Mitochondrial heat shock protein (HSP) 70 synergizes with HSP60 in transducing endothelial cell apoptosis induced by anti-HSP60 autoantibody.
Heat shock protein (HSP) 60, up-regulated by endothelial cells (ECs) to resist stress, is the target of a subgroup of apoptosis-inducing anti-EC autoantibodies (Abs) in human vasculitides. Given that HSP60 is not a transmembrane protein, the mechanism by which these auto-Abs induces apoptosis is unclear. EC membrane proteins were analyzed using bidimensional electrophoresis and Far Western blot, and the HSP60 receptor was identified by mass spectrometry. Heat stress-dependent synthesis of HSP60 and receptor was examined by semiquantitative RT-PCR, and expression was examined by flow cytometry and indirect immunofluorescence. Interaction was demonstrated by coimmunoprecipitations. Lipid rafts were purified to evaluate specific localization, and the apoptotic response was investigated by blocking monoclonal Ab. Mitochondrial HSP70 (mtHSP70) was identified as an HSP60 receptor. Stress was required for ECs to up-regulate mRNA and express mtHSP70 on their surface. HSP60 and mtHSP70 colocalized and interacted within lipid rafts. They were associated with chemokine CC motif receptor 5 (CCR5), also induced at the mRNA and protein levels in stressed ECs. CCR5 was involved in the anti-HSP60-triggered apoptosis of ECs. These results provide new insights into the mechanism by which anti-EC auto-Abs from vasculitides induce apoptosis of ECs.
Université Européenne de Bretagne, Brest, France.
Jean-Eric
JE
Alard
Maryvonne
M
Dueymes
Rizgar A
RA
Mageed
Alain
A
Saraux
Pierre
P
Youinou
Christophe
C
Jamin
D017209:Apoptosis / Q000502:physiology*; D001323:Autoantibodies / Q000008:administration & dosage; D001483:Base Sequence; D002460:Cell Line; D002478:Cells, Cultured; D018834:Chaperonin 60 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D021962:Membrane Microdomains / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D019713:Receptors, CCR5 / Q000235:genetics / Q000378:metabolism; D013312:Stress, Physiological
D001323:Autoantibodies; D018834:Chaperonin 60; D017931:DNA Primers; D018840:HSP70 Heat-Shock Proteins; D012333:RNA, Messenger; D019713:Receptors, CCR5