Plasma membrane-enriched samples were extracted from pepino fruit (cv.
El Camino) by phase partitioning. H+-ATPase (EC 3.6.1.35) activity in
these samples increased during late fruit development (immediately be
fore the onset of ripening) and western blotting confirmed there was a
n increase in enzyme abundance at this time. H+-ATPase activity decrea
sed during early ripening and then increased again in the final phase
of ripening. Immunolocalisation showed the plasma membrane H+-ATPase w
as most abundant in the outer cell layers of the fruit, which are cons
idered to have a major role in determining fruit texture. Fruit soften
ing was not accelerated by harvest and there was no stimulation of H+-
ATPase activity by harvest. An in vitro tensile test using fruit rings
showed tissue softening proceeded faster at low apoplastic pH (4.5) t
han pH 6.5; and tissue buffered at pH 6.5 softened less than unbuffere
d rings. Erythrosin B, an inhibitor of the plasma membrane H+-ATPase,
also retarded softening in vitro. These data suggest that plasma membr
ane H+-ATPase activity may contribute to the onset of pepino softening
through a reduction in apoplastic pH.