Structural insights into the function of type VI secretion system TssA subunits.

The type VI secretion system (T6SS) is a multi-protein complex that injects bacterial effector proteins into target cells. It is composed of a cell membrane complex anchored to a contractile bacteriophage tail-like apparatus consisting of a sharpened tube that is ejected by the contraction of a sheath against a baseplate. We present structural and biochemical studies on TssA subunits from two different T6SSs that reveal radically different quaternary structures in comparison to the dodecameric E. coli TssA that arise from differences in their C-terminal sequences. Despite this, the different TssAs retain equivalent interactions with other components of the complex and position their highly conserved N-terminal ImpA_N domain at the same radius from the centre of the sheath as a result of their distinct domain architectures, which includes additional spacer domains and highly mobile interdomain linkers. Together, these variations allow these distinct TssAs to perform a similar function in the complex

Expression of horseradish peroxidase in transgenic tobacco

Peroxidases are common oxidoreductases present in plants, animals and micro-organisms. Plant peroxidases are classified as basic, neutral or acidic enzymes, depending on their profiles of elution during ion-exchange column chromatography or isoelectric points [ 11. Horseradish (Amtoru~hru sticuna) peroxidases (HRPs; EC 1.1 1.1.7), are probably the best characterized of these enzymes. They are believed to be involved in plant responses to a number of biotic and abiotic stresses, notably in the disruption of free radicals and active derivatives of oxygen. The effects of these molecules have been well described [2], and in general the presence of these compounds are ‘highly destructive’ for cell metabolism. However, these oxygen derivatives are involved in halogenation and hydroxylation reactions in peroxisomes and in the cell wall, suggesting that the plant cell needs a fine regulation between production and destruction of oxygen radicals. A key to the comprehension of the role of peroxidase activity in this and other stress responses could be the study of the effect of modified expression and targeting of peroxidase activity in transgenic plants. The present report describes the production of such tobacco plants using a synthetic HRP gene [ 31, with a range of regulatory and targeting sequences

Direct gene transfer in potato: A comparison of particle bombardment of leaf explants and PEG-mediated transformation of protoplasts

Direct gene transfer methods in potato would facilitate the transfer of multiple genes and the manipulation of metabolic pathways in this species. In this study, up to 1.8 transformation events per shot (=0.5 per bombarded leaf) and 67.2 events per million protoplasts treated were obtained with particle bombardment and PEG-mediated direct DNA uptake, respectively. Limited disassociation of both HPT and GUS genes appeared to occur during the process of integration in only 19% of transformants. A large number of transformed potato plants with transgene expression at levels comparable to Agrobacterium-mediated transformation was obtained. High levels of GUS expression were only obtained in lines derived from PEG treatment. No correlation between the number of gene insertions and gene expression levels was found, suggesting that multiple insertions may have little or no effect on transgene expression