The Expanded mtDNA Phylogeny of the Franco-Cantabrian Region Upholds the Pre-Neolithic Genetic Substrate of Basques

The European genetic landscape has been shaped by several human migrations occurred since Paleolithic times. The accumulation of archaeological records and the concordance of different lines of genetic evidence during the last two decades have triggered an interesting debate concerning the role of ancient settlers from the Franco-Cantabrian region in the postglacial resettlement of Europe. Among the Franco-Cantabrian populations, Basques are regarded as one of the oldest and more intriguing human groups of Europe. Recent data on complete mitochondrial DNA genomes focused on macrohaplogroup R0 revealed that Basques harbor some autochthonous lineages, suggesting a genetic continuity since pre-Neolithic times. However, excluding haplogroup H, the most representative lineage of macrohaplogroup R0, the majority of maternal lineages of this area remains virtually unexplored, so that further refinement of the mtDNA phylogeny based on analyses at the highest level of resolution is crucial for a better understanding of the European prehistory. We thus explored the maternal ancestry of 548 autochthonous individuals from various Franco-Cantabrian populations and sequenced 76 mitogenomes of the most representative lineages. Interestingly, we identified three mtDNA haplogroups, U5b1f, J1c5c1 and V22, that proved to be representative of Franco-Cantabria, notably of the Basque population. The seclusion and diversity of these female genetic lineages support a local origin in the Franco-Cantabrian area during the Mesolithic of southwestern Europe, ∼10,000 years before present (YBP), with signals of expansions at ∼3,500 YBP. These findings provide robust evidence of a partial genetic continuity between contemporary autochthonous populations from the Franco-Cantabrian region, specifically the Basques, and Paleolithic/Mesolithic hunter-gatherer groups. Furthermore, our results raise the current proportion (≈15%) of the Franco-Cantabrian maternal gene pool with a putative pre-Neolithic origin to ≈35%, further supporting the notion of a predominant Paleolithic genetic substrate in extant European populations.     

Colorectal Cancers Mimic Structural Organization of Normal Colonic Crypts

Colonic crypts are stereotypical structures with distinct stem cell, proliferating, and differentiating compartments. Colorectal cancers derive from colonic crypt epithelia but, in contrast, form morphologically disarrayed glands. In this study, we investigated to which extent colorectal cancers phenocopy colonic crypt architecture and thus preserve structural organization of the normal intestinal epithelium. A subset of colon cancers showed crypt-like compartments with high WNT activity and nuclear β-Catenin at the leading tumor edge, adjacent proliferation, and enhanced Cytokeratin 20 expression in most differentiated tumor epithelia of the tumor center. This architecture strongly depended on growth conditions, and was fully reproducible in mouse xenografts of cultured and primary colon cancer cells. Full crypt-like organization was associated with low tumor grade and was an independent prognostic marker of better survival in a collection of 221 colorectal cancers. Our findings suggest that full activation of preserved intestinal morphogenetic programs in colon cancer requires in vivo growth environments. Furthermore, crypt-like architecture was linked with less aggressive tumor biology, and may be useful to improve current colon cancer grading schemes.     

A three-dimensional model of the U1 small nuclear ribonucleoprotein particle

Most of the pre-mRNAs in the eukaryotic cell are comprised of protein-coding exons and non-protein-coding introns. The introns are removed and the exons are ligated together, or spliced, by a large, macromolecular complex known as the spliceosome. This RNA-protein assembly is made up of five uridine-rich small nuclear RNAs (U1-, U2-, U4-, U5- and U6-snRNA) as well over 300 proteins, which form small nuclear ribonucleoprotein particles (snRNPs). Initial recognition of the 5′ exon/intron splice site is mediated by the U1 snRNP, which is composed of the U1 snRNA as well as at least ten proteins. By combining structural informatics tools with the available biochemical and crystallographic data, we attempted to simulate a complete, three dimensional U1 snRNP from the silk moth, Bombyx mori. Comparison of our model with empirically derived crystal structures and electron micrographs pinpoints both the strengths and weaknesses in the in silico determination of macromolecular complexes. One of the most striking differences between our model and experimentally generated structures is in the positioning of the U1 snRNA stem-loops. This highlights the continuing difficulties in generating reliable, complex RNA structures; however, three-dimensional modeling of individual protein subunits by threading provided models of biological significance and the use of both automated and manual docking strategies generated a complex that closely reflects the assembly found in nature. Yet, without utilizing experimentally-derived contacts to select the most likely docking scenario, ab initio docking would fall short of providing a reliable model. Our work shows that the combination of experimental data with structural informatics tools can result in generation of near-native macromolecular complexes.     

Free-living dinitrogen-fixing bacteria isolated from petroleum refinery oily sludge

Dinitrogen-fixing activity (acetylene reduction and N(2) fixation) was found in an oily sludge originating from a petroleum refinery. Two representative dinitrogen-fixing bacterial strains were isolated from this oily waste. Their nitrogenase activity was effective when they were cultivated on sterilized sludge or simple carbon substrates (organic acid salts, sugars). Using the classical methods, these strains could not be unambiguously related to other diazotrophic taxa. The landfarming process is widely used for oily sludge disposal; this study shows that oily sludges are more than a simple carbon input into the soil but that they must also be considered as real sources of dinitrogen-fixing and probably degradative microorganisms.     

Sequence diversity of pistil S-proteins associated with gametophytic self-incompatibility in Nicotiana alata

Summary In order to study the extent and nature of differences among various S-allele-associated proteins in N. alata, we carried out comparative studies of seven such proteins. We first isolated and sequenced cDNA clones for the S_z-, S_F11-, S₁-, and S_a-alleles, and then we compared the deduced amino acid sequences both of these four S-proteins and of three previously published S₂-, S₃-, and S₆-proteins. This comparison revealed (1) an average homology of 53.8% among the seven proteins and (2) two homology classes, with S_z and S_F11 in one class and S₁, S₂, S₃, and S₆ in the other class. There are 60 conserved residues, including 9 cysteines. Of the 144 variable residues, 50 were identified as hypervariable based on a calculation of their Similarity Indices. Although conserved, variable, and hypervariable residues are dispersed throughout the protein, some are clustered to form five conserved, five hypervariable, and a number of variable regions. Those variable sites which contain residues conserved within one class of S-proteins but different between classes might provide a clue to the evolutionary relationship of these two classes of S-proteins. The hypervariable residues, which account for sequence variability, may contribute to allelic specificity.     

Stability of Bradyrhizobium japonicum Inoculants after Introduction into Soil

Bradyrhizobium japonicum USDA 125-Sp, USDA 138, and USDA 138-Sm had been used as inoculants for soybean (Glycine max (L.) Merr.) in soils previously free of B. japonicum. At 8 to 13 years after their release, these strains were reisolated from soil samples. A total of 115 isolates were obtained through nodules, and seven colonies were obtained directly by a serological method. The stability of the inoculants was confirmed by comparing the reisolated cultures with their respective parental strains which had been preserved by being lyophilized or stored on a yeast extract-mannitol agar slant at 4°C. Comparisons were made on morphological and serological characters, carbon compound utilization (8 tested), intrinsic antibiotic resistance (9 tested), and enzymatic activity (19 tested). Mucous and nonmucous isolates of serogroup 125 were analyzed for symbiotic effectiveness and restriction fragment hybridization with a DNA probe. Our data suggest that the B. japonicum inoculants have survived for up to 13 years in the soils without significant mutation except for two reisolates with a slightly increased kanamycin resistance level.     

There are two forms of androgen binding protein in human testes. Comparison of their protomeric variants with serum testosterone-estradiol binding globulin

To determine how the androgen binding protein in human testes (hABP) is related to the serum protein, testosterone-estradiol binding globulin (hTeBG), both proteins were isolated and compared. The hABP in extracts of human testes was composed of two molecular species based on concanavalin A (ConA)-Sepharose chromatography. Form I hABP did not interact with ConA while Form II hABP bound to ConA and eluted with alpha-methylmannoside. Form I and Form II hABP from five batches of testes were then purified approximately 30,500- and 30,000-fold to apparent homogeneity by high-performance liquid chromatography and compared with hTeBG isolated from human pregnancy serum. Fractionation of both forms of hABP and hTeBG by polyacrylamide gel electrophoresis in the absence of sodium dodecyl sulfate suggested that the native forms of these proteins were indistinguishable. However, analysis of the purified proteins on sodium dodecyl sulfate-containing polyacrylamide gels indicated that all three were dimers and that each was composed of monomers of at least two sizes which were not present in equimolar concentrations. Two distinctive monomers or protomers of each protein were designated as heavy (H) and light (L) according to their electrophoretic mobilities on sodium dodecyl sulfate-polyacrylamide gels. The H and L protomers of Form I hABP showed apparent molecular weights of 55,000 and 52,000, respectively, in all preparations and were usually present in a 4:5 ratio (H:L). The two components of Form II hABP had apparent molecular weights of 53,000 and 48,000, respectively, and existed in a ratio of approximately 20:1. These two components could not be distinguished in some preparations where Form II hABP migrated as a broad band rather than as distinct protomers. By contrast, hTeBG, which was similar to Form II hABP with respect to ConA binding, always exhibited discrete H and L protomers in a 10:1 ratio. Photolysis of these highly purified proteins with delta 6-[3H]testosterone resulted in specific covalent labeling of their binding sites, confirming that the products identified by silver staining and immunoblotting were indeed steroid binding proteins. The H and L protomers of Form I hABP and hTeBG were separated and examined by peptide mapping using Staphylococcus aureus protease V8 and chymotrypsin. The comparison of the respective fragmentation patterns of protomers indicated that Form I hABP and hTeBG contained distinctive peptides.(ABSTRACT TRUNCATED AT 400 WORDS)     

The role of the carbohydrate moiety on the size heterogeneity and immunologic determinants of human testosterone-estradiol-binding globulin

Human testosterone-estradiol-binding globulin (hTeBG) has been purified to apparent homogeneity by several laboratories using procedures which, in most instances, were labor intensive. In this report, hTeBG was purified from pregnancy serum by a newly developed two step procedure involving sequential affinity chromatography and ion-exchange high performance liquid chromatography (ion-exchange HPLC). The purity of the final product was confirmed by silver stained SDS-polyacrylamide gel and reverse phase HPLC monitored at 206 nm. hTeBG purified by ion-exchange-HPLC maintained binding activity by Dextran coated charcoal (DCC) assay and size heterogeneity on SDS-polyacrylamide gels which were indistinguishable from those of the proteins purified by conventional chromatography. Removal of the carbohydrate moiety from the molecule by both enzymatic and chemical treatment reduced the apparent molecular size and eliminated lectin binding of hTeBG subunits. Deglycosylation did not, however, abolish or alter the distribution of the protomeric forms of this subunit. We conclude that hTeBG is a dimer whose monomer exhibits two protomeric forms which is not a result of carbohydrate heterogeneity. In addition, disialylated and deglycosylated hTeBG exhibited antigenic determinants identical to the native protein.     

The relationship between hydrogen metabolism,sulfate reduction and nitrogen fixation in sulfate reducers

Summary Hydrogenase and nitrogenase activities of sulfate-reducing bacteria allow their adaptation to different nutritional habits even under adverse conditions. These exceptional capabilities of adaptation are important factors in the understanding of their predominant role in problems related to anaerobic metal corrosion. Although the D₂–H⁺ exchange reaction indicated thatDesulfovibrio desulfuricans strain Berre-Sol andDesulfovibrio gigas hydrogenases were reversible, the predominant activity in vivo was hydrogen uptake. Hydrogen production was restricted to some particular conditions such as sulfate or nitrogen starvation. Under diazotrophic conditions, a transient hydrogen evolution was followed by uptake when dinitrogen was effectively fixed. In contrast, hydrogen evolution proceeded when acetylene was substituted as the nitrogenase substrate. Hydrogen can thus serve as an electron donor in sulfate reduction and nitrogen metabolism.