The Role of Pseudo-Orthocaspase (SyOC) of Synechocystis sp. PCC 6803 in Attenuating the Effect of Oxidative Stress.

Caspases are proteases, best known for their involvement in the execution of apoptosis-a subtype of programmed cell death, which occurs only in animals. These proteases are composed of two structural building blocks: a proteolytically active p20 domain and a regulatory p10 domain. Although structural homologs appear in representatives of all other organisms, their functional homology, i.e., cell death depending on their proteolytical activity, is still much disputed. Additionally, pseudo-caspases and pseudo-metacaspases, in which the catalytic histidine-cysteine dyad is substituted with non-proteolytic amino acid residues, were shown to be involved in cell death programs. Here, we present the involvement of a pseudo-orthocaspase (SyOC), a prokaryotic caspase-homolog lacking the p10 domain, in oxidative stress in the model cyanobacterium Synechocystis sp. PCC 6803. To study the in vivo impact of this pseudo-protease during oxidative stress its gene expression during exposure to H2O2 was monitored by RT-qPCR. Furthermore, a knock-out mutant lacking the pseudo-orthocaspase gene was designed, and its survival and growth rates were compared to wild type cells as well as its proteome. Deletion of SyOC led to cells with a higher tolerance toward oxidative stress, suggesting that this protein may be involved in a pro-death pathway.

Type III Secretion-Dependent and -Independent Phenotypes Caused by Ralstonia solanacearum in Arabidopsis Roots.

The causal agent of bacterial wilt, Ralstonia solanacearum, is a soilborne pathogen that invades plants through their roots, traversing many tissue layers until it reaches the xylem, where it multiplies and causes plant collapse. The effects of R. solanacearum infection are devastating, and no effective approach to fight the disease is so far available. The early steps of infection, essential for colonization, as well as the early plant defense responses remain mostly unknown. Here, we have set up a simple, in vitro Arabidopsis thaliana-R. solanacearum pathosystem that has allowed us to identify three clear root phenotypes specifically associated to the early stages of infection: root-growth inhibition, root-hair formation, and root-tip cell death. Using this method, we have been able to differentiate, on Arabidopsis plants, the phenotypes caused by mutants in the key bacterial virulence regulators hrpB and hrpG, which remained indistinguishable using the classical soil-drench inoculation pathogenicity assays. In addition, we have revealed the previously unknown involvement of auxins in the root rearrangements caused by R. solanacearum infection. Our system provides an easy-to-use, high-throughput tool to study R. solanacearum aggressiveness. Furthermore, the observed phenotypes may allow the identification of bacterial virulence determinants and could even be used to screen for novel forms of early plant resistance to bacterial wilt.

Dying two deaths - programmed cell death regulation in development and disease.

Programmed cell death (PCD) is a fundamental cellular process that has adopted a plethora of vital functions in multicellular organisms. In plants, PCD processes are elicited as an inherent part of regular development in specific cell types or tissues, but can also be triggered by biotic and abiotic stresses. Although over the last years we have seen progress in our understanding of the molecular regulation of different plant PCD processes, it is still unclear whether a common core machinery exists that controls cell death in development and disease. In this review, we discuss recent advances in the field, comparing some aspects of the molecular regulation controlling developmental and pathogen-triggered PCD in plants.

Detection and Quantification of Protein Aggregates in Plants.

Plants are constantly exposed to a complex and changing environment that challenges their cellular homeostasis. Stress responses triggered as a consequence of unfavorable conditions result in increased protein aggregate formation at the cellular level. When the formation of misfolded proteins surpasses the capacity of the cell to remove them, insoluble protein aggregates accumulate. In the animal field, an enormous effort is being placed to uncover the mechanisms regulating aggregate formation because of its implications in many important human diseases. Because of its importance for cellular functionality and fitness, it is equally important to expand plant research in this field. Here, we describe a cell fractionation-based method to obtain very pure insoluble protein aggregate fractions that can be subsequently semiquantified using image analysis. This method can be used as a first step to evaluate whether a particular condition results in an alteration of protein aggregate formation levels.