patterns could be linked to the expected blood group phenotype; in particular, the main mutation giving rise to the null O allele is likely to have appeared at least three times in human evolution, giving rise to allele lineages O02, O01, and O09. However, the genealogy changes along the gene and variations of both numbers of branches and of their time depth were observed, which could result from a combined action of recombination and selection. Several neutrality tests clearly demonstrated deviations compatible with balancing selection, peaking at several locations along the gene. The time depth of the genealogy was also incompatible with neutral evolution, particularly in the region from exons 6 to 7, which codes for most of the catalytic domain.

Patulin immunotoxicology: effect on phagocyte activation and the cellular and humoral immune system of mice and rabbits

Patulin is a mycotoxin frequently found in rotten apples or molded corn. We have investigated the effect of sublethal doses of patulin on the immune system in mice and rabbits. A significant suppression of the chemiluminescence response of peritoneal leucocytes was observed in both species. Mouse spleen lymphocytes showed a decrease in absolute number, most pronounced for the B-cell population whereas the Ts population showed a relative increase after patulin treatment. The mitogenic response to PHA, Con A and, in particular, PWM was also depressed by patulin. This was paralleled by decreasing serum immunoglobulin levels in the mice and rabbits. The immunosuppressive effect of patulin is reversible and is probably due to interaction with cellular free SH groups since the action of patulin can be circumvented, at least partially, by the prior administration of cystein. Under natural circumstances, patulin may constitute a health risk for animals.

Evolution of the O alleles of the human ABO blood group gene

BACKGROUND: To date, at least 40 different alleles O have been characterized on the basis of exon 6 and exon 7 sequences but not always for intron 6.

Molecular polymorphism of O alleles in five populations of different ethnic origins.

Sequences of exons 6 and 7 of the O allele of the ABO gene were studied in 317 individuals of the O phenotype from five different ethnic groups (Basques, Berbers, Akans from the Ivory Coast, and Amerindians: Cayapas from Ecuador and Aymaras from Bolivia). Twenty-one O alleles were characterized, among which 9 differed from all O alleles reported to date. The nine alleles differed from either the O01 allele (four out of nine) or O02 allele (five out of nine) by one to three point mutations. The number of different O alleles in population samples varied greatly: the highest number (13) was observed in Akans, and the lowest (5) in Amerindians. Some rare alleles previously reported by others at low frequencies were found with high frequencies in the Akans. The results also revealed a decreasing frequency of Ov7 alleles from south to north (Akans, Berbers, Basques). Berbers and Basques share two rare alleles, Ov6 and O03, which were not encountered in the other populations studied here.

A novel cis AB allele derived from a B allele through a single point mutation

BACKGROUND: The very rare cis AB phenotype, first described in 1964, corresponds to a special ABO allele encoding a glycosyltransferase that is capable of synthesizing both A and B substances. Until now, gene sequences of only two cis AB alleles were partially characterized. One involved the A1*02 allele with a single nonsynonymous substitution at codon 268, whereas the second arose from a single nonsynonymous substitution at codon 266 in exon 7 of a B1*01 allele.

A human monoclonal anti‐D antibody which detects a nonconformation‐dependent epitope on the RhD protein by immunoblot

We describe the first human monoclonal anti‐D (LOR‐15C9) which reacts with a D‐specific motif exposed either on a native form on intact D‐positive red cells or on a denatured form of the RhD protein (33u2003kD), and detected by immunoblotting. LOR‐15C9 was able to precipitate RhD but not RhcE proteins produced by in vitro transcription‐translation assays. The reactivity of the antibody, using panels of red cells with various partial D phenotypes known to lack some D epitopes and corresponding in RHD gene variants, suggested that LOR‐15C9 reactivity depends on the portion of the RhD polypeptide encoded by the exon 7 (amino acids 314–358). These findings correlate well with the reactivity of LOR‐15C9 with erythrocytes of some nonhuman primates (Dgor‐positive gorillas), but not of chimpanzee and Old or New World monkeys.

Variable region gene segment utilization in rhesus monkey hybridomas producing human red blood cell-specific antibodies: predominance of the VH4 family but not VH4-21 (V4-34).

Structural analyses of human immunoglobulin gene segments from monoclonal cell lines provide valuable information regarding the antibody repertoire. This information, in conjunction with a nearly complete knowledge of the human immunoglobulin germline repertoire, now allows further investigation into the underlying molecular basis responsible for some of the observed biases found in the expressed repertoire. One human heavy chain variable region gene segment, V4-34 (VH4-21), is one of the most prevalent gene segments in the expressed repertoire. The overwhelming presence of the V4-34 gene segment suggests that it may play an important role in immune responses. However, there is currently little information regarding its presence and potential importance in nonhuman primates. In order to determine if this gene segment is used by lower primates in a similar manner we determined the molecular structure of the variable region gene segments that are expressed by macaque monoclonal heterohybridomas that are specific for human red blood cell antigens. Eleven of the 12 hybridomas are derived from Rhesus monkeys (Macaca mulatta) and one is from a cynomologous monkey (Macaca fascicularis), all of which have been immunized with human red blood cells. The predominance of a VH4-like family and the specific absence of a VH4-21 equivalent led us to further characterize the macaque VH4 gene family at the germline level. Therefore, germline gene segments from the macaque equivalent to the human VH4 gene family are also described.

Erythrocyte‐magnetized technology: an original and innovative method for blood group serology

BACKGROUND: Erythrocyte‐magnetized technology (EMT) is a new fully automated method for ABO‐RH‐K phenotyping and antibody detection. The magnetization of red cells avoids centrifugation and washing phases. This report describes the results of an evaluation of this new technology on its specific automated system.