A dual-omics approach on the effects of fibroblast growth factor-2 (FGF-2) on ventral tegmental area dopaminergic neurons in response to alcohol consumption in mice.

Harmful alcohol consumption is a major socioeconomic burden to the health system, as it can be the cause of mortality of heavy alcohol drinkers. The dopaminergic (DAergic) system is thought to play an important role in the pathogenesis of alcohol drinking behaviour; however, its exact role remains elusive. Fibroblast growth factor 2 (FGF-2), a neurotrophic factor, associated with both the DAergic system and alcohol consumption, may play an important role in DAergic neuroadaptations during alcohol abuse. Within this study, we aimed to clarify the role of endogenous FGF-2 on the DAergic system and whether there is a possible link to alcohol consumption. We found that lack of FGF-2 reduces the alcohol intake of mice. Transcriptome analysis of DAergic neurons revealed that FGF-2 knockout (FGF-2 KO) shifts the molecular fingerprint of midbrain dopaminergic (mDA) neurons to DA subtypes of the ventral tegmental area (VTA). In line with this, proteomic changes predominantly appear also in the VTA. Interestingly, these changes led to an altered regulation of the FGF-2 signalling cascades and DAergic pathways in a region-specific manner, which was only marginally affected by voluntary alcohol consumption. Thus, lack of FGF-2 not only affects the gene expression but also the proteome of specific brain regions of mDA neurons. Our study provides new insights into the neuroadaptations of the DAergic system during alcohol abuse and, therefore, comprises novel targets for future pharmacological interventions.

Obesity impacts hypoxia adaptation of the lung.

Obesity is mostly associated with adverse health consequences, but may also elicit favorable effects under chronic conditions. This "obesity paradox" is under debate for pulmonary diseases. As confounding factors complicate conclusions from human studies, this study used a controlled animal model combining diet-induced obesity and chronic hypoxia as a model for pulmonary hypertension and chronic obstructive pulmonary disease. Male C57BL/6 mice were fed control or high-fat diet for 30 wk, and half of the animals were exposed to chronic hypoxia (13% O2) for 3 wk. Hypoxia induced right ventricular hypertrophy, thickening of pulmonary arterial and capillary walls, higher lung volumes, and increased hemoglobin concentrations irrespective of the body weight. In contrast, lung proteomes differed substantially between lean- and obese-hypoxic mice. Many of the observed changes were linked to vascular and extracellular matrix (ECM) proteins. In lean-hypoxic animals, circulating platelets were reduced and abundances of various clotting-related proteins were altered, indicating a hypercoagulable phenotype. Moreover, the septal ECM composition was changed, and airspaces were significantly distended pointing to lung hyperinflation. These differences were mostly absent in the obese-hypoxic group. However, the obesity-hypoxia combination induced the lowest blood CO2 concentrations, indicating hyperventilation for sufficient oxygen supply. Moreover, endothelial surface areas were increased in obese-hypoxic mice. Thus, obesity exerts differential effects on lung adaptation to hypoxia, which paradoxically include not only adverse but also rather protective changes. These differences have a molecular basis in the lung proteome and may influence the pathogenesis of lung diseases. This should be taken into account for future individualized prevention and therapy.NEW & NOTEWORTHY An "obesity paradox" is discussed for pulmonary diseases. By linking lung proteome analyses to pulmonary structure and function, we demonstrate that diet-induced obesity affects lung adaptation to chronic hypoxia in various ways. The observed changes include not only adverse but also protective effects and are associated with altered abundances of vascular and extracellular matrix proteins. These results highlight the existence of relevant differences in individuals with obesity that may influence the pathogenesis of lung diseases.

Clostridiumnovyi's Alpha-Toxin Changes Proteome and Phosphoproteome of HEp-2 Cells.

C. novyi type A produces the alpha-toxin (TcnA) that belongs to the large clostridial glucosylating toxins (LCGTs) and is able to modify small GTPases by N-acetylglucosamination on conserved threonine residues. In contrast, other LCGTs including Clostridioides difficile toxin A and toxin B (TcdA; TcdB) modify small GTPases by mono-o-glucosylation. Both modifications inactivate the GTPases and cause strong effects on GTPase-dependent signal transduction pathways and the consequent reorganization of the actin cytoskeleton leading to cell rounding and finally cell death. However, the effect of TcnA on target cells is largely unexplored. Therefore, we performed a comprehensive screening approach of TcnA treated HEp-2 cells and analyzed their proteome and their phosphoproteome using LC-MS-based methods. With this data-dependent acquisition (DDA) approach, 5086 proteins and 9427 phosphosites could be identified and quantified. Of these, 35 proteins were found to be significantly altered after toxin treatment, and 1832 phosphosites were responsive to TcnA treatment. By analyzing the TcnA-induced proteomic effects of HEp-2 cells, 23 common signaling pathways were identified to be altered, including Actin Cytoskeleton Signaling, Epithelial Adherens Junction Signaling, and Signaling by Rho Family GTPases. All these pathways are also regulated after application of TcdA or TcdB of C. difficile. After TcnA treatment the regulation on phosphorylation level was much stronger compared to the proteome level, in terms of both strength of regulation and the number of regulated phosphosites. Interestingly, various signaling pathways such as Signaling by Rho Family GTPases or Integrin Signaling were activated on proteome level while being inhibited on phosphorylation level or vice versa as observed for the Role of BRCA1 in DNA Damage Response. ZIP kinase, as well as Calmodulin-dependent protein kinases IV & II, were observed as activated while Aurora-A kinase and CDK kinases tended to be inhibited in cells treated with TcnA based on their substrate regulation pattern.