Effects of Ga and Ca on barley aleurone protoplasts: a freeze-fracture study.

Freeze-fracture electron microscopy was used to study changes in the endomembrane system of barley ( Hordeum vulgare L. cv. Himalaya) aleurone protoplasts. Protoplasts were used for this study because their response to calcium and the plant hormone gibberellic acid (Ga) can be monitored prior to rapid freezing of cells for electron microscopy. Protoplasts incubated in Ga plus Ca secrete elevated levels of a-amylase relative to cells incubated in Ga or Ca alone. The endoplasmic reticulum (ER) and Golgi apparatus of protoplasts incubated in Ga plus Ca undergo changes that are well correlated with the synthesis and secretion of a-amylase. The ER, which appears as short, single sheets of membrane in Ca-and Ga-treated protoplasts, exists as a series of long fenestrated stacks of membranes following incubation in Ga plus Ca. The Golgi apparatus is also more highly developed in protoplasts treated with Ga plus Ca. This organelle is larger and has more vesicles associated with its periphery in protoplasts that actively secrete a-amylase. Evidence that the Golgi apparatus participates in a-amylase secretion is also provided by experiments with the ionophore monensin, which causes pronounced swelling of Golgi cisternae and inhibits the secretion of a-amylase. We interpret these observations as showing that the ER and Golgi apparatus of barley aleurone participate in the intracellular transport and secretion of a-amylase. The plasmalemma (PF face) of barley aleurone protoplasts shows a high density of intramembranous particles (IMPs) which, in general, are evenly distributed. Occasionally, ordered arrays of IMPs are observed, possibly resulting fro m osmotic stress. after 48 hours the plasmalemma of some Ga-treated protoplasts show particle-free areas considered to be indications of senescence. [ABSTRACT FROM AUTHOR]