Lack of glutathione peroxidase-8 in the ER impacts on lipid composition of HeLa cells microsomal membranes

GPx8 is a glu​tathione per​ox​i​dase ho​molog in​serted in the mem​branes of en​do​plas​mic retic​u​lum (ER), where it seem​ingly plays a role in con​trol​ling re​dox sta​tus by pre​vent​ing the spill of H2O2. We ad​dressed the im​pact of GPx8 si​lenc​ing on the lipidome of mi​cro​so​mal mem​branes, us​ing sta​bly GPx8-si​lenced HeLa cells. The two cell lines were clearly sep​a​rated by Prin​ci​pal Com​po​nent Analy​sis (PCA) and Par​tial Least Square Dis​crim​i​nant analy​sis (PLS-DA) of lipidome. Con​sid​er​ing in de​tail the in​di​vid​ual lipid classes, we ob​served that un​sat​u​rated glyc​erophos​pho​lipids (GPL) de​creased, while only in phos​phatidyli​nos​i​tols (PI) a sub​sti​tu​tion of mo​noun​sat​u​rated fatty acids (MUFA) for polyun​sat​u​rated fatty acids (PUFA) was ob​served. Among sphin​golipids (SL), ce​ramides (CER) de​creased while sphin​gomyelins (SM) and neu​tral gly​coph​in​! golipids (nGSL) in​creased. Here, in ad​di​tion, longer chains than in con​trols in the amide fatty acid were pre​sent. The in​crease up to four folds of the CER (d18:1; c24:0) con​tain​ing three hex​ose units, was the most re​mark​able species in​creas​ing in the dif​fer​en​tial lipidome of siG​Px8 cells. Quan​ti​ta​tive RT-PCR com​plied with lipidomic analy​sis specif​i​cally show​ing an in​creased ex​pres​sion of: i) acyl-CoA syn​thetase 5 (AC​SL5); ii) CER syn​thase 2 and 4; iii) CER trans​porter (CERT); iv) UDP-glu​co​syl trans​ferase (UDP-GlcT), as​so​ci​ated to a de​creased ex​pres​sion of UDP-galac​to​syl trans​ferase (UDP-GalT). A role of the un​folded pro​tein re​! sponse (UPR) and the spliced form of the tran​scrip​tion fac​tor XBP1 on the tran​scrip​tional changes of GPx8 si​lenced cells was ruled-out. Sim​i​larly, also the in​volve​ment of Nrf2 and NF-!B. Al​to​gether our re​sults in​di​! cate that GPx8-si​lenc​ing of HeLa yields a mem​brane de​pleted by about 24% of polyun​sat​u​rated GPL and a cor​re​spond​ing in​crease of sat​u​rated or mo​noun​sat​u​rated SM and spe​ci$c nGSL. This is ten​ta​tively in​ter​! preted as an adap​tive mech​a​nism lead​ing to an in​creased re​sis​tance to rad​i​cal ox​i​da​tions. More​over, the marked shift of fatty acid com​po​si​tion of PI emerges as a pos​si​bly rel​e​vant is​sue in re​spect to the im​pact of GPx8 on sig​nal​ing path​ways.