SidL, an Aspergillus fumigatus transacetylase involved in biosynthesis of the siderophores ferricrocin and hydroxyferricrocin

The opportunistic fungal pathogen Aspergillus fumigatus produces four types of siderophores, low-molecular-mass iron chelators: it excretes fusarinine C (FsC) and triacetylfusarinine C (TAFC) for iron uptake and accumulates ferricrocin (FC) for hyphal and hydroxyferricrocin (HFC) for conidial iron distribution and storage. Siderophore biosynthesis has recently been shown to be crucial for fungal virulence. Here we identified a new component of the fungal siderophore biosynthetic machinery: AFUA_1G04450, termed SidL. SidL is conserved only in siderophore-producing ascomycetes and shows similarity to transacylases involved in bacterial siderophore biosynthesis and the N(5)-hydroxyornithine:anhydromevalonyl coenzyme A-N(5)-transacylase SidF, which is essential for TAFC biosynthesis. Inactivation of SidL in A. fumigatus decreased FC biosynthesis during iron starvation and completely blocked FC biosynthesis during iron-replete growth. In agreement with these findings, SidL deficiency blocked conidial accumulation of FC-derived HFC under iron-replete conditions, which delayed germination and decreased the size of conidia and their resistance to oxidative stress. Remarkably, the sidL gene is not clustered with other siderophore-biosynthetic genes, and its expression is not affected by iron availability. Tagging of SidL with enhanced green fluorescent protein suggested a cytosolic localization of the FC-biosynthetic machinery. Taken together, these data suggest that SidL is a constitutively active N(5)-hydroxyornithine-acetylase required for FC biosynthesis, in particular under iron-replete conditions. Moreover, this study revealed the unexpected complexity of siderophore biosynthesis, indicating the existence of an additional, iron-repressed N(5)-hydroxyornithine-acetylase.     

Hybrid speciation through sorting of parental incompatibilities in Italian sparrows

Speciation by hybridization is emerging as a significant contributor to biological diversification. Yet, little is known about the relative contributions of (i) evolutionary novelty and (ii) sorting of pre‐existing parental incompatibilities to the build‐up of reproductive isolation under this mode of speciation. Few studies have addressed empirically whether hybrid animal taxa are intrinsically isolated from their parents, and no study has so far investigated by which of the two aforementioned routes intrinsic barriers evolve. Here, we show that sorting of pre‐existing parental incompatibilities contributes to intrinsic isolation of a hybrid animal taxon. Using a genomic cline framework, we demonstrate that the sex‐linked and mitonuclear incompatibilities isolating the homoploid hybrid Italian sparrow at its two geographically separated hybrid–parent boundaries represent a subset of those contributing to reproductive isolation between its parent species, house and Spanish sparrows. Should such a sorting mechanism prove to be pervasive, the circumstances promoting homoploid hybrid speciation may be broader than currently thought, and indeed, there may be many cryptic hybrid taxa separated from their parent species by sorted, inherited incompatibilities.     

Impact of miR-21, miR-126 and miR-221 as Prognostic Factors of Clear Cell Renal Cell Carcinoma with Tumor Thrombus of the Inferior Vena Cava

Clear cell renal cell carcinoma (ccRCC) characterized by a tumor thrombus (TT) extending into the inferior vena cava (IVC) generally indicates poor prognosis. Nevertheless, the risk for tumor recurrence after nephrectomy and thrombectomy varies. An applicable and accurate prediction system to select ccRCC patients with TT of the IVC (ccRCC/TT) at high risk after nephrectomy is urgently needed, but has not been established up to now. To our knowledge, a possible role of microRNAs (miRs) for the development of ccRCC/TT or their impact as prognostic markers in ccRCC/TT has not been explored yet. Therefore, we analyzed the expression of the previously described onco-miRs miR-200c, miR-210, miR-126, miR-221, let-7b, miR-21, miR-143 and miR-141 in a study collective of 74 ccRCC patients. Using the expression profiles of these eight miRs we developed classification systems that accurately differentiate ccRCC from non-cancerous renal tissue and ccRCC/TT from tumors without TT. In the subgroup of 37 ccRCC/TT cases we found that miR-21, miR-126, and miR-221 predicted cancer related death (CRD) accurately and independently from other clinico-pathological features. Furthermore, a combined risk score based on the expression of miR-21, miR-126 and miR-221 was developed and showed high sensitivity and specificity to predict cancer specific survival (CSS) in ccRCC/TT. Using the combined risk score we were able to classify ccRCC/TT patients correctly into high and low risk cases. The risk stratification by the combined risk score (CRS) will benefit from further cohort validation and might have potential for clinical application as a molecular prediction system to identify high- risk ccRCC/TT patients.     

Endosomal translocation of vertebrate DNA activates dendritic cells via TLR9-dependent and -independent pathways

TLRs discriminate foreign from self via their specificity for pathogen-derived invariant ligands, an example being TLR9 recognizing bacterial unmethylated CpG motifs. In this study we report that endosomal translocation of CpG DNA via the natural endocytotic pathway is inefficient and highly saturable, whereas endosomal translocation of DNA complexed to the cationic lipid N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium methylsulfate (DOTAP) is not. Interestingly, DOTAP-mediated enhanced endosomal translocation of otherwise nonstimulatory vertebrate DNA or of certain noncanonical CpG motifs triggers robust dendritic cell activation in terms of both up-regulation of CD40/CD69 and cytokine production, such as type I IFN and IL-6. We report that the stimulatory activity of phosphorothioated noncanonical CpG oligodeoxynucleotides is TLR9 dependent, whereas phosphodiester DNA, such as vertebrate DNA, in addition trigger TLR9-independent pathways. We propose that the inefficiency of the natural route for DNA internalization hinders low affinity TLR9 ligands in endosomes to reach threshold concentrations required for TLR9 activation. Endosomal compartmentalization of TLR9 may thus reflect an evolutionary strategy to avoid TLR9 activation by self-DNA.     

Odoriferous compounds from the flowers of the conifers Picea abies,pinus sylvestris and Larix sibirica

A GC/MS analysis of the volatile constituents from the flowers of Norway Spruce, Picea abies, has been carried out. The volatile constituents of the female flowers were distinctly different from those of the male flowers and the twigs. Characteristic constituents are methyl and ethyl benzoate, methyl and ethyl salicylate, methyl and ethyl butanoate, borneol and bornyl acetate. In the scent from the male flowers we could only detect the same monoterpenes as in the twigs. In Larix sibirica methyl benzoate, methyl salicylate, borneol and bornyl acetate were detected in the female flowers and, in the female flowers of Pinus sylvestris, methyl salicylate was found.     

Recognition by human Vγ9/Vδ2 T cells of melanoma cells upon fusion with Daudi cells

 Daudi Burkitt’s lymphoma cells, unlike other tumor cell lines, stimulate human T cells coexpressing the variable (V) region genes TCRG-V9 and V TCRD-V2 to proliferate and secrete lymphokines. Hybrids, derived by the fusion of Daudi cells with the human melanoma cell line MZ2-MEL 2.2, retain the morphology of melanoma cells. Unlike the parental melanoma cell line, these Daudi × MZ2-MEL 2.2 hybrids stimulate secretion of tumor necrosis factor (TNF) and granulocyte/macrophage colony stimulating factor (GM-CSF) by CD4-positive Vγ9/Vδ2 T-cell clones. Whereas the stimulator phenotype of Daudi cells behaves as a dominant trait in Daudi × melanoma hybrids, the expression of B-cell differentiation markers is suppressed. Thus, the γ/δ T-cell ligand expressed by Daudi cells behaves as a dominant tumor antigen in Daudi × melanoma hybrids and is unrelated to the differentiated B-cell phenotype. Dominant expression of the Daudi ligand for human Vγ9/Vδ2 T cells in these hybrids may provide a basis for defining the stimulatory principle at the molecular level. Received: 2 May 1996 / Revised: 15 July 1996     

Callus induction and plant regeneration from gladiolus

A method for the initiation of callus capable of plant regeneration from in vivo grown cormels of gladiolus (Gladiolus x grandiflorus Hort.) is described. Sliced cormels of the large-flowering hybrid, Peter Pears were cultured in vitro on a modified Murashige and Skoog medium, supplemented with various auxins. Yellow callus, which was either friable or compact, could be induced on all media tested. Callus induced on media with naphthaleneacetic acid failed to proliferate. Callus induced on media with 9 mM 2,4-dichlorophenoxyacetic acid showed the best growth. Addition of micro-elements and vitamins increased the induction and growth of callus capable of plant regeneration. Explants taken from the middle part of the cormels had the highest competence for callus initiation. Callus was induced on several gladiolus hybrids and the South African species G. garnierii Klatt. Callus induction was genotype dependent and among the cultivars tested, Peter Pears and White Prosperity were superior with respect to callus production on the media with either 2,4-dichlorophenoxyacetic acid or picloram. Plants were regenerated from yellow compact callus of all genotypes on media containing zeatin and benzyladenine in various concentrations.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - MS Murashige and Skoog basal salt and vitamins (1962) - CI callus induction medium - NAA -naphthaleneacetic acid - BA 6-benzyladenine - picloram 4-amino-3,5,6-trichloro-2-pyridinecarboxylic acid - zeatin 6-[4-hydroxy-3-methylbut-2-enylamino]purine