Changes in the relative amount of subunits of methionine adenosyltransferase in human lymphocytes upon stimulation with a polyclonal T cell mitogen.
Activation of resting human peripheral blood T lymphocytes by the lectin phytohemagglutinin results in an increase in methionine adenosyltransferase (MAT) activity, accompanied by an increase in the amount of the alpha/alpha' catalytic subunits of the enzyme. In contrast, the amount of the noncatalytic beta subunit remains constant throughout the course of the response. Using both polyclonal antibodies to the holoenzyme and monoclonal antibodies to the alpha/alpha' subunits, we detected a cross-reactive 68-kDa protein, which we refer to as lambda. This protein is present in high abundance in resting T cells but decreases upon cell stimulation, as both MAT activity and the amount of the catalytic alpha/alpha' subunits increase. The decrease in lambda and increase in alpha/alpha' occurs after interleukin-2 production and before DNA synthesis. lambda virtually disappears when the cells are actively dividing. Several continuous T cell lines (HPB-ALL, MOLT-4, and Jurkat) as well as a freshly isolated T cell leukemia (ALL-2) had no detectable lambda. The Km for L-methionine for enzyme from resting peripheral blood mononuclear cells was 19-23 microM, which is 3-8-fold higher than purified MAT from fresh leukemic cells or enzyme from Jurkat cells, both of which have a Km of 3.5-3.8 microM. Kinetic analysis of enzyme activity from activated peripheral blood mononuclear cells suggested the presence of two forms of enzyme catalyzing the synthesis of AdoMet. After separation of lambda from the alpha and beta subunits by hydrophobic chromatography, it was determined that lambda has MAT activity but that it is significantly less active than the form containing the alpha subunit. It therefore appears that in resting T cells MAT is sequestered as a less active form. We hypothesize that lambda is a precursor to the catalytic subunits of human lymphocyte MAT and propose that the transition from lambda to alpha/alpha' may be important in the response of T cells to mitogenic signals.
De La Rosa
J
J
Research Service, Veterans Administration Medical Center, Memphis, Tennessee 38104.
LeGros
H L
HL
Geller
A M
AM
Kotb
M
M
D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D002384:Catalysis; D002853:Chromatography, Liquid; D003429:Cross Reactions; D004247:DNA / Q000096:biosynthesis; D004591:Electrophoresis, Polyacrylamide Gel; D004792:Enzyme Precursors / Q000378:metabolism; D006801:Humans; D007376:Interleukin-2 / Q000096:biosynthesis; D007700:Kinetics; D015458:Leukemia, T-Cell / Q000201:enzymology; D008716:Methionine Adenosyltransferase / Q000378:metabolism*; D010835:Phytohemagglutinins; D013601:T-Lymphocytes / Q000187:drug effects / Q000201:enzymology*; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D004792:Enzyme Precursors; D007376:Interleukin-2; D010835:Phytohemagglutinins; D004247:DNA; D008716:Methionine Adenosyltransferase