PubMed

Cranial irradiation in juvenile mice leads to early and sustained defects in the stem and progenitor cell pools and late cognitive impairments.

Brain Res. 2019 Nov 09;:146548

Authors: Ruddy RM, Derkach D, Dadwal P, Morshead CM

Abstract
Cranial irradiation is used in combination with other therapies as a treatment for brain tumours and is thought to contribute to long-term cognitive deficits. Several rodent models have demonstrated that these cognitive deficits may be correlated with damage to neural progenitor cells in the subventricular zone (SVZ) and dentate gyrus (DG), the two neurogenic niches of the brain. Studies in rodent models typically assess the proliferating progenitor population, but rarely investigate the effect of cranial irradiation on the neural stem cell pool. Further, few studies evaluate the effects in juveniles, an age when children typically receive this treatment. Herein, we examine the cellular and behavioural effects of juvenile cranial irradiation on stem and progenitor populations in the two neurogenic regions of the brain and assess cognitive outcomes. We found regionally distinct effects of cranial irradiation in the juvenile brain. In the SVZ, we observed a defect in the stem cell pool and a concomitant decrease in proliferating cells that were maintained for at least one week. In the DG, a similar defect in the stem cell pool and proliferating cells was observed and persisted in the stem cell population. Finally, we demonstrated that cranial irradiation resulted in late cognitive deficits. This study demonstrates that juvenile cranial irradiation leads to regionally distinct defects in the stem and progenitor populations, and late cognitive deficits, which may be important factors in determining therapeutic targets and timing of interventions following cranial irradiation.

PMID: 31715143 [PubMed - as supplied by publisher]

A potent anti-SpuE antibody allosterically inhibits type III secretion system and attenuates virulence of Pseudomonas aeruginosa.

J Mol Biol. 2019 Nov 01;:

Authors: Zhang Y, Sun X, Qian Y, Yi H, Song K, Zhu H, Zonta F, Chen W, Ji Q, Miersch S, Sidhu SS, Wu D

Abstract
Multidrug-resistant Gram-negative bacteria infection is particularly severe within the designated ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species), which underscores the urgent need to explore alternative therapeutic strategies. The type III secretion system (T3SS) is considered to be a key virulence factor in many Gram-negative bacteria, and T3SS is in turn regulated by SpuE in P. aeruginosa, which is a spermidine binding protein from an ATP-binding cassette transporter family and highly conserved within ESKAPE pathogens. Here, we identified a potent anti-SpuE antagonistic antibody that allosterically inhibits the expression of T3SS and attenuates virulence of P. aeruginosa. X-ray crystallography and molecular dynamics simulations revealed that binding of antibody to SpuE induces a change in the dynamics of SpuE, which in turn may reduce spermidine uptake by P. aeruginosa. The antibody could serve as a template for developing novel biologics to target a broad spectrum of Gram-negative bacteria.

PMID: 31682834 [PubMed - as supplied by publisher]

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Highly Combinatorial Genetic Interaction Analysis Reveals a Multi-Drug Transporter Influence Network.

Cell Syst. 2019 Oct 14;:

Authors: Celaj A, Gebbia M, Musa L, Cote AG, Snider J, Wong V, Ko M, Fong T, Bansal P, Mellor JC, Seesankar G, Nguyen M, Zhou S, Wang L, Kishore N, Stagljar I, Suzuki Y, Yachie N, Roth FP

Abstract
Many traits are complex, depending non-additively on variant combinations. Even in model systems, such as the yeast S. cerevisiae, carrying out the high-order variant-combination testing needed to dissect complex traits remains a daunting challenge. Here, we describe "X-gene" genetic analysis (XGA), a strategy for engineering and profiling highly combinatorial gene perturbations. We demonstrate XGA on yeast ABC transporters by engineering 5,353 strains, each deleted for a random subset of 16 transporters, and profiling each strain's resistance to 16 compounds. XGA yielded 85,648 genotype-to-resistance observations, revealing high-order genetic interactions for 13 of the 16 transporters studied. Neural networks yielded intuitive functional models and guided exploration of fluconazole resistance, which was influenced non-additively by five genes. Together, our results showed that highly combinatorial genetic perturbation can functionally dissect complex traits, supporting pursuit of analogous strategies in human cells and other model systems.

PMID: 31668799 [PubMed - as supplied by publisher]

Comprehensive analysis of all evolutionary paths between two divergent PDZ domain specificities.

Protein Sci. 2019 Oct 25;:

Authors: Teyra J, Ernst A, Singer A, Sicheri F, Sidhu SS

Abstract
To understand the molecular evolution of functional diversity in protein families, we comprehensively investigated the consequences of all possible mutation combinations separating two peptide-binding domains with highly divergent specificities. We analyzed the Erbin PDZ domain (Erbin-PDZ), which exhibits canonical type I specificity, and a synthetic Erbin-PDZ variant (E-14) that differs at six positions and exhibits an atypical specificity that closely resembles that of the natural Pdlim4 PDZ domain (Pdlim4-PDZ). We constructed a panel of 64 PDZ domains covering all possible transitions between Erbin-PDZ and E-14 (i.e. the panel contained variants with all possible combinations of either the Erbin-PDZ or E-14 sequence at the six differing positions). We assessed the specificity profiles of the 64 PDZ domains using a C-terminal phage-displayed peptide library containing all possible genetically encoded heptapeptides. The specificity profiles clustered into six distinct groups, showing that intermediate domains can be nodes for the evolution of divergent functions. Remarkably, three substitutions were sufficient to convert the specificity of Erbin-PDZ to that of Pdlim4-PDZ, whereas Pdlim4-PDZ contains 71 differences relative to Erbin-PDZ. X-ray crystallography revealed the structural basis for specificity transition: a single substitution in the center of the binding site, supported by contributions from auxiliary substitutions, altered the main chain conformation of the peptide ligand to resemble that of ligands bound to Pdlim4-PDZ. Our results show that a very small set of mutations can dramatically alter protein specificity, and these findings support the hypothesis whereby complex protein functions evolve by gene duplication followed by cumulative mutations.

PMID: 31654425 [PubMed - as supplied by publisher]