The insect V-ATPase, a plasma membrane proton pump energizing secondary active transport: immunological evidence for the occurrence of a V-ATPase in insect ion-transporting epithelia

Active electrogenic K⁺ transport in insects serves as the energy source for secretion or absorption in gastrointestinal epithelia or for the receptor current in sensory epithelia. In the larval midgut of the tobacco hornworm Manduca sexta, a vacuolar-type proton pump (V-ATPase) and a K⁺/nH⁺ antiport represent the functional elements of the potassium pump. Several immunological findings support the hypothesis that active K⁺ transport in other insect epithelia may also be energized by a V-ATPase. In immunoblots, crude homogenates of sensilla-rich antennae and Malpighian tubules of M. sexta cross reacted with an immune serum directed to the purified plasma membrane V-ATPase from the midgut; the M. sexta midgut V-ATPase cross reacted with polyclonal antibodies to endomembrane V-ATPases from xenic origin. In immunocytochemical investigations of larvae of M. sexta and adults of Antheraea pernyi, monoclonal antibodies to defined subunits of the purified midgut V-ATPase or polyclonal antibodies to xenic endomembrane V-ATPase labelled the sites of active K⁺ transport: the goblet cell apical membrane in the midgut, the brush border of Malpighian tubules and the apical projections of the auxiliary cells in antennal sensilla. The functional mechanism of a primary H⁺-pumping V-ATPase and a secondary H⁺-dependent K⁺ transport postulated for K⁺-transporting insect epithelia may be further applicable to active Na⁺ or Cl⁻ transport and would provide a unifying concept for all ouabain-insensitive electrogenic ion transport in insects. The findings from the midgut investigations, however, are the first instance in which a V-ATPase provides an alternative to the Na⁺/K⁺-ATPase in energizing secondary active transport in animal plasma membranes.