Genome-wide CRISPR Screens in T Helper Cells Reveal Pervasive Crosstalk between Activation and Differentiation

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Nematodes found in Nile crocodiles in the Kruger National Park,South Africa,with redescriptions of Multicaecum agile (Wedl, 1861) (Heterocheilidae) and Camallanus kaapstaadi Southwell & Kirshner, 1937 (Camallanidae)

Sixteen Nile crocodiles were collected in the Kruger National Park, South Africa and vicinity during 2010 and 2011. A total of 11 nematode species representing six families were recovered. Heterocheilids were the dominant group, comprising five species, with Dujardinascaris madagascariensis (Chabaud & Caballero, 1966) being the most prevalent (75%), followed by Ingwenascaris sprenti Junker & Mutafchiev, 2017 (68.8%), which was also the second most numerous nematode. While less prevalent (31.3%), Typhlophoros kwenae Junker & Mutafchiev, 2017 was the most abundant species. Micropleura huchzermeyeri Junker & Mutafchiev, 2017 (Micropleuridae) was collected from five crocodiles and Crocodylocapillaria sp. (Capillariidae) occurred in a single host. Three nematodes, Camallanus kaapstaadi Southwell & Kirshner, 1937, Spirocamallanus sp. (both Camallanidae) and Ascarophis sp. (Cystidicolidae), are considered accidental infections, likely ingested with the hosts’ prey. Our findings of D. dujardini (Travassos, 1920), D. madagascariensis and Multicaecum agile (Wedl, 1861) in South Africa constitute new geographical records. Crocodylocapillaria sp. represents a new host and geographical record, while T. kwenae, I. sprenti and M. huchzermeyeri have been described as new species during the course of this survey. Multicaecum agile is here redescribed based on light and scanning electron microscopy. Previously undescribed morphological characters of C. kaapstaadi, typically a parasite of Xenopus spp. (Amphibia: Pipidae), but here found in two Nile crocodiles, are also presented.

     

Reconstitution of EBV-directed T cell immunity by adoptive transfer of peptide-stimulated T cells in a patient after allogeneic stem cell transplantation for AITL

Reconstitution of the T cell repertoire after allogeneic stem cell transplantation is a long and often incomplete process. As a result, reactivation of Epstein-Barr virus (EBV) is a frequent complication that may be treated by adoptive transfer of donor-derived EBV-specific T cells. We generated donor-derived EBV-specific T cells by stimulation with peptides representing defined epitopes covering multiple HLA restrictions. T cells were adoptively transferred to a patient who had developed persisting high titers of EBV after allogeneic stem cell transplantation for angioimmunoblastic T-cell lymphoma (AITL). T cell receptor beta (TCRβ) deep sequencing showed that the T cell repertoire of the patient early after transplantation (day 60) was strongly reduced and only very low numbers of EBV-specific T cells were detectable. Manufacturing and in vitro expansion of donor-derived EBV-specific T cells resulted in enrichment of EBV epitope-specific, HLA-restricted T cells. Monitoring of T cell clonotypes at a molecular level after adoptive transfer revealed that the dominant TCR sequences from peptide-stimulated T cells persisted long-term and established an EBV-specific TCR clonotype repertoire in the host, with many of the EBV-specific TCRs present in the donor. This reconstituted repertoire was associated with immunological control of EBV and with lack of further AITL relapse.     

Expression of the Gm1-Species, [NeuN]-GM1, in a Case of Human Glioma

Altered glycosylation is a common feature in tumors of various kind and particular interest has been focused on the expression of tumor-associated gangliosides. We have previously identified some human glioma-associated gangliosides and in this study yet another, not previously described, ganglioside has been isolated. The ganglioside was prepared from human glioma tissue taken at autopsy. The new ganglioside bound cholera-toxin B-subunit and its structure was confirmed by fast atom bombardment—mass spectrometry to be NeuN-GM1 (II³NeuNH₂-GgOse₄Cer). In the dissected tumor specimen, the concentration of NeuN-GM1 was 0.1 mol/g wet weight and accounted for approximately 20% of the monosialoganglioside fraction. Normal human brain tissue specimens (n = 10) did not contain detectable (>0.5 nmol/g wet weight of tissue) amounts of NeuN-GM1, indicating that this ganglioside might be associated with human glioma. However, none of the 17 other tumour specimens reveal any detectable amounts of this ganglioside. In conclusion, NeuN GM1 is a glioma-associated ganglioside but its exceptional expression limits its relevance as a molecule involved in general tumor biology.     

Thioredoxin can influence gene expression by affecting gyrase activity

The expression of many genes of facultatively photosynthetic bacteria of the genus Rhodobacter is controlled by the oxygen tension. Among these are the genes of the puf and puc operons, which encode proteins of the photosynthetic apparatus. Previous results revealed that thioredoxins are involved in the regulated expression of these operons, but it remained unsolved as to the mechanisms by which thioredoxins affect puf and puc expression. Here we show that reduced TrxA of Rhodobacter capsulatus and Rhodobacter sphaeroides and oxidized TrxC of R.capsulatus interact with DNA gyrase and alter its DNA supercoiling activity. While TrxA enhances supercoiling, TrxC exerts a negative effect on this activity. Furthermore, inhibition of gyrase activity strongly reduces puf and puc expression. Our results reveal a new signaling pathway by which oxygen can affect the expression of bacterial genes.     

Inhibition of basal and interleukin-1-induced VCAM-1 expression by phospholipid hydroperoxide glutathione peroxidase and 15-lipoxygenase in rabbit aortic smooth muscle cells

Cytokines or hydroperoxides upregulate cell adhesion molecules (CAM) in early stages of atherosclerosis. VCAM-1 expression was therefore investigated in rabbit aortic smooth muscle cells (SMC) stably transfected either with phospholipid hydroperoxide glutathione peroxidase (PHGPx; SMCPHGPx) as a hydroperoxide-reducing enzyme or with 15-lipoxygenase (15-LOX; SMCLOX) as a hydroperoxide-producing enzyme. Transfected cells showed up to 3-fold enhanced PHGPx and a marked LOX activity, respectively, that was absent in controls. Intracellular hydroperoxides were 6-fold higher in SMCLOX than in SMC or SMCPHGPx. Intracellular protein thiols were decreased by 50 and 90% in SMCPHGPx and SMCLOX, respectively. Glutathione mixed disulfides were tentatively increased from SMC via SMCPHGPx to SMCLOX, accordingly. Thiol reduction with tris(2-carboxyethyl)phosphine completely restored protein thiols in SMCPHGPx, whereas in SMCLOX only 60% of control values were recovered. Basal VCAM-1 mRNA levels were decreased by 50% in SMCPHGPx and 75% in SMCLOX. VCAM-1-inducibility was abrogated in SMCLOX but not in SMCPHGPx. Accordingly, NFkappaB-driven reporter gene activation by IL-1 was unaffected in SMCPHGPx but abolished in SMCLOX. The data confirm that PHGPx overexpression dampens CAM expression either by lowering stimulatory hydroperoxides or by using hydroperoxides for protein modification. But hydroperoxides, when constitutively overproduced as in SMCLOX, inhibit CAM expression and render cells refractory to IL-1 stimulation likely due to oxidation of protein thiols of the signaling system.     

Resource pulses,species interactions,and diversity maintenance in arid and semi-arid environments

Arid environments are characterized by limited and variable rainfall that supplies resources in pulses. Resource pulsing is a special form of environmental variation, and the general theory of coexistence in variable environments suggests specific mechanisms by which rainfall variability might contribute to the maintenance of high species diversity in arid ecosystems. In this review, we discuss physiological, morphological, and life-history traits that facilitate plant survival and growth in strongly water-limited variable environments, outlining how species differences in these traits may promote diversity. Our analysis emphasizes that the variability of pulsed environments does not reduce the importance of species interactions in structuring communities, but instead provides axes of ecological differentiation between species that facilitate their coexistence. Pulses of rainfall also influence higher trophic levels and entire food webs. Better understanding of how rainfall affects the diversity, species composition, and dynamics of arid environments can contribute to solving environmental problems stemming from land use and global climate change.     

The structure of bovine F1-ATPase inhibited by ADP and beryllium fluoride

The structure of bovine F1-ATPase inhibited with ADP and beryllium fluoride at 2.0 angstroms resolution contains two ADP.BeF3- complexes mimicking ATP, bound in the catalytic sites of the beta(TP) and beta(DP) subunits. Except for a 1 angstrom shift in the guanidinium of alphaArg373, the conformations of catalytic side chains are very similar in both sites. However, the ordered water molecule that carries out nucleophilic attack on the gamma-phosphate of ATP during hydrolysis is 2.6 angstroms from the beryllium in the beta(DP) subunit and 3.8 angstroms away in the beta(TP) subunit, strongly indicating that the beta(DP) subunit is the catalytically active conformation. In the structure of F1-ATPase with five bound ADP molecules (three in alpha-subunits, one each in the beta(TP) and beta(DP) subunits), which has also been determined, the conformation of alphaArg373 suggests that it senses the presence (or absence) of the gamma-phosphate of ATP. Two catalytic schemes are discussed concerning the various structures of bovine F1-ATPase.