CD26/DPPIV signal transduction function, but not proteolytic activity, is directly related to its expression level on human Th1 and Th2 cell lines as detected with living cell cytochemistry.
CD26/DPPIV is a cell surface glycoprotein that functions both in signal transduction and as a proteolytic enzyme, dipeptidyl peptidase IV (DPPIV). To investigate how two separate functions of one molecule are regulated, we analyzed CD26 protein expression and DPPIV enzyme activity on living human T-helper 1 (Th1) and Th2 cells that express different levels of CD26/DPPIV. DPPIV activity was specifically determined with the synthetic fluorogenic substrate ala-pro-cresyl violet and CD26 protein expression was demonstrated with an FITC-conjugated CD26-specific antibody. Fluorescence of liberated cresyl violet (red) and FITC (green) was detected simultaneously on living T-cells using flow cytometry and spectrofluorometry. Th1 cells expressed three- to sixfold more CD26 protein than Th2 cells. The signal transduction function of the CD26/DPPIV complex, tested by measuring its co-stimulatory potential for proliferation, was directly related to the amount of CD26 protein at the cell surface. However, DPPIV activity was similar in both cell populations at physiological substrate concentrations because of differences in K(m) and V(max) values of DPPIV on Th1 and Th2 cells. Western blotting and zymography of Th1 and Th2 whole-cell lysates demonstrated similar patterns. This study shows that two functions of one molecule can be controlled differentially.
Boonacker
Emil P
EP
Academic Medical Center, University of Amsterdam, Department of Cell Biology and Histology, Amsterdam, The Netherlands.
Wierenga
Eddy A
EA
Smits
Hermelijn H
HH
Van Noorden
Cornelis J F
CJ
D002460:Cell Line; D016207:Cytokines / Q000096:biosynthesis; D018819:Dipeptidyl Peptidase 4 / Q000096:biosynthesis / Q000502:physiology*; D005434:Flow Cytometry; D006801:Humans; D007700:Kinetics; D018613:Microscopy, Confocal; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence; D018417:Th1 Cells / Q000201:enzymology / Q000378:metabolism*; D018418:Th2 Cells / Q000201:enzymology / Q000378:metabolism*
D016207:Cytokines; D018819:Dipeptidyl Peptidase 4