Differential phosphorylation of c-Jun and JunD in response to the epidermal growth factor is determined by the structure of MAPK targeting sequences

MAPK phosphorylation of various substrates is mediated by the presence of docking sites, including the D domain and the DEF motif. Depending on the number and sequences of these domains, substrates are phosphorylated by specific subsets of MAPKs. For example, a D domain targets JNK to c-Jun, whereas a DEF motif is required for ERK phosphorylation of c-Fos. JunD, in contrast, contains both D and DEF domains. Here we show that these motifs mediate JunD phosphorylation in response to either ERK or JNK activation. An intact D domain is required for phosphorylation and activation of JunD by both subtypes of MAPK. The DEF motif acts together with the D domain to elicit efficient phosphorylation of JunD in response to the epidermal growth factor (EGF) but has no function on JunD phosphorylation and activation by JNK signaling. Furthermore, we show that conversion of a c-Jun sequence to a canonical DEF domain, as it is present in JunD, elicits c-Jun activation in response to EGF. Our results suggest that evolution of a particular modular system of MAPK targeting sequences has determined a differential response of JunD and c-Jun to ERK activation

Thermodynamic analysis of porphyrin binding to Momordica charantia (bitter gourd) lectin

Owing to the use of porphyrins in photodynamic therapy for the treatment of malignant tumors, and the preferential interaction of lectins with tumor cells, studies on lectin-porphyrin interaction are of significant interest. In this study, the interaction of several free-base and metalloporphyrins with Momordica charantia (bitter gourd) lectin (MCL) was investigated by absorption spectroscopy. Difference absorp-tion spectra revealed that significant changes occur in the Soret band region of the porphyrins on binding to MCL. These changes were monitored to obtain association constants (Ka) and stoichiometry of binding. The tetrameric MCL binds four porphyrin molecules, and the stoichiometry was unaffected by the presence of the specific sugar, lactose. In addition, the agglutination activity of MCL was unaffected by the presence of the porphyrins used in this study, clearly indicating that porphyrin and carbohydrate ligands bind at different sites. Both cationic and anionic porphyrins bind to the lectin with comparable affinity (Ka =103-105 m-1). The thermodynamic parameters associated with the interaction of several porphyrins, obtained from the temperature dependence of the Ka values, were found to be in the range: Δ H° = -98.1 to -54.4 kJ·mol-1 and Δ S° =-243.9 to -90.8 J·mol-1·K-1. These results indicate that porphyrin binding to MCL is governed by enthalpic forces and that the contribution from binding entropy is negative. Enthalpy-entropy compensation was observed in the interaction of different porphyrins with MCL, underscoring the role of water structure in the overall binding process. Analysis of CD spectra of MCL indicates that this protein contains about 13%α-helix, 36%β-sheet, 21%β-turn, and the rest unordered structures. Binding of porphyrins does not significantly alter the secondary and tertiary structures of MCL

Differential Scanning calorimetric studies on the interaction of N-acylethanolamines with cholesterol

Earlier studies have suggested the formation of a 1 : 1 (mol/mol) complex between N-myristoylethanolamine (NMEA) and cholesterol in aqueous dispersion. In this study, this interaction has been investigated further by differential scanning calorimetry (DSC) on dry mixtures of NMEA, N-palmitoylethanolamine (NPEA) and N-stearoylethanolamine (NSEA) with cholesterol. The results obtained indicate that addition of cholesterol to NMEA leads to a new phase transition at 86.5°C, besides the solid-liquid phase transition of NMEA at 95°C. The intensity of the peak corresponding to the new transition increases with cholesterol content up to 50 mol%, but decreases thereafter, whereas the intensity of the peak corresponding to the melting of NMEA decreases with increasing cholesterol content, with concomitant and gradual shift to lower temperatures and vanishes at 50 mol% cholesterol. These results are consistent with the formation of a 1:1 molar complex between NMEA and cholesterol proposed earlier and indicate that these two amphiphiles are associated in the solid state as well. DSC studies on hydrated mixtures of NPEA and NSEA with cholesterol yielded results that parallel those obtained with the NMEA/cholesterol system, indicating that these two long-chain NAEs also form 1:1 (mol/mol) complexes with cholesterol

Sputum induced cellularity in a group of traffic policemen.

It has been demonstrated that a group of workers (e.g. waste handlers) daily exposed to a traffic related air pollution present airway inflammation in term of an increase of neutrophilic inflammation. The aim of our study was to determine the presence of airways inflammation detected by induced sputum in a population of traffic policemen (TP) in the city of Bari, compared to a group of healthy subjects (HS) without any occupational exposure to inhalation of traffic-related air pollution. Twelve non smokers, non atopics, healthy traffic policemen with a history of exposure to airway pollution and 12 HS underwent sputum induction. TP show a statistically significant increase in the percentage neutrophil cell count (median and IQ range) compared to the HS (65 and 13.5 vs. 40.5 and 9.5; p<0.01). In conclusion we have found that policemen chronically exposed to air pollution presented airway neutrophilic inflammation and the results of this pilot study could be strictly considered for the long term effect of a traffic pollution in airway inflammation and the lung function

Inverse relationship between chitobiase and transglycosylation activities of chitinase-D from Serratia proteamaculans revealed by mutational and biophysical analyses

Serratia proteamaculans chitinase-D (SpChiD) has a unique combination of hydrolytic and transglycosylation (TG) activities. The TG activity of SpChiD can be used for large-scale production of chito-oligosaccharides (CHOS). The multiple activities (hydrolytic and/or chitobiase activities and TG) of SpChiD appear to be strongly influenced by the substrate-binding cleft. Here, we report the unique property of SpChiD substrate-binding cleft, wherein, the residues Tyr28, Val35 and Thr36 control chitobiase activity and the residues Trp160 and Trp290 are crucial for TG activity. Mutants with reduced (V35G and T36G/F) or no (SpChiD&#916;30–42 and Y28A) chitobiase activity produced higher amounts of the quantifiable even-chain TG product with degree of polymerization (DP)-6, indicating that the chitobiase and TG activities are inversely related. In addition to its unprecedented catalytic properties, unlike other chitinases, the single modular SpChiD showed dual unfolding transitions. Ligand-induced thermal stability studies with the catalytically inactive mutant of SpChiD (E153A) showed that the transition temperature increased upon binding of CHOS with DP2–6. Isothermal titration calorimetry experiments revealed the exceptionally high binding affinities for E153A to CHOS with DP2–6. These observations strongly support that the architecture of SpChiD substrate-binding cleft adopted to control chitobiase and TG activities, in addition to usual chitinase-mediated hydrolysis

Effect of a Park-Based Physical Activity Intervention on Psychological Wellbeing at the Time of COVID-19

Practicing regular physical activity in green spaces has been invocated as a promising strategy for improving wellbeing in urban settings. The aim of the study was to assess the effect of a structured park-based physical activity intervention, the “Moving Parks” project, on citizens’ wellbeing at the time of COVID-19. The intervention was carried out in six public parks in Bologna (Italy) and administered by qualified instructors (from May 2021 to September 2021). The Psychological General Well Being Index short form questionnaire was administered before and after the three months of outdoor activities. A total of 328 participants completed the questionnaire at the beginning and at the end of the project. In September 2021, all psychosocial domains of the questionnaire (anxiety, depressed mood, self-control, positive well-being, vitality energy, and vitality-tiredness) significantly improved in the female sample (p value &lt; 0.01) and only the last two in the male sample (p value &lt; 0.05). The “Moving Parks” project seems to be able to improve citizens’ psychological wellbeing, particularly in women

The v-Ki-Ras Oncogene Alters cAMP Nuclear Signaling by Regulating the Location and the Expression of cAMP-dependent Protein Kinase IIβ

The v-Ki-Ras oncoprotein dedifferentiates thyroid cells and inhibits nuclear accumulation of the catalytic subunit of cAMP-dependent protein kinase. After activation of v-Ras or protein kinase C, the regulatory subunit of type II protein kinase A, RIIbeta, translocates from the membranes to the cytosol. RIIbeta mRNA and protein were eventually depleted. These effects were mimicked by expressing AKAP45, a truncated version of the RII anchor protein, AKAP75. Because AKAP45 lacks membrane targeting domains, it induces the translocation of PKAII to the cytoplasm. Expression of AKAP45 markedly decreased thyroglobulin mRNA levels and inhibited accumulation of C-PKA in the nucleus. Our results suggest that: 1) The localization of PKAII influences cAMP signaling to the nucleus; 2) Ras alters the localization and the expression of PKAII; 3) Translocation of PKAII to the cytoplasm reduces nuclear C-PKA accumulation, resulting in decreased expression of cAMP-dependent genes, including RIIbeta, TSH receptor, and thyroglobulin. The loss of RIIbeta permanently down-regulates thyroid-specific gene expression