The 86-kDa subunit of autoantigen Ku is a somatostatin receptor regulating protein phosphatase-2A activity.
We previously reported the immunopurification of a somatostatin receptor from the human tumoral gastric cell HGT1 using the monoclonal antibody 30F3 (Reyl-Desmars, F., Le Roux, S., Linard, C., Benkouka, F., and Lewin, M. J. M. (1989) J. Biol. Chem. 264, 18789-18795). Screening of a lambda gt11 HGT1-cDNA library with 30F3 led us to isolate a cDNA encoding an 86-kDa polypeptide displaying 100% structural identity with the 86-kDa subunit (p86-Ku) of the Ku autoantigen. Recombinant p86 expressed in Escherichia coli cross-reacted with 30F3 and specifically bound [125I-Tyr11]somatstatin-14. Binding was totally displaced by somatostatin-14, somatostatin-28, and SMS 201-995, with IC50 values of 0.7, 1.0, and 1.2 nM, respectively. In a search for a biological effect associated with binding, we purified a 36-kDa, okadaic acid-sensitive phosphatase (protein phosphatase-2A (PP2A)) from rat gastric cytosol. PP2A catalyzed 32P release from p34cdc2-phosphorylated histone H1. However, PP2A-induced 32P release was concentration dependently inhibited by recombinant p86-Ku, with a decrease in maximal velocity without a change in Km. Steric exclusion high pressure chromatography indicated that the inhibition resulted from direct interaction of the enzyme with p86-Ku. Furthermore, it was antagonized by increased concentrations of somatostatin-14 and prevented by preincubating p86-Ku with 30F3. Given the key role played by PP2A in cell cycle regulation, the current findings suggest that p86-Ku could be a physiological target of somatostatin antiproliferative action.
Unité de Recherches de Gastroentérologie, INSERM U10, Hôpital Bichat-Claude Bernard, Paris, France.
M
M
Le Romancer
F
F
Reyl-Desmars
Y
Y
Cherifi
C
C
Pigeon
S
S
Bottari
O
O
Meyer
M J
MJ
Lewin
D000818:Animals; D000906:Antibodies; D034961:Antigens, Nuclear; D001324:Autoantigens / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004265:DNA Helicases; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D006801:Humans; D000072200:Ku Autoantigen; D008297:Male; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010766:Phosphorylation; D054648:Protein Phosphatase 2; D051381:Rats; D017208:Rats, Wistar; D017481:Receptors, Somatostatin / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D000906:Antibodies; D034961:Antigens, Nuclear; D001324:Autoantigens; D004268:DNA-Binding Proteins; D006657:Histones; D009687:Nuclear Proteins; D017481:Receptors, Somatostatin; D011994:Recombinant Proteins; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; C000606511:Xrcc6 protein, rat; D000072200:Ku Autoantigen