The neighbouring genes <i>AvrLm10A</i> and <i>AvrLm10B</i> are part of a large multigene family of cooperating effector genes conserved in Dothideomycetes and Sordariomycetes

Fungal effectors (small-secreted proteins) have long been considered as species or even subpopulation-specific. The increasing availability of high-quality fungal genomes and annotations has allowed the identification of trans-species or trans-genera families of effectors. Two avirulence effectors, AvrLm10A and AvrLm10B, of Leptosphaeria maculans, the fungus causing stem canker of oilseed rape, are members of such a large family of effectors. AvrLm10A and AvrLm10B are neighbouring genes, organized in divergent transcriptional orientation. Sequence searches within the L. maculans genome showed that AvrLm10A/AvrLm10B belong to a multigene family comprising five pairs of genes with a similar tail-to-tail organization. The two genes, in a pair, always had the same expression pattern and two expression profiles were distinguished, associated with the biotrophic colonization of cotyledons and/or petioles and stems. Of the two protein pairs further investigated, AvrLm10A_like1/AvrLm10B_like1 and AvrLm10A_like2/AvrLm10B_like2, the second one had the ability to physically interact, similarly to what was previously described for the AvrLm10A/AvrLm10B pair, and cross-interactions were also detected for two pairs. AvrLm10A homologues were identified in more than 30 Dothideomycete and Sordariomycete plant-pathogenic fungi. One of them, SIX5, is an effector from Fusarium oxysporum f. sp. lycopersici physically interacting with the avirulence effector Avr2. We found that AvrLm10A/SIX5 homologues were associated with at least eight distinct putative effector families, suggesting that AvrLm10A/SIX5 is able to cooperate with different effectors. These results point to a general role of the AvrLm10A/SIX5 proteins as “cooperating proteins”, able to interact with diverse families of effectors whose encoding gene is co-regulated with the neighbouring AvrLm10A homologue

Antiproliferative activity and mode of action analysis of novel amino and amido substituted phenantrene and naphtho[2,1-b]thiophene derivatives

Herein we present and describe the design and synthesis of novel phenantrene derivatives substituted with either amino or amido side chains and their biological activity. Antiproliferative activities were assessed in vitro on a panel of human cancer cell lines. Tested compounds showed moderate activity against cancer cells in comparison with 5-fluorouracile. Among all tested compounds, some compounds substituted with cyano groups showed a pronounced and selective activity in the nanomolar range of inhibitory concentrations against HeLa and HepG2. The strongest selective activity against HeLa cells was observed for acrylonitriles 8 and 11 and their cyclic analogues 15 and 17 substituted with two cyano groups with a corresponding IC50 = 0.33, 0.21, 0.65 and 0.45 μM, respectively. Compounds 11 showed the most pronounced selectivity being almost non cytotoxic to normal fibroblasts. Additionally, mode of biological action analysis was performed in silico and in vitro by Western blot analysis of HIF-1-α relative expression for compounds 8 and 11

Interfacial charge transfer in nanoscale polymer transistors

Interfacial charge transfer plays an essential role in establishing the relative alignment of the metal Fermi level and the energy bands of organic semiconductors. While the details remain elusive in many systems, this charge transfer has been inferred in a number of photoemission experiments. We present electronic transport measurements in very short channel ($L < 100$ nm) transistors made from poly(3-hexylthiophene) (P3HT). As channel length is reduced, the evolution of the contact resistance and the zero-gate-voltage conductance are consistent with such charge transfer. Short channel conduction in devices with Pt contacts is greatly enhanced compared to analogous devices with Au contacts, consistent with charge transfer expectations. Alternating current scanning tunneling microscopy (ACSTM) provides further evidence that holes are transferred from Pt into P3HT, while much less charge transfer takes place at the Au/P3HT interface.Comment: 19 preprint pages, 6 figure

Transition Metal-Mediated Hydrolysis of the Imine Bond in 2-Azomethine Benzothiazoles

An ultrasound-assisted synthesis has been used for the preparation of novel benzothiazole Schiff bases (1–3) as ligands for Zn(II) and Cu(II) metal complexes. In this class of Schiff base ligands, the isolated complexes 1a and 2a appear to be the only ones that have structures which include both the Schiff base ligand and the hydrolysis product 2-aminobenzothiazole in the same complex molecule. The aldehydes formed by Schiff base hydrolysis formed a rare example of a trans isomer in pentacoordinated 2, 4-disubstituted benzaldehyde complexes

Protein–protein interactions as a proxy to monitor conformational changes and activation states of the tomato resistance protein I-2

Plant resistance proteins (R) are involved in pathogen recognition and subsequent initiation of defence responses. Their activity is regulated by inter- and intramolecular interactions. In a yeast two-hybrid screen two clones (I2I-1 and I2I-2) specifically interacting with I-2, a Fusarium oxysporum f. sp. lycopersici resistance protein of the CC-NB-LRR family, were identified. Sequence analysis revealed that I2I-1 belongs to the Formin gene family (SlFormin) whereas I2I-2 has homology to translin-associated protein X (SlTrax). SlFormin required only the N-terminal CC I-2 domain for binding, whereas SlTrax required both I-2 CC and part of the NB-ARC domain. Tomato plants stably silenced for these interactors were not compromised in I-2-mediated disease resistance. When extended or mutated forms of I-2 were used as baits, distinct and often opposite, interaction patterns with the two interactors were observed. These interaction patterns correlated with the proposed activation state of I-2 implying that active and inactive R proteins adopt distinct conformations. It is concluded that the yeast two hybrid system can be used as a proxy to monitor these different conformational states

Evolution of Linked Avirulence Effectors in Leptosphaeria maculans Is Affected by Genomic Environment and Exposure to Resistance Genes in Host Plants

Brassica napus (canola) cultivars and isolates of the blackleg fungus, Leptosphaeria maculans interact in a ‘gene for gene’ manner whereby plant resistance (R) genes are complementary to pathogen avirulence (Avr) genes. Avirulence genes encode proteins that belong to a class of pathogen molecules known as effectors, which includes small secreted proteins that play a role in disease. In Australia in 2003 canola cultivars with the Rlm1 resistance gene suffered a breakdown of disease resistance, resulting in severe yield losses. This was associated with a large increase in the frequency of virulence alleles of the complementary avirulence gene, AvrLm1, in fungal populations. Surprisingly, the frequency of virulence alleles of AvrLm6 (complementary to Rlm6) also increased dramatically, even though the cultivars did not contain Rlm6. In the L. maculans genome, AvrLm1 and AvrLm6 are linked along with five other genes in a region interspersed with transposable elements that have been degenerated by Repeat-Induced Point (RIP) mutations. Analyses of 295 Australian isolates showed deletions, RIP mutations and/or non-RIP derived amino acid substitutions in the predicted proteins encoded by these seven genes. The degree of RIP mutations within single copy sequences in this region was proportional to their proximity to the degenerated transposable elements. The RIP alleles were monophyletic and were present only in isolates collected after resistance conferred by Rlm1 broke down, whereas deletion alleles belonged to several polyphyletic lineages and were present before and after the resistance breakdown. Thus, genomic environment and exposure to resistance genes in B. napus has affected the evolution of these linked avirulence genes in L. maculans

The Nuclear Protein Sge1 of Fusarium oxysporum Is Required for Parasitic Growth

Dimorphism or morphogenic conversion is exploited by several pathogenic fungi and is required for tissue invasion and/or survival in the host. We have identified a homolog of a master regulator of this morphological switch in the plant pathogenic fungus Fusarium oxysporum f. sp. lycopersici. This non-dimorphic fungus causes vascular wilt disease in tomato by penetrating the plant roots and colonizing the vascular tissue. Gene knock-out and complementation studies established that the gene for this putative regulator, SGE1 (SIX Gene Expression 1), is essential for pathogenicity. In addition, microscopic analysis using fluorescent proteins revealed that Sge1 is localized in the nucleus, is not required for root colonization and penetration, but is required for parasitic growth. Furthermore, Sge1 is required for expression of genes encoding effectors that are secreted during infection. We propose that Sge1 is required in F. oxysporum and other non-dimorphic (plant) pathogenic fungi for parasitic growth

CaZF, a Plant Transcription Factor Functions through and Parallel to HOG and Calcineurin Pathways in Saccharomyces cerevisiae to Provide Osmotolerance

Salt-sensitive yeast mutants were deployed to characterize a gene encoding a C2H2 zinc finger protein (CaZF) that is differentially expressed in a drought-tolerant variety of chickpea (Cicer arietinum) and provides salinity-tolerance in transgenic tobacco. In Saccharomyces cerevisiae most of the cellular responses to hyper-osmotic stress is regulated by two interconnected pathways involving high osmolarity glycerol mitogen-activated protein kinase (Hog1p) and Calcineurin (CAN), a Ca2+/calmodulin-regulated protein phosphatase 2B. In this study, we report that heterologous expression of CaZF provides osmotolerance in S. cerevisiae through Hog1p and Calcineurin dependent as well as independent pathways. CaZF partially suppresses salt-hypersensitive phenotypes of hog1, can and hog1can mutants and in conjunction, stimulates HOG and CAN pathway genes with subsequent accumulation of glycerol in absence of Hog1p and CAN. CaZF directly binds to stress response element (STRE) to activate STRE-containing promoter in yeast. Transactivation and salt tolerance assays of CaZF deletion mutants showed that other than the transactivation domain a C-terminal domain composed of acidic and basic amino acids is also required for its function. Altogether, results from this study suggests that CaZF is a potential plant salt-tolerance determinant and also provide evidence that in budding yeast expression of HOG and CAN pathway genes can be stimulated in absence of their regulatory enzymes to provide osmotolerance

Integrating Phosphorylation Network with Transcriptional Network Reveals Novel Functional Relationships

Phosphorylation and transcriptional regulation events are critical for cells to transmit and respond to signals. In spite of its importance, systems-level strategies that couple these two networks have yet to be presented. Here we introduce a novel approach that integrates the physical and functional aspects of phosphorylation network together with the transcription network in S.cerevisiae, and demonstrate that different network motifs are involved in these networks, which should be considered in interpreting and integrating large scale datasets. Based on this understanding, we introduce a HeRS score (hetero-regulatory similarity score) to systematically characterize the functional relevance of kinase/phosphatase involvement with transcription factor, and present an algorithm that predicts hetero-regulatory modules. When extended to signaling network, this approach confirmed the structure and cross talk of MAPK pathways, inferred a novel functional transcription factor Sok2 in high osmolarity glycerol pathway, and explained the mechanism of reduced mating efficiency upon Fus3 deletion. This strategy is applicable to other organisms as large-scale datasets become available, providing a means to identify the functional relationships between kinases/phosphatases and transcription factors