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Melampsora larici-populina (Mlp), the causal agent of Populus leaf rust, secretes an array of effectors into the host through the haustorium to gain nutrients and suppress immunity. The precise mechanisms by which these effectors promote virulence remain unclear. To address this question, we developed a transgenic Arabidopsis line expressing a candidate effector, Mlp124357. Constitutive expression of the effector increased plant susceptibility to pathogens. A GxxxG motif present in Mlp124357 is required for its subcellular localization at the vacuolar membrane of the plant cell, as replacement of the glycine residues with alanines led to the delocalization of Mlp124357 to the nucleus and cytoplasm. We used immunoprecipitation and mass spectrometry (MS) to identify Mlp124357 interaction partners. Only one of the putative interaction partners knock-out line caused delocalization of the effector, indicating that Arabidopsis protein disulfide isomerase-11 (AtPDI-11) is required for the effector localization. This interaction was further confirmed by a complementation test, a yeast-two hybrid assay and a molecular modeling experiment. Moreover, localization results and infection assays suggest that AtPDI-11 act as a helper for Mlp124357. In summary, our findings established that one of Mlp effectors resides at the vacuole surface and modulates plant susceptibility.

The aim of this work was to demonstrate the first proof-of-concept for the use of ab initio -aided assembly strategy intensifying in vivo biosynthesis process to construct Escherichia coli cell factory overproducing highly active xanthine dehydrogenase (XDH). Three global regulator (IscS, TusA and NarJ) and four chaperone proteins (DsbA, DsbB, NifS and XdhC) were overexpressed to aid the formation and ordered assembly of three redox center cofactors of Rhodobacter capsulatus XDH in E. coli . The NifS, IscS and DsbB enhanced the specific activity of Rc XDH by 30%, 94% and 49%, respectively. The combinatorial expression of NarJ and IscS synergistically increased the specific activity by 129% and enhanced the total enzyme activity by a remarkable 3.9-fold. The crude enzyme showed nearly the same coupling efficiency of electron transfer and product formation as previously purified XDHs, indicating an integrity and efficient assembly of highly active XDH. [ABSTRACT FROM AUTHOR]

Bacillus thuringiensis (Bt) is the most successful, environmentally-friendly, and intensively studied microbial insecticide. The major characteristic of Bt is the production of proteinaceous crystals containing toxins with specific activity against many pests including dipteran, lepidopteran, and coleopteran insects, as well as nematodes, protozoa, flukes, and mites. These crystals allow large quantities of the protein toxins to remain stable in the environment until ingested by a susceptible host. It has been previously established that 135 kDa Cry proteins have a crystallization domain at their C-terminal end. In the absence of this domain, Cry proteins often need helper proteins or other factors for crystallization. In this review, we classify the Cry proteins based on their requirements for crystallization. [ABSTRACT FROM AUTHOR]

Purpose: Targeted immunotherapy using dendritic cells (DCs) has been employed in numerous investigations aiming at combating neoplasms. We previously showed that copulsing of an antigen with a helper protein could considerably enhance antigen presenting capacity of ex vivo-generated DCs. In this study, we attempted to administer an effective treatment in a murine model of colon cancer with DCs pulsed with the mixture of a tumor-specific gp70-derived peptide (AH1) and a helper protein, ovalbumin (OVA). Materials and Methods: First, the presence of gp70 in CT26 tumor cells and tumor tissues was verified using immunofluorescence and Western blot analyses. Next, DCs were purified from normal mice, loaded ex vivowith AH1 and OVA (DC-Pep-OVA), and injected into tumor-bearing mice. Tumor volume, in vitro antigen (Ag)-specific proliferation of splenic cells, and survival rate were measured to determine the efficacy of DC-Pep-OVA. As the control groups, tumor-bearing mice were vaccinated with DC-Pep, unpulsed DC, and DCs loaded with a mixture of OVA and an irrelevant peptide (P15), or were not vaccinated at all. Results: DC-Pep-OVA showed superior efficacy over other groups, as indicated by smaller tumor volume, higher Ag-specific proliferation rate of splenic cells, and prolonged survival. Conclusion: Overall, in the present study we showed for the first time that DCs copulsed with AH1 (tumor Ag) and OVA (helper molecule) could be considered as potentially robust weapons for use in future antitumor immunotherapies. [ABSTRACT FROM AUTHOR]