Pectin methyl esterases and rhamnogalacturonan hydrolases: weapons for successful Monilinia laxa infection in stone fruit?

The secretion of cell wall‐degrading enzymes is one of the mechanisms used by necrotrophic fungi to colonize host tissues. However, information about virulence factors of Monilinia spp., the causal agents of brown rot in stone fruit, is scarce. Plant cell walls have three main components that are broken down by fungal enzymes: cellulose, hemicellulose and pectin. In order to identify Monilinia laxa candidate proteins involved in pectin hydrolysis, two in vitro approaches were conducted: (i) phenotypic and ecophysiological characterization of growth of the pathogen at different pHs, in glucose‐ and pectin‐containing solid media for 7 days' incubation; and (ii) expression analysis of genes encoding M. laxa pectin methyl esterases (MlPMEs) and rhamnogalacturonan hydrolases (MlRG‐HYDs) after incubation for 0.5, 2, 6, 24 and 48 h in glucose‐ and pectin‐containing liquid media. Phenotypic tests showed the role of carbon source on M. laxa growth rate and aggressiveness, and indicated that pectinases were greatly affected by pH. Gene expression analyses uncovered differences among members of each family of pectinases and between the two families, defining sets of genes expressed at earlier (0.5–6 h) and later (48 h) phases. Notably, the up‐ or down‐regulation of these target genes was carbon source‐dependent. Finally, an in vivo study confirmed the synergistic and complementary role that these genes play in the M. laxa–stone fruit pathosystem. Based on these results, it is hypothesized that MlPME2, MlRG‐HYD1 and MlRG‐HYD2 may be potential virulence factors of M. laxa in the process from infection to colonization.info:eu-repo/semantics/acceptedVersio

Histone deacetylase inhibitors promote glioma cell death by G2 checkpoint abrogation leading to mitotic catastrophe

Glioblastoma multiforme is resistant to conventional anti-tumoral treatments due to its infiltrative nature and capability of relapse; therefore, research efforts focus on characterizing gliomagenesis and identifying molecular targets useful on therapy. New therapeutic strategies are being tested in patients, such as Histone deacetylase inhibitors (HDACi) either alone or in combination with other therapies. Here two HDACi included in clinical trials have been tested, suberanilohydroxamic acid (SAHA) and valproic acid (VPA), to characterize their effects on glioma cell growth in vitro and to determine the molecular changes that promote cancer cell death. We found that both HDACi reduce glioma cell viability, proliferation and clonogenicity. They have multiple effects, such as inducing the production of reactive oxygen species (ROS) and activating the mitochondrial apoptotic pathway, nevertheless cell death is not prevented by the pan-caspase inhibitor Q-VD-OPh. Importantly, we found that HDACi alter cell cycle progression by decreasing the expression of G2 checkpoint kinases Wee1 and checkpoint kinase 1 (Chk1). In addition, HDACi reduce the expression of proteins involved in DNA repair (Rad51), mitotic spindle formation (TPX2) and chromosome segregation (Survivin) in glioma cells and in human glioblastoma multiforme primary cultures. Therefore, HDACi treatment causes glioma cell entry into mitosis before DNA damage could be repaired and to the formation of an aberrant mitotic spindle that results in glioma cell death through mitotic catastrophe-induced apoptosis