Alpha A-crystallin and alpha B-crystallin, newly identified interaction proteins of protease-activated receptor-2, rescue astrocytes from C2-ceramide- and staurosporine-induced cell death.
Protease-activated receptor-2 (PAR-2) is a G protein-coupled receptor activated by trypsin and other trypsin-like serine proteases. The widely expressed PAR-2 is involved in inflammation response but the physiological/pathological roles of PAR-2 in the nervous system are still uncertain. In the present study, we report novel PAR-2 interaction proteins, alphaA-crystallin and alphaB-crystallin. These 20 kDa proteins have been implicated in neurodegenerative diseases like Alexander's disease, Creutzfeldt-Jacob disease, Alzheimer's disease, and Parkinson's disease. Results from yeast two-hybrid assay using the cytoplasmic C-tail of PAR-2 as bait suggested that alphaA-crystallin interacts with PAR-2. We further demonstrate the in vitro and cellular in vivo interaction of C-tail of PAR-2 as well as of full-length PAR-2 with alphaA(alphaB)-crystallins. We use pull-down, co-immunoprecipitation, and co-localization assays. Analysis of alphaA-crystallin deletion mutants showed that amino acids 120-130 and 136-154 of alphaA-crystallin are required for the interaction with PAR-2. Co-immunoprecipitation experiments ruled out an interaction of alphaA(alphaB)-crystallins with PAR-1, PAR-3, and PAR-4. This demonstrates that alphaA(alphaB)-crystallins are PAR-2-specific interaction proteins. Moreover, we investigated the functional role of PAR-2 and alpha-crystallins in astrocytes. Evidence is presented to show that PAR-2 activation and increased expression of alpha-crystallins reduced C2-ceramide- and staurosporine-induced cell death in astrocytes. Thus, both PAR-2 and alpha-crystallins are involved in cytoprotection in astrocytes.
Li
Rongyu
R
Medizinische Fakultät, Institut für Neurobiochemie, Otto-von-Guericke-Universität Magdeburg, Magdeburg 39120, Germany.
Rohatgi
Tanuja
T
Hanck
Theodor
T
Reiser
Georg
G
D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000502:physiology*; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002478:Cells, Cultured; D006801:Humans; D047468:Immunoprecipitation; D007313:Insecta; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology; D051381:Rats; D044464:Receptor, PAR-2 / Q000378:metabolism*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000633:toxicity; D019311:Staurosporine / Q000633:toxicity*; D038202:alpha-Crystallin A Chain / Q000378:metabolism* / Q000502:physiology; D038203:alpha-Crystallin B Chain / Q000378:metabolism* / Q000502:physiology
C064769:N-acetylsphingosine; D044464:Receptor, PAR-2; D038202:alpha-Crystallin A Chain; D038203:alpha-Crystallin B Chain; D019311:Staurosporine; D013110:Sphingosine