RBSEC1A AND RBSEC1B COLOCALIZE WITH SYNTAXIN-1 AND SNAP-25 THROUGHOUTTHE AXON, BUT ARE NOT IN A STABLE COMPLEX WITH SYNTAXIN

rbSec1 is a mammalian neuronal protein homologous to the yeast SEC1 ge ne product which is required for exocytosis. Mutations in Sec1 homolog ues in the nervous systems of C. elegans and D. melanogaster lead to d efective neurotransmitter secretion. Biochemical studies have shown th at recombinant rbSec1 binds syntaxin 1 but not SNAP-25 or synaptobrevi n/VAMP, the two proteins which together with syntaxin 1 form the synap tic SNARE complex. In this study we have examined the subcellular loca lization of rbSec1 and the degree of interaction between rbSec1 and sy ntaxin 1 in situ. rbsec1, which we show here to be represented by two alternatively spliced isoforms, rbSec1A and B, has a widespread distri bution in the axon and is not restricted to the nerve terminal. This d istribution parallels the localization of syntaxin 1 and SNAP-25 along the entire axonal plasmalemma. rbSec1 is found in a soluble and a mem brane-associated form. Although a pool of rbSec1 is present on the pla smalemma, the majority of membrane-bound rbSec1 is not associated with syntaxin 1. We also show that rbSec1 is not part of the synaptic SNAR E complex or of the syntaxin 1/SNAP-25 complex we show to be present i n non-synaptic regions of the axon. Thus, in spite of biochemical stud ies demonstrating the high affinity interaction of rbSec1 and syntaxin 1, our results indicate that rbSec1 and syntaxin 1 are not stably ass ociated. They also suggest that the function of rbSec1, syntaxin 1, an d SNAP-25 is not restricted to synaptic vesicle exocytosis at the syna pse.