title: Analysis of properties of small heat shock protein Hsp25 in MAPK-activated protein kinase 2 (MK2)-deficient cells: MK2-dependent insolubilization of Hsp25 oligomers correlates with susceptibility to stress.
abstract: Small heat shock proteins (sHsps) exist in dynamic oligomeric complexes and display diverse biological functions ranging from chaperone properties to modulator of apoptosis. So far, the role of stress-dependent phosphorylation of mammalian sHsps for its structure and function has been analyzed by using various phosphorylation site mutants overexpressed in different cell types as well as by non-exclusive inhibitors of the p38 MAPK cascade. Here we investigate the role of phosphorylation of endogenous sHsp in a genetic model lacking the major Hsp25 kinase, the MAP kinase-activated protein kinase MK2. We demonstrate that in MK2-deficient fibroblasts, where no stress-dependent phosphorylation of Hsp25 at Ser86 and no in vitro binding to 14-3-3 was detectable, stress-dependent disaggregation of endogenous Hsp25 complexes is impared and kinetics of arsenite-dependent, H2O2-dependent, and sublethal heat shock-induced insolubilization of Hsp25 is delayed. Similarly, green fluorescent protein-tagged Hsp25 shows retarded subcellular accumulation into stress granules in MK2-deficient cells after arsenite treatment. Decreased insolubilization of Hsp25 in MK2-deficient cells correlates with increased resistance against arsenite, H2O2, and sublethal heat shock treatment and with decreased apoptosis. In contrast, after severe, lethal heat shock MK2-deficient embryonic fibroblasts cells show fast and complete insolubilization of Hsp25 independent of MK2 and no increased stress resistance. Hence, MK2-dependent formation of insoluble stress granules and irreversible cell damage by oxidative stresses and sublethal heat shock correlate and only upon severe, lethal heat shock MK2-independent processes could determine insolubilization of Hsp25 and are more relevant for cellular stress damage.
authors.affiliation: Institute of Biochemistry, Medical School Hannover, Hannover 30625, Germany.
authors.forename: Anastassiia
authors.initials: A
authors.lastname: Vertii
authors.forename: Cosima
authors.initials: C
authors.lastname: Hakim
authors.forename: Alexey
authors.initials: A
authors.lastname: Kotlyarov
authors.forename: Matthias
authors.initials: M
authors.lastname: Gaestel
mesh_terms: D048948:14-3-3 Proteins / Q000737:chemistry; D000818:Animals; D017209:Apoptosis; D049109:Cell Proliferation; D002470:Cell Survival; D002850:Chromatography, Gel; D005347:Fibroblasts / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000737:chemistry*; D006861:Hydrogen Peroxide / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D020935:MAP Kinase Signaling System; D051379:Mice; D018832:Molecular Chaperones; D009363:Neoplasm Proteins / Q000737:chemistry*; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics* / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism; D000099190:MAP-Kinase-Activated Kinase 2
chemical_list: D048948:14-3-3 Proteins; D006360:Heat-Shock Proteins; D006861:Hydrogen Peroxide; D047908:Intracellular Signaling Peptides and Proteins; D018832:Molecular Chaperones; D009363:Neoplasm Proteins; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D000099190:MAP-Kinase-Activated Kinase 2; C486113:Hsbp1 protein, mouse