Secretagogues differentially activate endoplasmic reticulum stress responses in pancreatic acinar cells

Endoplasmic reticulum (ER) stress leads to the accumulation of misfolded proteins in the ER lumen and initiates the unfolded protein response (UPR). Components of the UPR are important in pancreatic development, and recent studies have indicated that the UPR is activated in the arginine model of acute pancreatitis. However, the effects of secretagogues on UPR components in the pancreas are unknown. The present study aimed to examine the effects of different types and concentrations of secretagogues on acinar cell function and specific components of the UPR. Rat pancreatic acini were stimulated with the CCK analogs CCK8 (10 pM-10 nM) or JMV-180 (10 nM-10 microM) or with bombesin (1-100 nM). Components of the UPR, including chaperone BiP expression, PKR-like ER kinase (PERK) phosphorylation, X box-binding protein 1 (XBP1) splicing, and CCAAT/enhancer binding protein homologous protein (CHOP) expression, were measured, as were effects on amylase secretion and intracellular trypsin activation. CCK8 generated a biphasic secretion dose-response curve, and high concentrations increased intracellular active trypsin levels. In contrast, JMV-180 and bombesin secretion dose-response curves were monophasic, and high concentrations did not increase intracellular trypsin activity. All three secretagogues increased BiP levels and XBP1 splicing. However, only supraphysiological levels of CCK8 associated with inhibited amylase secretion and trypsin activation stimulated PERK phosphorylation and expression of CHOP. The effects of CCK8 on UPR components were rapid, occurring within 5-20 min. In conclusion, ER stress response mechanisms appear to be involved in both pancreatic physiology and pathophysiology, and future efforts should be directed at understanding the roles of these mechanisms in the pancreas.     

The molecular origin and evolution of dim‐light vision in mammals

The nocturnal origin of mammals is a longstanding hypothesis that is considered instrumental for the evolution of endothermy, a potential key innovation in this successful clade. This hypothesis is primarily based on indirect anatomical inference from fossils. Here, we reconstruct the evolutionary history of rhodopsin—the vertebrate visual pigment mediating the first step in phototransduction at low‐light levels—via codon‐based model tests for selection, combined with gene resurrection methods that allow for the study of ancient proteins. Rhodopsin coding sequences were reconstructed for three key nodes: Amniota, Mammalia, and Theria. When expressed in vitro, all sequences generated stable visual pigments with λ_MAX values similar to the well‐studied bovine rhodopsin. Retinal release rates of mammalian and therian ancestral rhodopsins, measured via fluorescence spectroscopy, were significantly slower than those of the amniote ancestor, indicating altered molecular function possibly related to nocturnality. Positive selection along the therian branch suggests adaptive evolution in rhodopsin concurrent with therian ecological diversification events during the Mesozoic that allowed for an exploration of the environment at varying light levels.     

Toxoplasma gondii: analysis of the active site insertion of its ferredoxin-NADP(+)-reductase by peptide-specific antibodies and homology-based modeling

Apicomplexan parasites possess an apicoplast-localized redox system consisting of a plant-type ferredoxin-NADP(+)-reductase (FNR) and its redox partner ferredoxin, a small [2Fe-2S] protein. We show here that several apicomplexan FNRs contain unique amino acid insertions of various lengths which are located in close proximity to the enzymatically important FAD and ferredoxin-binding sites of these proteins. Using the insertion of the Toxoplasma gondii reductase as an example we raised epitope-specific antibodies against an 11 amino acids long peptide predicted to be surface-exposed within this insertion. This peptide was found to be immunogenic when presented to the immune system as part of a carrier protein, but also in its natural structural context in the whole recombinant protein, implying that the epitope is surface-exposed. Three-dimensional modeling of T. gondii FNR based on the known 3D-structure of maize root FNR predicts that the overall structure of plant and apicomplexan FNRs are very similar and that the 11 amino acids are part of an alpha-helix, looping out of the molecule. Collectively, these data suggest that the insertion in T. gondii FNR does not affect the overall structure of the protein but may have an effect on the binding dynamics of FAD, NADP(+), and/or ferredoxin to FNR.     

Non-Listening and Self Centered Leadership – Relationships to Socioeconomic Conditions and Employee Mental Health

Background

The way in which leadership is experienced in different socioeconomic strata is of interest per se, as well as how it relates to employee mental health.

Methods

Three waves of SLOSH (Swedish Longitudinal Occupational Survey of Health, a questionnaire survey on a sample of the Swedish working population) were used, 2006, 2008 and 2010 (n = 5141). The leadership variables were: “Non-listening leadership” (one question: “Does your manager listen to you?” - four response categories), “Self centered leadership” (sum of three five-graded questions – “non-participating”, “asocial” and “loner”). The socioeconomic factors were education and income. Emotional exhaustion and depressive symptoms were used as indicators of mental health.

Results

Non-listening leadership was associated with low income and low education whereas self-centered leadership showed a weaker relationship with education and no association at all with income. Both leadership variables were significantly associated with emotional exhaustion and depressive symptoms. “Self centered” as well as “non-listening” leadership in 2006 significantly predicted employee depressive symptoms in 2008 after adjustment for demographic variables. These predictions became non-significant when adjustment was made for job conditions (demands and decision latitude) in the “non-listening” leadership analyses, whereas predictions of depressive symptoms remained significant after these adjustments in the “self-centered leadership” analyses.

Conclusions

Our results show that the leadership variables are associated with socioeconomic status and employee mental health. “Non-listening” scores were more sensitive to societal change and more strongly related to socioeconomic factors and job conditions than “self-centered” scores.     

The Aspergillus nidulans kinesin-3 tail is necessary and sufficient to recognize modified microtubules

Posttranslational microtubule modifications (PTMs) are numerous; however, the biochemical and cell biological roles of those modifications remain mostly an enigma. The Aspergillus nidulans kinesin-3 UncA uses preferably modified microtubules (MTs) as tracks for vesicle transportation. Here, we show that a positively charged region in the tail of UncA (amino acids 1316 to 1402) is necessary for the recognition of modified MTs. Chimeric proteins composed of the kinesin-1 motor domain and the UncA tail displayed the same specificity as UncA, suggesting that the UncA tail is sufficient to establish specificity. Interaction between the UncA tail and alpha-tubulin was shown using a yeast two-hybrid assay and in A. nidulans by bimolecular fluorescence complementation. This is the first demonstration of how a kinesin-3 motor protein distinguishes among different MT populations in fungal cells, and how specificity determination depends on the tail rather than the motor domain, as has been demonstrated for kinesin 1 in neuronal cells.     

Sickness Presenteeism Predicts Suboptimal Self-Rated Health and Sickness Absence: A Nationally Representative Study of the Swedish Working Population

Background

Earlier studies have suggested that sickness presenteeism (SP) may be a risk factor for future health problems. The purpose of the present study was to test this in a nationally representative prospective study of Swedish workers.

Methods

Prospective cohort with a representative sample of the Swedish working population surveyed in 2008 and 2010. Odds ratios (OR) with 95% confidence intervals (CI) were calculated using logistic regression.

Results

Those who reported more than 7 days of SP had higher risk of suboptimal SRH compared to those who reported no SP (OR = 5.95; 95% CI 4.98–7.12), also after adjustment for confounders (OR = 1.64; 95% CI 1.30–2.06). Those who reported 1–7 days of SP also had an increased risk before and after adjustments. Inclusion of self-rated physical and psychological work capacity did not attenuate the associations, whereas of emotional exhaustion attenuated the ORs to non-significance for both outcomes, indicating that the health consequences associated with SP are largely related to mental health.

Conclusion

The results strengthen earlier findings suggesting that SP can be a risk factor for future suboptimal general health and sickness absence, particularly through mental health problems. This indicates that asking about SP could yield important information for employers, occupational health practitioners and GPs, possibly leading to more timely intervention that could decrease the risk of future sickness absence and more serious health problems, especially in the mental domain. Further studies of the possible causal pathways between SP and future health development are also warranted, especially since going to work is often seen as desirable also for those with poor health.     

Characterization of <Emphasis Type="Italic">Escherichia coli</Emphasis> [NiFe]-Hydrogenase Distribution During Fermentative Growth at Different pHs

The contribution made by each of the three active [NiFe]-hydrogenases (Hyd) of Escherichia coli during fermentation of glucose or glycerol in peptone-based medium at different pHs was analysed. The activities of the hydrogen-oxidizing Hyd-1 and Hyd-2 enzymes showed a reciprocal dependence on the pH of the medium while Hyd-3, a key component of the hydrogen-evolving formate hydrogenlyase complex, was mainly active at pH 6.5. Our findings identify the conditions during fermentation of glucose or glycerol under which each [NiFe]-hydrogenase is optimally active and demonstrate a previously unrecognized dependence on Hyd-1 activity at low pH.     

Regulation of the (pro)renin-renin receptor in cardiac remodelling

The (pro)renin-renin receptor [(P)RR] was discovered as an important novel component of the renin-angiotensin system (RAS). The functional significance of (P)RR is widely studied in renal and vascular pathologies and has sparked interest for a potential role in cardiovascular disease. To investigate the role of (P)RR in cardiac pathophysiology, we aimed to assess (P)RR regulation in adverse cardiac remodelling of the failing heart. In particular, we evaluated the expression of (P)RR in different models of heart failure and across different species. Significantly increased levels of (P)RR mRNA were found in post-myocardial infarcted (MI) hearts of rats (1.6-fold, P < 0.05) and mice (5-fold, P < 0.01), as well as in transgenic rats with overexpression of the mouse renin gene (Ren2) (2.2-fold, P < 0.01). Moreover, we observed a strong increase of (P)RR expression in hearts of dilated cardiomyopathy (DCM) patients (5.3-fold, P < 0.001). Because none of the tested commercially available antibodies appeared to detect endogenous (P)RR, a (P)RR-specific polyclonal antibody was generated to study (P)RR protein levels. (P)RR protein levels were significantly increased in the post-MI rat heart (1.4-fold, P < 0.05) as compared to controls. Most interestingly in DCM patients, a significant 8.7-fold (P < 0.05) increase was observed. Thus, protein expression paralleled gene expression. These results demonstrate that (P)RR expression is strongly up-regulated both in rodent models of heart failure and in the failing human heart, hinting to a potential role for (P)RR in cardiac pathophysiology.     

Green tea extracts interfere with the stress-protective activity of PrP and the formation of PrP

A hallmark in prion diseases is the conformational transition of the cellular prion protein (PrP(C)) into a pathogenic conformation, designated scrapie prion protein (PrP(Sc)), which is the essential constituent of infectious prions. Here, we show that epigallocatechin gallate (EGCG) and gallocatechin gallate, the main polyphenols in green tea, induce the transition of mature PrP(C) into a detergent-insoluble conformation distinct from PrP(Sc). The PrP conformer induced by EGCG was rapidly internalized from the plasma membrane and degraded in lysosomal compartments. Isothermal titration calorimetry studies revealed that EGCG directly interacts with PrP leading to the destabilizing of the native conformation and the formation of random coil structures. This activity was dependent on the gallate side chain and the three hydroxyl groups of the trihydroxyphenyl side chain. In scrapie-infected cells EGCG treatment was beneficial; formation of PrP(Sc) ceased. However, in uninfected cells EGCG interfered with the stress-protective activity of PrP(C). As a consequence, EGCG-treated cells showed enhanced vulnerability to stress conditions. Our study emphasizes the important role of PrP(C) to protect cells from stress and indicate efficient intracellular pathways to degrade non-native conformations of PrP(C).