Evidence of the involvement of a cyclase gene in the biosynthesis of ochratoxin A in Aspergillus carbonarius

Ochratoxin A (OTA) is a well-known mycotoxin with wide distribution in food and feed. Fungal genome sequencing has great utility for identifying secondary metabolites gene clusters for known and novel compounds. A comparative analysis of the OTA-biosynthetic cluster in A. steynii, A. westerdijkiae, A. niger, A. carbonarius, and P. nordicum has revealed a high synteny in OTA cluster organization in five structural genes (otaA, otaB, ota, otaR1, and otaD). Moreover, a recent detailed comparative genome analysis of Aspergilli OTA producers led to the identification of a cyclase gene, otaY, located in the OTA cluster between the otaA and otaB genes, encoding for a predicted protein with high similarity to SnoaLs domain. These proteins have been shown to catalyze ring closure steps in the biosynthesis of polyketide antibiotics produced in Streptomyces. In the present study, we demonstrated an upregulation of the cyclase gene in A. carbonarius under OTA permissive conditions, consistent with the expression trends of the other OTA cluster genes and their role in OTA biosynthesis by complete gene deletion. Our results pointed out the involvement of a cyclase gene in OTA biosynthetic pathway for the first time. They represent a step forward in the understanding of the molecular basis of OTA biosynthesis in A. carbonarius

Mechanisms of goethite dissolution in the presence of desferrioxamine B and Suwannee River fulvic acid at pH 6.5

Siderophores are Fe3+ specific low MW chelating ligands secreted by microorganisms in response to Fe stress. Low MW organic acids such as oxalate have been shown to enhance siderophore mediated dissolution of Fe3+ oxides. However, the effect of fulvic acid presence on siderophore function remains unknown. We used batch dissolution experiments to investigate Fe release from goethite in the goethite-fulvic acid desferrioxamine B (goethite-SRFA-DFOB) ternary system. Experiments were conducted at pH 6.5 while varying reagent addition sequence. FTIR and UV-Vis spectroscopy were employed to characterise the Fe-DFOB, Fe-SRFA and DFOB–SRFA complexes. Iron released from goethite in the presence of SRFA alone was below detection limit. In the presence of both SRFA and DFOB, dissolved Fe increased with reaction time, presence of the DFOB-SRFA complex, and where SRFA was introduced prior to DFOB. FTIR data show that in the ternary system, Fe3+ is complexed primarily to oxygen of the DFOB hydroxamate group, whilst the carboxylate C=O of SRFA forms an electrostatic association with the terminal NH3+ of DFOB. We propose that SRFA sorbed to goethite lowers the net positive charge of the oxide surface, thus facilitating adsorption of cationic DFOB and subsequent Fe3+ chelation and release. Furthermore, the sorbed SRFA weakens Fe-O bonds at the goethite surface, increasing the population of kinetically labile Fe. This work demonstrates the positive, though indirect role of SRFA in increasing the bioavailability of Fe3+

Effect of desferrioxamine B and Suwannee River fulvic acid on Fe(III) release and Cr(III) desorption from goethite

Siderophores are biogenic chelating ligands that facilitate the solubilization of Fe(III) and form stable complexes with a range of contaminant metals and therefore may significantly affect their biogeochemical cycling. Desferrioxamine B (DFOB) is a trihydroxamate siderophore that acts synergistically with fulvic acid and low molecular weight organic ligands to release Fe from Fe(III) oxides. We report the results of batch dissolution experiments in which we determine the rates of Cr(III) desorption and Fe(III) release from Cr(III)-treated synthetic goethite as influenced by DFOB, by fulvic acid, and by the two compounds in combination. We observed that adsorbed Cr(III) at 3% surface coverage significantly reduced Fe(III) release from goethite for all combinations of DFOB and fulvic acid. When DFOB (270 µM) was the only ligand present, dissolved Fe(III) and Cr(III) increased approximately 1000-fold and 16-fold, respectively, as compared to the ligand-free system, a difference we attribute to the slow rate of water exchange of Cr(III). Suwannee River fuvic acid (SRFA) acts synergistically with DFOB by (i) reducing the goethite surface charge leading to increased HDFOB+ surface excess and by (ii) forming aqueous Fe(III)-SRFA species whose Fe(III) is subsequently removed by DFOB to yield aqueous Fe(III)-DFOB complexes. These observations shed new light on the synergistic relationship between DFOB and fulvic acid and reveal the mechanisms of Fe(III) acquisition available to plants and micro-organisms in Cr(III) contaminated environments

Sailing into the wind : new disciplines in Australian higher education

Much is made of the potential of lifelong learning for individuals and organisations. In this article we tend to make much less of it, certainly with respect to its use in universities to discipline academics. Nevertheless, we argue that academics now need to re-learn the positions they occupy and the stances they take in response to the marketisation of Australian universities. In particular, we suggest that the position of (pure) critique no longer commands attention in Australian contexts of higher education, although the paper does not suggest a disregard for a critical stance purely for the sake of participation. It is in understanding the interconnections between position and stance , and how they might be strategically performed during the everyday practices of academics, that a more promising way of engaging with the venalities of the market is envisaged; a strategy that could be described as \u27sailing into the wind\u27. In discussing these matters, the paper draws on semi-structured interviews with academics located in university faculties/departments/schools of education along Australia\u27s eastern seaboard

Novel Strains of Mice Deficient for the Vesicular Acetylcholine Transporter: Insights on Transcriptional Regulation and Control of Locomotor Behavior

Defining the contribution of acetylcholine to specific behaviors has been challenging, mainly because of the difficulty in generating suitable animal models of cholinergic dysfunction. We have recently shown that, by targeting the vesicular acetylcholine transporter (VAChT) gene, it is possible to generate genetically modified mice with cholinergic deficiency. Here we describe novel VAChT mutant lines. VAChT gene is embedded within the first intron of the choline acetyltransferase (ChAT) gene, which provides a unique arrangement and regulation for these two genes. We generated a VAChT allele that is flanked by loxP sequences and carries the resistance cassette placed in a ChAT intronic region (FloxNeo allele). We show that mice with the FloxNeo allele exhibit differential VAChT expression in distinct neuronal populations. These mice show relatively intact VAChT expression in somatomotor cholinergic neurons, but pronounced decrease in other cholinergic neurons in the brain. VAChT mutant mice present preserved neuromuscular function, but altered brain cholinergic function and are hyperactive. Genetic removal of the resistance cassette rescues VAChT expression and the hyperactivity phenotype. These results suggest that release of ACh in the brain is normally required to “turn down” neuronal circuits controlling locomotion

Kaposi's Sarcoma Herpesvirus Upregulates Aurora A Expression to Promote p53 Phosphorylation and Ubiquitylation

Aberrant expression of Aurora A kinase has been frequently implicated in many cancers and contributes to chromosome instability and phosphorylation-mediated ubiquitylation and degradation of p53 for tumorigenesis. Previous studies showed that p53 is degraded by Kaposi's sarcoma herpesvirus (KSHV) encoded latency-associated nuclear antigen (LANA) through its SOCS-box (suppressor of cytokine signaling, LANASOCS) motif-mediated recruitment of the EC5S ubiquitin complex. Here we demonstrate that Aurora A transcriptional expression is upregulated by LANA and markedly elevated in both Kaposi's sarcoma tissue and human primary cells infected with KSHV. Moreover, reintroduction of Aurora A dramatically enhances the binding affinity of p53 with LANA and LANASOCS-mediated ubiquitylation of p53 which requires phosphorylation on Ser215 and Ser315. Small hairpin RNA or a dominant negative mutant of Aurora A kinase efficiently disrupts LANA-induced p53 ubiquitylation and degradation, and leads to induction of p53 transcriptional and apoptotic activities. These studies provide new insights into the mechanisms by which LANA can upregulate expression of a cellular oncogene and simultaneously destabilize the activities of the p53 tumor suppressor in KSHV-associated human cancers