Ancient DNA and the tropics: a rodent's tale

Most genetic studies of Holocene fauna have been performed with ancient samples from dry and cold regions, in which preservation of fossils is facilitated and molecular damage is reduced. Ancient DNA work from tropical regions has been precluded owing to factors that limit DNA preservation (e.g. temperature, hydrolytic damage). We analysed ancient DNA from rodent jawbones identified as Ototylomys phyllotis, found in Holocene and Late Pleistocene stratigraphic layers from Loltún, a humid tropical cave located in the Yucatan peninsula. We extracted DNA and amplified six short overlapping fragments of the cytochrome b gene, totalling 666 bp, which represents an unprecedented success considering tropical ancient DNA samples. We performed genetic, phylogenetic and divergence time analyses, combining sequences from ancient and modern O. phyllotis, in order to assess the ancestry of the Loltún samples. Results show that all ancient samples fall into a unique clade that diverged prior to the divergence of the modern O. phyllotis, supporting it as a distinct Pleistocene form of the Ototylomys genus. Hence, this rodent''s tale suggests that the sister group to modern O. phyllotis arose during the Miocene–Pliocene, diversified during the Pleistocene and went extinct in the Holocene.     

Immunotherapy of murine leukemia following non-myeloablative conditioning with naïve or G-CSF mobilized blood or bone marrow stem cells

Allogeneic stem cell transplantation (SCT) is the treatment of choice for a large number of hematologic malignancies. Its major advantage over conventional chemotherapy lies in the graft-versus-leukemia (GVL) effects mediated by allo- or tumor-reactive donor lymphocytes given in the course of SCT or post transplantation as donor lymphocyte infusions (DLI). The benefits of cell-mediated immunotherapy over myeloablative radiochemotherapy have also made it possible to reduce the intensity of conditioning regimens. Mobilized peripheral blood has proved preferable to bone marrow (BM) as a source of stem cells for transplantation, since it provides a larger number of stem cells on the one hand and immunologically competent lymphocytes on the other. The use of granulocyte colony stimulating factor (G-CSF), which is necessary to mobilize and increase the number of stem cells, may down-regulate the GVL effect by suppression of donor effector T lymphocytes by inducing Th₁Th₂ cytokine switch. It has previously been shown that GVL effects may be amplified by both in vivo and in vitro activation of donor lymphocytes with human recombinant interleukin-2 (rIL-2). Our studies using a leukemic murine model prepared for transplantation with low intensity conditioning prior to infusion of G-CSF-mobilized peripheral blood stem cells (PBSC) have demonstrated that mobilization of blood cells with G-CSF and in vivo treatment with rIL-2 following low-intensity conditioning enhances the GVL effects and prolongs survival of recipients inoculated with BCL1. Activation of donor lymphocytes with rIL-2 may thus be useful for amplifying GVL effects following mobilization with G-CSF.     

The crystal structure of mouse phosphoglucose isomerase at 1.6A resolution and its complex with glucose 6-phosphate reveals the catalytic mechanism of sugar ring opening

Phosphoglucose isomerase (PGI) is an enzyme of glycolysis that interconverts glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P) but, outside the cell, is a multifunctional cytokine. High-resolution crystal structures of the enzyme from mouse have been determined in native form and in complex with the inhibitor erythrose 4-phosphate, and with the substrate glucose 6-phosphate. In the substrate-bound structure, the glucose sugar is observed in both straight-chain and ring forms. This structure supports a specific role for Lys518 in enzyme-catalyzed ring opening and we present a "push-pull" mechanism in which His388 breaks the O5-C1 bond by donating a proton to the ring oxygen atom and, simultaneously, Lys518 abstracts a proton from the C1 hydroxyl group. The reverse occurs in ring closure. The transition from ring form to straight-chain substrate is achieved through rotation of the C3-C4 bond, which brings the C1-C2 region into close proximity to Glu357, the base catalyst for the isomerization step. The structure with G6P also explains the specificity of PGI for glucose 6-phosphate over mannose 6-isomerase (M6P). To isomerize M6P to F6P requires a rotation of its C2-C3 bond but in PGI this is sterically blocked by Gln511.     

Identification of HLA-DRPheβ47 as the susceptibility marker of hypersensitivity to beryllium in individuals lacking the berylliosis-associated supratypic marker HLA-DPGluβ69

Background

Susceptibility to beryllium (Be)-hypersensitivity (BH) has been associated with HLA-DP alleles carrying a glutamate at position 69 of the HLA-DP β-chain (HLA-DPGlu69) and with several HLA-DP, -DQ and -DR alleles and polymorphisms. However, no genetic associations have been found between BH affected subjects not carrying the HLA-DPGlu69 susceptibility marker.

Methods

In this report, we re-evaluated an already described patient populations after 7 years of follow-up including new 29 identified BH subjects. An overall population 36 berylliosis patients and 38 Be-sensitization without lung granulomas and 86 Be-exposed controls was analysed to assess the role of the individual HLA-class II polymorphisms associated with BH-susceptibility in HLA-DPGlu69 negative subjects by univariate and multivariate analysis.

Results

As previously observed in this population the HLA-DPGlu69 markers was present in higher frequency in berylliosis patients (31 out of 36, 86%) than in Be-sensitized (21 out of 38, 55%, p = 0.008 vs berylliosis) and 41 out of 86 (48%, p < 0.0001 vs berylliosis, p = 0.55 vs Be-sensitized) Be-exposed controls.However, 22 subjects presenting BH did not carry the HLA-DPGlu69 marker. We thus evaluated the contribution of all the HLA-DR, -DP and -DQ polymorphisms in determining BH susceptibility in this subgroup of HLA-Glu69 subjects. In HLA-DPGlu69-negatives a significant association with BH was found for the HLA-DQLeu26, for the HLA-DRB1 locus residues Ser13, Tyr26, His32, Asn37, Phe47 and Arg74 and for the HLA-DRB3 locus clusterized residues Arg11, Tyr26, Asp28, Leu38, Ser60 and Arg74. HLA-DRPhe47 (OR 2.956, p < 0.05) resulting independently associated with BH. Further, Be-stimulated T-cell proliferation in the HLA-DPGlu69-negative subjects (all carrying HLA-DRPhe47) was inhibited by the anti-HLA-DR antibody (range 70–92% inhibition) significantly more than by the anti-HLA-DP antibody (range: 6–29%; p < 0.02 compared to anti-HLA-DR) while it was not affected by the anti-HLA-DQ antibody.

Conclusion

We conclude that HLA-DPGlu69 is the primary marker of Be-hypersensitivity and HLA-DRPhe47 is associated with BH in Glu69-negative subjects, likely playing a role in Be-presentation and sensitization.     

Rat MYH,a glycosylase for repair of oxidatively damaged DNA,has brain-specific isoforms that localize to neuronal mitochondria

Mitochondrial genomes are exposed to a heavy load of reactive oxygen species (ROS) that damage DNA. Since in neurons, mitochondrial DNA integrity must be maintained over the entire mammalian life span, neuronal mitochondria most likely repair oxidatively damaged DNA. We show that the Escherichia coli MutY DNA glycosylase homolog (MYH) in rat (rMYH) involved in repair of oxidative damage is abundantly expressed in the rat brain, with isoforms that are exclusive to brain tissue. Confocal microscopy and western analyses reveal localization of rMYH in neuronal mitochondria. To assess involvement of MYH in the neuronal response to oxidative DNA damage, we used a rat model of respiratory hypoxia, in which acutely reduced blood oxygenation leads to generation of superoxide, and formation and subsequent removal of 8-hydroxy-2'-deoxyguanosine (8OHdG). Removal of 8OHdG is accompanied by a spatial increase in rMYH immunoreactivity in the brain and an increase in levels of one of the three mitochondrial MYH isoforms, suggesting that inducible and non-inducible MYH isoforms exist in the brain. The mitochondrial localization of oxidative DNA damage repair enzymes in neurons may represent a specialized neuronal mechanism that safeguards mitochondrial genomes in the face of routine and accidental exposures to heavy loads of injurious ROS.     

Analysis of Genomic DNA from Medieval Plague Victims Suggests Long-Term Effect of Yersinia pestis on Human Immunity Genes

Pathogens and associated outbreaks of infectious disease exert selective pressure on human populations, and any changes in allele frequencies that result may be especially evident for genes involved in immunity. In this regard, the 1346-1353 Yersinia pestis-caused Black Death pandemic, with continued plague outbreaks spanning several hundred years, is one of the most devastating recorded in human history. To investigate the potential impact of Y. pestis on human immunity genes, we extracted DNA from 36 plague victims buried in a mass grave in Ellwangen, Germany in the 16th century. We targeted 488 immune-related genes, including HLA, using a novel in-solution hybridization capture approach. In comparison with 50 modern native inhabitants of Ellwangen, we find differences in allele frequencies for variants of the innate immunity proteins Ficolin-2 and NLRP14 at sites involved in determining specificity. We also observed that HLA-DRB1*13 is more than twice as frequent in the modern population, whereas HLA-B alleles encoding an isoleucine at position 80 (I-80+), HLA C*06:02 and HLA-DPB1 alleles encoding histidine at position 9 are half as frequent in the modern population. Simulations show that natural selection has likely driven these allele frequency changes. Thus, our data suggest that allele frequencies of HLA genes involved in innate and adaptive immunity responsible for extracellular and intracellular responses to pathogenic bacteria, such as Y. pestis, could have been affected by the historical epidemics that occurred in Europe.     

Agricultural development and maize diversity in Mexico

Mexico is within the center of origin of Zea mays and has among the highest levels of maize genetic diversity in the world. This diversity is traced to factors at the regional and farm levels. Loss of crop genetic diversity has been related to economic and agricultural development, although opposed views of this exist for the Mexican case. Agricultural development appears to be affecting virtually all types of farms in Mexico. A case study in Chiapas suggests that the adoption of some improved varieties has enhanced genetic diversity in maize, but one improved type competes with landraces in the most favorable land. The adoption of this improved type is associated with greater access to capital and with lower risk.The research for this project was carried out with the support of the UC/MEXUS program of the University of California and CONACYT. We wish to acknowledge the advice and assistance provided by Esteban Betanzos of the Chiapas office of the Instituto Nacional de Investigaciones Forestales y Agropecuarios, Efraim Hernandez of the Colegio de Postgraduados at Chapingo, Robert Tripp of the Centro Internacional de Mejoramiento de Maiz y Trigo, and to Manuel Parra of the Centro de Investigaciones Ecologicas del Sureste in San Cristobal de las Casas. The authors wish to thank J. Edward Taylor, Andraes Buerkert, Aaron Zazueta, and Daniel Mountjoy for their useful comments on an earlier draft.     

Genetic Evidence for Multiple Sources of the Non-Native Fish Cichlasoma urophthalmus (Günther; Mayan Cichlids) in Southern Florida

The number and diversity of source populations may influence the genetic diversity of newly introduced populations and affect the likelihood of their establishment and spread. We used the cytochrome b mitochondrial gene and nuclear microsatellite loci to identify the sources of a successful invader in southern Florida, USA, Cichlasoma urophthalmus (Mayan cichlid). Our cytochrome b data supported an introduction from Guatemala, while our microsatellite data suggested movement of Mayan Cichlids from the upper Yucatán Peninsula to Guatemala and introductions from Guatemala and Belize to Florida. The mismatch between mitochondrial and nuclear genomes suggests admixture of a female lineage from Guatemala, where all individuals were fixed for the mitochondrial haplotype found in the introduced population, and a more diverse but also relatively small number of individuals from Belize. The Florida cytochrome b haplotype appears to be absent from Belize (0 out of 136 fish screened from Belize had this haplotype). Genetic structure within the Florida population was minimal, indicating a panmictic population, while Mexican and Central American samples displayed more genetic subdivision. Individuals from the Upper Yucatán Peninsula and the Petén region of Guatemala were more genetically similar to each other than to fish from nearby sites and movement of Mayan Cichlids between these regions occurred thousands of generations ago, suggestive of pre-Columbian human transportation of Mayan Cichlids through this region. Mayan Cichlids present a rare example of cytonuclear disequilibrium and reduced genetic diversity in the introduced population that persists more than 30 years (at least 7–8 generations) after introduction. We suggest that hybridization occurred in ornamental fish farms in Florida and may contribute their establishment in the novel habitat. Hybridization prior to release may contribute to other successful invasions.