PREHARVEST AND POSTHARVEST MUSKMELON FRUIT CALCIUM- STIMULATED PROTEIN KINASE ACTIVITY

In orange-fleshed netted muskmelon (Cucumis melo L. var. reticulatus Naud.), hypodermal mesocarp plasma membrane senescence is retarded by low exogenous CaCl₂ concentrations (Lester, 1996). The effect of calcium on muskmelon plasma membrane senescence is directly related to maintenance of phospholipids, proteins, and H⁺-ATPase activity.
Calcium can regulate plant senescence through calcium-dependent protein kinase (CDPK) activity, which phosphorylates H⁺-ATPase (Schaller and Sussman, 1988). CDPK phosphorylation of mature and postharvest melon plasma membrane H⁺-ATPase is the objective of this study.
Melon plasma membrane isolation and H⁺-ATPase assays were as previously described (Lester and Stein, 1993). Kinase assays and gel electrophoresis were as previously described (Baizabal-Aguirre and Gonzalez de la Vara, 1994). Coincidently, melon plasma membrane H⁺-ATPase and protein kinase activity decrease following fruit maturation and postharvest storage.
Melon plasma membrane protein kinase activity is stimulated by calcium and promotes the phosphorylation of many proteins.
Among these the most conspicuously phosphorylated are 100 kDa proteins corresponding to the weight of all known H⁺-ATPase (Schaller and Sussman, 1988). However, the 100 kDa proteins disappear in postharvest plasma membranes.
A known protein kinase substrate, Histone III-S, ±Ca²⁺ increases kinase activity in preharvest and postharvest fruit plasma membrane, demonstrating both a stimulatory effect of calcium and a loss of suitable kinase substrate.
H⁺-ATPase is a CDPK substrate (Schaller and Sussman, 1988). Washing plasma membrane with EGTA to remove calcium and calcium-binding proteins decreases plasma membrane kinase activity in preharvest plasma membrane.
In senescent postharvest plasma membrane, a 2.3-fold increase in activity occurs indicating the presence of a unknown kinase inhibitor.
Our study is the first to show that CDPK activity occurs in pre- and postharvest muskmelon plasma membranes.
Even though there is a decline in CDPK activity in postharvest fruit plasma membrane coinciding with a loss in H⁺-ATPase activity, the loss in CDPK activity does not appear due to a loss in protein, but rather to a decrease in a suitable substrate (H⁺-ATPase), a decrease in calcium (Lester, 1996) and an increase in a unknown inhibitor.
Thus, the decline in H⁺-ATPase activity observed in postharvest melon fruit is better explained by a decline in the amount of H⁺-ATPase than by its phosphorylation status.
The beneficial effect of calcium (Lester, 1996) is, most likely, through the maintenance of the structural integrity of the plasma membrane.

International Postharvest Science Conference Postharvest 96

G.E. Lester, V.M. Baizabal-Aguirre, L.E. Gonzalez de la Vara

https://doi.org/10.17660/ActaHortic.1998.464.99