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Membranes and Bioenergetics

Immunocytochemical Localization of the Vacuolar H⁺-ATPase in Maize Root Tip Cells ¹

Department of Biology, University of California, Santa Cruz, Santa Cruz, California 95064, Thimann Laboratories, University of California, Santa Cruz, Santa Cruz, California 95064

The vacuolar H⁺-ATPase of maize (Zea mays L.) root tip cells has been localized at the EM level using rabbit polyclonal antibodies to the 69 kilodalton subunit and protein A-colloidal gold. Intracellular gold particles were detected mainly on the tonoplast and Golgi membranes. Only about 27% of the vacuoles were labeled above background. The absence of gold particles on the majority of vacuoles suggests either that the tonoplast H⁺-ATPase is degraded during tissue preparation or that the small vacuoles of root tip cells are specialized with respect to H⁺-ATP ase activity. The pattern of gold particles on the labeled vacuoles ranged from uniform to patchy. Virtually all of the Golgi bodies were labeled by the antibody, but the particle densities were too low to determine whether the H⁺-ATPase was associated with specific regions, such as the trans-face. Cell wall-labeling was also observed which could be partially prevented by the inclusion of gelatin as a blocking agent. The immunocytochemical results confirm previous biochemical studies with isolated membrane fractions (A Chanson, L Taiz 1985 Plant Physiol 78: 232-240).

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