Anti-CD3 activation of human CD4+ T cells increases expression of the intracellular beta-endorphin endopeptidase (IDE/gamma-EpGE).
In this study, increased expression of an endopeptidase hydrolyzing beta-endorphin (beta-Ep) to gamma-endorphin (gamma-Ep, beta-Ep1-17) was observed upon immobilized anti-CD3 stimulated activation of human peripheral blood CD4+ T cells (hCD4+ T cells). Although freshly isolated hCD4+ T cells are devoid of significant beta-Ep endopeptidase activity ( < 0.1 nmol h(-1) 10(6) cells (-1)), activation of these cells with immobilized anti-CD3 results in a time dependent appearance of beta-Ep endopeptidase activity which reaches a maximal value of 17.4+/-0.48 nmol h(-1) 10(6) cells(-1) after 48 h of culture. Significant up-regulation of both mRNA encoding IDE/gamma-EpGE and immunoreactive protein are observed in anti-CD3 stimulated hCD4+ T cells, indicating transcription and translation of IDE/gamma-EpGE may be elevated. No significant hydrolysis of exogenous beta-Ep is observed with intact hCD4+ T cells whether quiescent or activated or from preparations of hCD4+ T cell membranes. Therefore, this activity appears to be intracellular. Immunoreactive IDE/gamma-EpGE is detected inside activated hCD4+ T cells. Analysis of metabolites generated upon hydrolysis of beta-Ep with lysed activated hCD4+ T cell preparations identified the presence of: beta-Ep1-18, beta-Ep2-18, beta-Ep1-17, beta-Ep2-17, beta-Ep18-31, beta-Ep19-31, beta-Ep1-13, beta-Ep2-13, beta-Ep18-26, and beta-Ep20-31 as major metabolites and the majority of these are consistent with beta-Ep hydrolytic activity attributable to IDE/gamma-EpGE.
Sarada
B
B
Department of Biochemistry, The University of Texas Southwestern Medical Center at Dallas, 75235-9038, USA.
Thiele
D
D
Dang
T
T
Lee
J
J
Safavia
A
A
Hersh
L B
LB
Cottam
G L
GL
D017252:CD3 Complex / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000201:enzymology* / Q000276:immunology*; D002851:Chromatography, High Pressure Liquid; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D007425:Intracellular Membranes / Q000201:enzymology*; D010446:Peptide Fragments / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D001615:beta-Endorphin / Q000378:metabolism
D017252:CD3 Complex; D010446:Peptide Fragments; D012333:RNA, Messenger; D001615:beta-Endorphin; D010450:Endopeptidases; C043537:gamma-endorphin generating enzyme