URIDINE-DIPHOSPHATE GLUCOSE-METABOLISM AND CALLOSE SYNTHESIS IN CULTURED POLLEN TUBES OF NICOTIANA-ALATA LINK ET OTTO

Membrane preparations from cultured pollen tubes of Nicotiana alata Li nk et Otto contain a Ca2+-independent (1-3)-beta-D-glucan (callose) sy nthase activity that has a low affinity for UDP-glucose, even when act ivated by treatment with trypsin (H. Schlupmann, A. Bacic, S.M. Read [ 1993] Planta 191: 470-481). Therefore, we investigated whether UDP-glu cose was a likely substrate for callose synthesis in actively growing pollen tubes. Deposition of (1-3)-beta-glucan occurred at a constant r ate, 1.4 to 1.7 nmol glucose min(-1), in tubes from 1 mg of pollen fro m 3 h after germination; however, the rate of incorporation of radioac tivity from exogenous [C-14]sucrose into wall polymers was not constan t, but increased until at least 8 h after germination, probably due to decreasing use of internal reserves. UDP-glucose was a prominent ultr aviolet-absorbing metabolite in pollen-tube extracts, with 1.6 nmol pr esent in tubes from 1 mg of pollen, giving a calculated cytoplasmic co ncentration of approximately 3.5 mM. Radioactivity from [C-14]sucrose was rapidly incorporated into sugar monophosphates and UDP-glucose by the growing tubes, consistent with a turnover time for UDP-glucose of less than 1 min; the specific radioactivity of extracted UDP-[C-14]glu cose was equal to that calculated from the rate of incorporation of [C -14]sucrose into wall glucans. Large amounts of less metabolically act ive neutral sugars were also present. The rate of synthesis of (1-3)-b eta-glucan by nontrypsin-treated pollen-tube membrane preparations inc ubated with 3.5 mM UDP-glucose and a beta-glucoside activator was slig htly greater than the rate of deposition of (1-3)-beta-glucan by intac t pollen tubes. These data are used to assess the physiological signif icance of proteolytic activation of pollen-tube callose synthase.