Abstract
Concanavalin A added to intact cells at 37 degrees caused rapid and reversible inactivation of a soluble enzyme, tyrosine aminotransferase, in two lines of rat hepatoma tissue culture cells grown in monolayer culture. This temperature-dependent process was independent of de novo protein and RNA synthesis and independent of increased uptake of Ca2+ and Mg2+ or glucose. The inactivation could be reversed by adding alpha-methyl-D-mannopyranoside a competing sugar for concanavalin A binding. Other lectins known to bind to different sugars did not bring about the inactivation of tyrosine aminotransferase. Addition of concanavalin A did not result in the inactivation of another soluble enzyme, lactic dehydrogenase. The maintenance of tyrosine aminotransferase in an inactive form after the binding of concanavalin A to the cells required the continued presence of concanavalin A. This effect of concanavalin A could not be mimicked either by dibutyryl cyclic adenosine or guanosine monophosphoric acid. Incubation of cell extracts with concanavalin A did not result in inactivation nor did mixing of extracts from concanavalin A-treated cells with extracts from untreated cells. On the basis of these results we conclude that the following are the essential requirements for concanavalin A to bring about the inactivation of tyrosine aminotransferase: (a) the binding of native concanavalin A to the cells; (b) integrity of certain structural elements of the cells.
Highlights
Concanavalin A added to intact cells at 3’7” caused rapid and reversible inactivation of a soluble enzyme, tyrosine aminotransferase, in two lines of rat hepatoma tissue cdture cells grown in monolayer culture
Samples were harvested for tyrosine aminotransferase assay as described previously at different times
The medium was replaced in the rest of the dishes with fresh medium containing 50 rnM a-methyl-n-mannopyranoside along with (O-0)
Summary
Binding of the lectin to either of two lines of rat hepatoma cells in tissue culture resulted in a rapid and reversible inactivation of the intracellular enzyme, tyrosine aminotransferase (EC 2.6.1.5). This enzyme, found in the soluble fraction of cell extracts, is induced in both lines by glucocorticosteroid hormones [10, 11] and insulin [12, 13], and in one of them (FU-5-5) by analogs of cyclic adenosine monophosphate [14, 15].
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