Electronic cigarette use has raised concern worldwide regarding potential health risks and its position in tobacco cessation strategies. As part of any toxicity assessment, the chemical characterization of e-liquids and their related vapors are among fundamental data to be determined. Considering the lack of available reference methods, we developed and validated several analytical procedures in order to conduct a multicomponent analysis of six e-liquid refills and their resultant vapor emissions (generated by a smoking machine), and compared them with tobacco smoke. We combined several techniques including gas-chromatography, high and ultra-performance liquid chromatography and inductively coupled plasma with mass spectrometry or ultraviolet and flame ionization detection in order to identify the main e-liquid constituents (propylene glycol, glycerol and nicotine), as well as multiple potentially harmful components (trace elements, polycyclic aromatic hydrocarbons (PAHs), pesticides and carbonyl compounds). Regarding propylene glycol, glycerol and nicotine concentrations, the six tested e-liquids comply with the advertised composition and contain only traces of pollutants. Noticeable lower concentrations of trace elements (≤3.4 pg/mL puff), pesticides (

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International audienc

The Golgi ion homeostasis is tightly regulated to ensure essential cellular processes such as glycosylation, yet our understanding of this regulation remains incomplete. Gdt1p is a member of the conserved Uncharacterized Protein Family (UPF0016). Our previous work suggested that Gdt1p may function in the Golgi by regulating Golgi Ca/Mn homeostasis. NMR structural analysis of the polymannan chains isolated from yeasts showed that the gdt1Δ mutant cultured in presence of high Ca concentration, as well as the pmr1Δ and gdt1Δ/pmr1Δ strains presented strong late Golgi glycosylation defects with a lack of α-1,2 mannoses substitution and α-1,3 mannoses termination. The addition of Mn confirmed the rescue of these defects. Interestingly, our structural data confirmed that the glycosylation defect in pmr1Δ could also completely be suppressed by the addition of Ca. The use of Pmr1p mutants either defective for Ca or Mn transport or both revealed that the suppression of the observed glycosylation defect in pmr1Δ strains by the intraluminal Golgi Ca requires the activity of Gdt1p. These data support the hypothesis that Gdt1p, in order to sustain the Golgi glycosylation process, imports Mn inside the Golgi lumen when Pmr1p exclusively transports Ca. Our results also reinforce the functional link between Gdt1p and Pmr1p as we highlighted that Gdt1p was a Mn sensitive protein whose abundance was directly dependent on the nature of the ion transported by Pmr1p. Finally, this study demonstrated that the aspartic residues of the two conserved motifs E-x-G-D-[KR], likely constituting the cation binding sites of Gdt1p, play a crucial role in Golgi glycosylation and hence in Mn/Catransport.

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Congenital disorders of glycosylation are severe inherited diseases in which aberrant protein glycosylation is a hallmark. Transmembrane protein 165 (TMEM165) is a novel Golgi transmembrane protein involved in type II congenital disorders of glycosylation. Although its biologic function is still a controversial issue, we have demonstrated that the Golgi glycosylation defect due to TMEM165 deficiency resulted from a Golgi Mn2+ homeostasis defect. The goal of this study was to delineate the cellular pathway by which extracellular Mn2+ rescues N‐glycosylation in TMEM165 knockout (KO) cells. We first demonstrated that after extracellular exposure, Mn2+ uptake by HEK293 cells at the plasma membrane did not rely on endocytosis but was likely done by plasma membrane transporters. Second, we showed that the secretory pathway Ca2+‐ATPase 1, also known to mediate the influx of cytosolic Mn2+ into the lumen of the Golgi apparatus, is not crucial for the Mn2+‐induced rescue glycosylation of lysosomal‐associated membrane protein 2 (LAMP2). In contrast, our results demonstrate the involvement of cyclopiazonic acid—and thapsigargin (Tg)‐sensitive pumps in the rescue of TMEM165‐associated glycosylation defects by Mn2+. Interestingly, overexpression of sarco/endoplasmic reticulum Ca2+‐ATPase (SERCA) 2b isoform in TMEM165 KO cells partially rescues the observed LAMP2 glycosylation defect. Overall, this study indicates that the rescue of Golgi N‐glycosylation defects in TMEM165 KO cells by extracellular Mn2+ involves the activity of Tg and cyclopiazonic acid–sensitive pumps, probably the SERCA pumps.—Houdou, M., Lebredonchel, E., Garat, A., Duvet, S., Legrand, D., Decool, V., Klein, A., Ouzzine, M., Gasnier, B., Potelle, S., Foulquier, F. Involvement of thapsigargin—and cyclopiazonic acid–sensitive pumps in the rescue of TMEM165‐associated glycosylation defects by Mn2+. FASEB J. 33, 2669–2679 (2019). http://www.fasebj.org