Recent findings indicate that proteins in the SNARE superfamily are essential for cell signaling, in addition to facilitating vesicle traffic in plant cell homeostasis, growth, and development. We previously identified SNAREs SYP121/Syr1 from tobacco (Nicotiana tabacum) and the Arabidopsis thaliana homolog SYP121 associated with abscisic acid and drought stress. Disrupting tobacco SYP121 function by expressing a dominant-negative Sp2 fragment had severe effects on growth, development, and traffic to the plasma membrane, and it blocked K(+) and Cl(-) channel responses to abscisic acid in guard cells. These observations raise questions about SNARE control in exocytosis and endocytosis of ion channel proteins and their organization within the plane of the membrane. We have used a dual, in vivo tagging strategy with a photoactivatable green fluorescent protein and externally exposed hemagglutinin epitopes to monitor the distribution and trafficking dynamics of the KAT1 K(+) channel transiently expressed in tobacco leaves. KAT1 is localized to the plasma membrane within positionally stable microdomains of approximately 0.5 microm in diameter; delivery of the K(+) channel, but not of the PMA2 H(+)-ATPase, to the plasma membrane is suppressed by Sp2 fragments of tobacco and Arabidopsis SYP121, and Sp2 expression leads to profound changes in KAT1 distribution and mobility within the plane of the plasma membrane. These results offer direct evidence for SNARE-mediated traffic of the K(+) channel and a role in its distribution within subdomains of the plasma membrane, and they implicate a role for SNAREs in positional anchoring of the K(+) channel protein.