Association between genetic variants of the cholinergic system and postoperative delirium and cognitive dysfunction in elderly patients.

BACKGROUND: Postoperative delirium (POD) and postoperative cognitive dysfunction (POCD) are frequent and serious complications after surgery. We aim to investigate the association between genetic variants in cholinergic candidate genes according to the Kyoto encyclopedia of genes and genomes - pathway: cholinergic neurotransmission with the development of POD or POCD in elderly patients. METHODS: This analysis is part of the European BioCog project ( www.biocog.eu ), a prospective multicenter observational study with elderly surgical patients. Patients with a Mini-Mental-State-Examination score ≤ 23 points were excluded. POD was assessed up to seven days after surgery using the Nursing Delirium Screening Scale, Confusion Assessment Method and a patient chart review. POCD was assessed three months after surgery with a neuropsychological test battery. Genotyping was performed on the Illumina Infinium Global Screening Array. Associations with POD and POCD were analyzed using logistic regression analysis, adjusted for age, comorbidities and duration of anesthesia (for POCD analysis additionally for education). Odds ratios (OR) refer to minor allele counts (0, 1, 2). RESULTS: 745 patients could be included in the POD analysis, and 452 in the POCD analysis. The rate of POD within this group was 20.8% (155 patients), and the rate of POCD was 10.2% (46 patients). In a candidate gene approach three genetic variants of the cholinergic genes CHRM2 and CHRM4 were associated with POD (OR [95% confidence interval], rs8191992: 0.61[0.46; 0.80]; rs8191992: 1.60[1.22; 2.09]; rs2067482: 1.64[1.10; 2.44]). No associations were found for POCD. CONCLUSIONS: We found an association between genetic variants of CHRM2 and CHRM4 and POD. Further studies are needed to investigate whether disturbances in acetylcholine release and synaptic plasticity are involved in the development of POD. TRIAL REGISTRATION: ClinicalTrials.gov: NCT02265263.

Targeting GSK3 from Ustilago maydis: type-II kinase inhibitors as potential antifungals.

Protein kinases are key enzymes in the complex regulation of cellular processes in almost all living organisms. For this reason, protein kinases represent attractive targets to stop the growth of eukaryotic pathogens such as protozoa and fungi. However, using kinase inhibitors to fight against these organisms bears several challenges since most of them are unselective and will also affect crucial host kinases. Here we present the X-ray structure of glycogen synthase kinase 3 from the fungal plant pathogen Ustilago maydis (UmGSK3) and its inhibition by type-II kinase inhibitors. Despite the high sequence homology between the human and the fungal variant of this vital kinase, we found substantial differences in the conformational plasticity of their active sites. Compounds that induced such conformational changes could be used to selectively inhibit the fungal kinase. This study serves as an example of how species-specific selectivity of inhibitors can be achieved by identifying and addressing the inactive state of a protein kinase. In addition to this, our study gives interesting insights into the molecular plasticity of UmGSK3 by revealing a previously unknown inactive conformation of this important kinase family.

Identification of Ustilago maydis Aurora kinase as a novel antifungal target.

Infestation of crops by pathogenic fungi has continued to have a major impact by reducing yield and quality, emphasizing the need to identify new targets and develop new agents to improve methods of crop protection. Here we present Aurora kinase from the phytopathogenic fungus Ustilago maydis as a novel target for N-substituted diaminopyrimidines, a class of small-molecule kinase inhibitors. We show that Aurora kinase is essential in U. maydis and that diaminopyrimidines inhibit its activity in vitro. Furthermore, we observed an overall good correlation between in vitro inhibition of Aurora kinase and growth inhibition of diverse fungi in vivo. In vitro inhibition assays with Ustilago and human Aurora kinases indicate that some compounds of the N-substituted diaminopyrimidine class show specificity for the Ustilago enzyme, thus revealing their potential as selective fungicides.

Identification and characterization of secreted and pathogenesis-related proteins in Ustilago maydis.

Interactions between plants and fungal pathogens require a complex interplay at the plant-fungus interface. Extracellular effector proteins are thought to play a crucial role in establishing a successful infection. To identify pathogenesis-related proteins in Ustilago maydis we combined the isolation of secreted proteins using a signal sequence trap approach with bioinformatic analyses and the subsequent characterization of knock-out mutants. We identified 29 secreted proteins including hydrophobins and proteins with a repetitive structure similar to the repellent protein Rep1. Hum3, a protein containing both, a hydrophobin domain and a repetitive Rep1-like region, is shown to be processed during passage through the secretory pathway. While single knock-outs of hydrophobin or repellent-like genes did not affect pathogenicity, we found a strong effect of a double knock-out of hum3 and the repetitive rsp1. Yeast-like growth, mating, aerial hyphae formation and surface hydrophobicity were unaffected in this double mutant. However, pathogenic development in planta stops early after penetration leading to a complete loss of pathogenicity. This indicates that Hum3 and Rsp1 are pathogenicity proteins that share an essential function in early stages of the infection. Our results demonstrate that focusing on secreted proteins is a promising way to discover novel pathogenicity proteins that might be broadly applied to a variety of fungal pathogens.

Peptide-mediated broad-spectrum plant resistance to tospoviruses.

Plant viruses have a significant impact on agronomic losses worldwide. A new strategy for engineering virus-resistant plants by transgenic expression of a dominant interfering peptide is presented here. This peptide of 29 aa strongly interacts with the nucleocapsid proteins (N) of different tospoviruses. Transgenic Nicotiana benthamiana lines expressing the peptide fused to a carrier protein were challenged with five different tospoviruses that have a nucleocapsid protein interacting with the peptide. In the transgenic plants, strong resistance to tomato spotted wilt virus, tomato chlorotic spot virus, groundnut ring spot virus, and chrysanthemum stem necrosis virus was observed. This therefore demonstrates the feasibility of using peptide "aptamers" as an in vivo tool to control viral infection in higher plants.