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The effects of chronic stress on learning are highly variable across individuals. This variability stems from gene–environment interactions. However, the mechanisms by which stress affects genetic predictors of learning are unclear. Thus, we aim to determine whether the genetic pathways that predict spatial memory performance are altered by previous exposure to chronic stress. Sixty-two BXD recombinant inbred strains of mice, as well as parent strains C57BL/6J and DBA/2J, were randomly assigned as behavioral control or to a chronic variable stress paradigm and then underwent behavioral testing to assess spatial memory and learning performance using the Morris water maze. Quantitative trait loci (QTL) mapping was completed for average escape latency times for both control and stress animals. Loci on chromosomes 5 and 10 were found in both control and stress environmental populations; eight additional loci were found to be unique to either the control or stress environment. In sum, results indicate that certain genetic loci predict spatial memory performance regardless of prior stress exposure, while exposure to stress also reveals unique genetic predictors of training during the memory task. Thus, we find that genetic predictors contributing to spatial learning and memory are susceptible to the presence of chronic stress. [ABSTRACT FROM AUTHOR]

The enzymatic A1 chain of cholera toxin retrotranslocates across the endoplasmic reticulum membrane into the cytosol, where it induces toxicity. Almost all other retrotranslocation substrates are modified by the attachment of polyubiquitin chains and moved into the cytosol by the ubiquitin-interacting p97 ATPase complex. The cholera toxin A1 chain, however, can induce toxicity in the absence of ubiquitination, and the motive force that drives retrotranslocation is not known. Here, we use adenovirus expressing dominant-negative mutants of p97 to test whether p97 is required for toxin action. We find that cholera toxin still functions with only a small decrease in potency in cells that cannot retrotranslocate other substrates at all. These results suggest that p97 does not provide the primary driving force for extracting the A1 chain from the endoplasmic reticulum, a finding that is consistent with a requirement for polyubiquitination in p97 function. [ABSTRACT FROM AUTHOR]

ONC201/TIC10 is a first-in-class small molecule inducer of TRAIL that causes early activation of the integrated stress response. Its promising safety profile and broad-spectrum efficacy in vitro have been confirmed in Phase I/II trials in several advanced malignancies. Binding and reporter assays have shown that ONC201 is a selective antagonist of the dopamine D2-like receptors, specifically, DRD2 and DRD3. We hypothesized that ONC201's interaction with DRD2 plays a role in ONC201's anticancer effects. Using cBioportal and quantitative reversetranscription polymerase chain reaction analyses, we confirmed that DRD2 is expressed in different cancer cell types in a cell type-specific manner. On the other hand, DRD3 was generally not detectable. Overexpressing DRD2 in cells with low DRD2 levels increased ONC201-induced PARP cleavage, which was preceded and correlated with an increase in ONC201-induced CHOP mRNA expression. On the other hand, knocking out DRD2 using CRISPR/Cas9 in three cancer cell lines was not sufficient to abrogate ONC201's anticancer effects. Although ONC201's anticancer activity was not dependent on DRD2 expression in the cancer cell types tested, we assessed the cytotoxic potential of DRD2 blockade. Transient DRD2 knockdown in HCT116 cells activated the integrated stress response and reduced cell number. Pharmacological antagonism of DRD2 significantly reduced cell viability. Thus, we demonstrate in this study that disrupting dopamine receptor expression and activity can have cytotoxic effects that may at least be in part due to the activation of the integrated stress response. On the other hand, ONC201's anticancer activity goes beyond its ability to antagonize DRD2, potentially due to ONC201's ability to activate other pathways that are independent of DRD2. Nevertheless, blocking the dopamine D1-like receptor DRD5 via siRNA or the use of a pharmacological antagonist promoted ONC201-induced anticancer activity. [ABSTRACT FROM AUTHOR]

Cholera toxin causes diarrheal disease by binding ganglioside GM1 on the apical membrane of polarized intestinal epithelial cells and trafficking retrograde through sorting endosomes, the trans-Golgi network (TGN), and into the endoplasmic reticu-lum. A fraction of toxin also moves from endosomes across the cell to the basolateral plasma membrane by transcytosis, thus breeching the intestinal barrier. Here we find that sorting of cholera toxin into this transcytotic pathway bypasses retrograde transport to the TGN. We also find that GM1 sphingolipids can traffic from apical to basolateral membranes by transcytosis in the absence of toxin binding but only if the GM1 species contain cis-unsaturated or short acyl chains in the ceramide domain. We found previously that the same GM1 species are needed to efficiently traffic retrograde into the TGN and endoplasmic reticulum and into the recycling endosome, implicating a shared mechanism of action for sorting by lipid shape among these pathways. [ABSTRACT FROM AUTHOR]

Summary: The glycosphingolipid GM1 binds cholera toxin (CT) on host cells and carries it retrograde from the plasma membrane (PM) through endosomes, the trans-Golgi (TGN), and the endoplasmic reticulum (ER) to induce toxicity. To elucidate how a membrane lipid can specify trafficking in these pathways, we synthesized GM1 isoforms with alternate ceramide domains and imaged their trafficking in live cells. Only GM1 with unsaturated acyl chains sorted efficiently from PM to TGN and ER. Toxin binding, which effectively crosslinks GM1 lipids, was dispensable, but membrane cholesterol and the lipid raft-associated proteins actin and flotillin were required. The results implicate a protein-dependent mechanism of lipid sorting by ceramide structure and provide a molecular explanation for the diversity and specificity of retrograde trafficking by CT in host cells. [Copyright &y& Elsevier]

Abstract: Proteosome-based vaccines have TLR2-based adjuvant activity and show promise for mucosal immunization. We examined the effects of proteosomes on mucosal uptake in Peyer''s patches in vivo. Proteosomes accelerated transepithelial transport of microparticles by M cells and induced migration of dendritic cells (DCs) into the follicle-associated epithelium (FAE); both effects were dependent on TLR2. Proteosomes induced the release of the DC-attracting chemokine MIP3α from Caco-2 epithelial cells in vitro. In HEK cells, proteosome-mediated MIP3α release was dependent on TLR2 expression and matrix metalloproteinase activation. Thus, TLR2 activation by proteosomes may promote mucosal uptake of particulate vaccines, and this may contribute to their adjuvanticity. [Copyright &y& Elsevier]