More constraint on ParaHox than Hox gene families in early metazoan evolution

Hox and ParaHox (H/P) genes belong to evolutionary-sister clusters that arose through duplication of a ProtoHOX cluster early in animal evolution. In contrast to bilaterians, cnidarians express, beside PG1, PG2 and Gsx orthologs, numerous Hox-related genes with unclear origin. We characterized from marine hydrozoans three novel Hox-related genes expressed at medusa and polyp stages, which include a Pdx/Xlox ParaHox ortholog induced 1 day later than Gsx during embryonic development. To reconstruct H/P genes' early evolution, we performed multiple systematic comparative phylogenetic analyses, which identified derived sequences that blur the phylogenetic picture, recorded dramatically different evolutionary rates between ParaHox and Hox in cnidarians and showed the unexpected grouping of [Gsx-Pdx/Xlox-PG2-PG3] families in a single metagroup distinct from PG1. We propose a novel more parsimonious evolutionary scenario whereby H/P genes originated from a [Gsx-Pdx/Xlox-PG2-PG3]-related ProtoHox gene, the «posterior» and «anterior» H/P genes appearing secondarily. The ProtoHOX cluster would have contained the three Gsx/PG2, Pdx/PG3, Cdx/PG9 paralogs and produced through tandem duplication the primordial HOX and ParaHOX clusters in the Cnidaria-Bilateria ancestor. The stronger constraint on cnidarian ParaHox genes suggests that the primary function of pre-bilaterian H/P genes was to drive cellular evolutionary novelties such as neurogenesis rather than axis specification.     

Mechanism of cell killing after ionizing radiation by a dominant negative DNA polymerase beta

Several types of DNA lesion are induced after ionizing irradiation (IR) of which double strand breaks (DSBs) are expected to be the most lethal, although single strand breaks (SSBs) and DNA base damages are quantitatively in the majority. Proteins of the base excision repair (BER) pathway repair these numerous lesions. DNA polymerase beta has been identified as a crucial enzyme in BER and SSB repair (SSBR). We showed previously that inhibition of BER/SSBR by expressing a dominant negative DNA polymerase beta (polβDN) resulted in radiosensitization. We hypothesized increased kill to result from DSBs arising from unrepaired SSBs and BER intermediates. We find here higher numbers of IR-induced chromosome aberrations in polβDN expressing cells, confirming increased DSB formation. These aberrations did not result from changes in DSB induction or repair of the majority of lesions. SSB conversion to DSBs has been shown to occur during replication. We observed an increased induction of chromatid aberrations in polβDN expressing cells after IR, suggesting such a replication-dependence of secondary DSB formation. We also observed a pronounced increase of chromosomal deletions, the most likely cause of the increased kill. After H₂O₂ treatment, polβDN expression only resulted in increased chromatid (not chromosome) aberrations. Together with the lack of sensitization to H₂O₂, these data further suggest that the additional secondarily induced lethal DSBs resulted from repair attempts at complex clustered damage sites, unique to IR. Surprisingly, the polβDN induced increase in residual γH2AX foci number was unexpectedly low compared with the radiosensitization or induction of aberrations. Our data thus demonstrate the formation of secondary DSBs that are reflected by increased kill but not by residual γH2AX foci, indicating an escape from γH2AX-mediated DSB repair. In addition, we show that in the polβDN expressing cells secondary DSBs arise in a radiation-specific and partly replication-dependent manner.     

Caldinitratiruptor microaerophilus, gen. nov., sp. nov. isolated from a French hot spring (Chaudes-Aigues, Massif Central): a novel cultivated facultative microaerophilic anaerobic thermophile pertaining to the Symbiobacterium branch within the Firmicutes

A novel facultative microaerophilic nitrate-reducing bacterium designated CA62N^T was isolated from a thermal spring in France. Cells were non-motile rods (2–3 × 0.2 μm) and showed low cytoplasmic density when observed under a phase-contrast microscope. Strain CA62N^T grew at temperatures between 50 and 75°C (optimum 65°C) and at a pH between 6.3 and 7.9 (optimum 7.0). NaCl was not required for growth but was tolerated up to 10 gl⁻¹. Sulfate, thiosulfate, elemental sulfur, sulfite, and nitrite were not used as electron acceptors. Nitrate was reduced to nitrite. Strain CA62N^T used lactate, pyruvate, glucose, mannose, fructose, and casamino acids and some amino acids as electron donors only in the presence of nitrate as electron acceptor. None of these substrates was fermented. The main end-products of glucose oxidation were acetate, CO₂, and traces of H₂. The G + C content of the genomic DNA was 70.3 mol% (HPLC techniques). Phylogenetic analysis of the small-subunit (SSU) ribosomal RNA (rRNA) gene sequence indicated that strain CA62N^T was affiliated to the Symbiobacterium branch within the Firmicutes and had Symbiobacterium thermophilum and “S. toebii” as its closest phylogenetic relatives. On the basis of phylogenetical and physiological characteristics, strain CA62N^T is proposed to be the type strain for the novel species in the novel genus, Caldinitratiruptor microaerophilus gen. nov., sp. nov. (DSM 22660, JCM 16183).     

Desulfosporosinus acidiphilus sp. nov.: a moderately acidophilic sulfate-reducing bacterium isolated from acid mining drainage sediments

An obligately anaerobic, spore-forming, acidophilic sulfate-reducing bacterium, strain SJ4^T, was isolated from an acid mining effluent decantation pond sediment sample (pH around 3.0). Cells were Gram negative, non-motile, curved rods occurring singly. Strain SJ4^T grew at pH 3.6–5.5 with an optimum at pH 5.2. Strain SJ4^T utilized H₂, lactate, pyruvate, glycerol, glucose, and fructose as electron donors. Lactate and glucose were weakly used. Sulfate was used as electron acceptors, but not sulfite, elemental sulfur, arsenate (V), and fumarate. The G + C content of genomic DNA was 42.3 mol% (HPLC). 16S rRNA gene sequence analysis indicated that strain SJ4^T belonged to the genus Desulfosporosinus within the family Peptococcaceae in the phylum Firmicutes. The level of 16S rRNA gene sequence similarity with other Desulfosporosinus species was 94.7–96.2%, D. orientis DSM 765^T (similarity of 96.2%) and D. auripigmenti DSM 13351^T (similarity of 95%) being its closest relatives. DNA–DNA relatedness values with D. orientis and D. auripigmenti were 16.5 and 31.8%, respectively. On the basis of phenotypic, phylogenetic, and genetic characteristics, strain SJ4^T represents a novel species within the genus Desulfosporosinus, for which the name Desulfosporosinus acidiphilus sp. nov. is proposed. The type strain is SJ4^T (=DSM 22704^T = JCM 16185^T).     

Fatty Acid- and Cholesterol Transporter Protein Expression along the Human Intestinal Tract

Background

Protein distribution profiles along the human intestinal tract of transporters involved in the absorption of cholesterol and long-chain fatty acids (LCFA) have been scarcely evaluated.

Methodology/Principal Findings

In post-mortem samples from 11 subjects, intestinal transporter distribution profiles were determined via Western Blot. Differences in transporter protein levels were statistically tested using ANOVA and Tukey''s Post Hoc comparisons. Levels in all segments were expressed relative to those in duodenum. Except for ABCG5 and FATP4, levels (mean±SEM) were the highest in the ileum. For ABCA1, ileal levels (1.80±0.26) differed significantly from those in duodenum (P = 0.049) and proximal colon (0.92±0.14; P = 0.029). ABCG8 levels in ileum (1.91±0.30) differed from those in duodenum (P = 0.041) and distal colon (0.84±0.22; P = 0.010) and jejunum (1.64±0.26) tended to be higher than distal colon (0.84±0.22; P = 0.087). Ileal NPC1L1 levels (2.56±0.51) differed from duodenum levels (P = 0.019) and from distal colon (1.09±0.22; P = 0.030). There was also a trend (P = 0.098) for higher jejunal (2.23±0.37) than duodenal NPC1L1 levels. The levels of ABCG5 did not correlate with those of ABCG8. FAT/CD36 levels in ileum (2.03±0.42) differed from those in duodenum (P = 0.017), and proximal and distal colon (0.89±0.13 and 0.97±0.15 respectively; P = 0.011 and P = 0.014). FABPpm levels in ileum (1.04±0.13) differed from proximal (0.64±0.07; P = 0.026) and distal colon (0.66±0.09; P = 0.037).

Conclusions/Significance

The distribution profiles showed a bell-shape pattern along the GI-tract with the highest levels in ileum for ABCA1, ABCG8, NPC1L1, FATCD36 and FABPm, suggesting a prominent role for ileum in transporter-mediated uptake of cholesterol and LCFAs.     

Fine mapping of Co-x,an anthracnose resistance gene to a highly virulent strain of Colletotrichum lindemuthianum in common bean

Key message

The Co - x anthracnose R gene of common bean was fine-mapped into a 58 kb region at one end of chromosome 1, where no canonical NB-LRR-encoding genes are present in G19833 genome sequence.

Abstract

Anthracnose, caused by the phytopathogenic fungus Colletotrichum lindemuthianum, is one of the most damaging diseases of common bean, Phaseolus vulgaris. Various resistance (R) genes, named Co-, conferring race-specific resistance to different strains of C. lindemuthianum have been identified. The Andean cultivar JaloEEP558 was reported to carry Co-x on chromosome 1, conferring resistance to the highly virulent strain 100. To fine map Co-x, 181 recombinant inbred lines derived from the cross between JaloEEP558 and BAT93 were genotyped with polymerase chain reaction (PCR)-based markers developed using the genome sequence of the Andean genotype G19833. Analysis of RILs carrying key recombination events positioned Co-x at one end of chromosome 1 to a 58 kb region of the G19833 genome sequence. Annotation of this target region revealed eight genes: three phosphoinositide-specific phospholipases C (PI-PLC), one zinc finger protein and four kinases, suggesting that Co-x is not a classical nucleotide-binding leucine-rich encoding gene. In addition, we identified and characterized the seven members of common bean PI-PLC gene family distributed into two clusters located at the ends of chromosomes 1 and 8. Co-x is not a member of Co-1 allelic series since these two genes are separated by at least 190 kb. Comparative analysis between soybean and common bean revealed that the Co-x syntenic region, located at one end of Glycine max chromosome 18, carries Rhg1, a major QTL contributing to soybean cyst nematode resistance. The PCR-based markers generated in this study should be useful in marker-assisted selection for pyramiding Co-x with other R genes.     

Modulation of Bax mitochondrial insertion and induced cell death in yeast by mammalian protein kinase Cα

Protein kinase Cα (PKCα) is a classical PKC isoform whose involvement in cell death is not completely understood. Bax, a major member of the Bcl-2 family, is required for apoptotic cell death and regulation of Bax translocation and insertion into the outer mitochondrial membrane is crucial for regulation of the apoptotic process. Here we show that PKCα increases the translocation and insertion of Bax c-myc (an active form of Bax) into the outer membrane of yeast mitochondria. This is associated with an increase in cytochrome c (cyt c) release, reactive oxygen species production (ROS), mitochondrial network fragmentation and cell death. This cell death process is regulated, since it correlates with an increase in autophagy but not with plasma membrane permeabilization. The observed increase in Bax c-myc translocation and insertion by PKCα is not due to Bax c-myc phosphorylation, and the higher cell death observed is independent of the PKCα kinase activity. PKCα may therefore have functions other than its kinase activity that aid in Bax c-myc translocation and insertion into mitochondria. Together, these results give a mechanistic insight on apoptosis regulation by PKCα through regulation of Bax insertion into mitochondria.