Quantitation of Starch and ADP-Glucose Pyrophosphorylase in non-Embryogenic Cells and Embryogenic Cell Clusters from Carrot Suspension Cultures

Abstract

Summary Cultures of carrot cells ( Daucus carota L. var. Danver) in a medium containing auxin were fractionated by size and density into embryogenic cell clusters and non-embryogenic cells. The embryogenic potentials of the embryogenic cell clusters were 800- to 3000-fold greater than those of the non-embryogenic cells. These two fractions were analyzed for their starch content and ADP-glucose pyrophosphorylase (E.C. 2.7.7.27) activity and subunit composition. Starch content was significantly greater (15- to 40-fold) in the embryogenic cell clusters. Therefore, the embryogenic potential of these two populations of cells, in the same medium, can be distinguished by their cellular starch content. However, this difference in starch content was not reflected in the starch biosynthetic capacity of these fractions, as inferred from the ADP-glucose pyrophosphorylase activity, which was similar in the two fractions. This implies that the level of ADP-glucose pyrophosphorylase is independent of embryogenic potential and does not determine the steady-state starch content for these cells. Immunoblot analysis with antiserum to the purified spinach leaf enzyme revealed a 50 kDa subunit of ADP-glucose pyrophosphorylase in both non-embryogenic cells and embryogenic cell clusters. A 100 kDa polypeptide which cross reacted with antiserum to ADP-glucose pyrophosphorylase was present only in embryogenic cell clusters. This 100 kDa polypeptide is one of the few examples of a polypeptide whose accumulation is correlated with somatic embryogenic potential.