J. Neurochem. (2012) 121, 861–880.AbstractVesicular transport in neurons plays a vital role in neuronal function and survival. Nesca is a novel protein that we previously identified and herein describe its pattern of expression, subcellular localization and protein–protein interactions both in vitro and in vivo. Specifically, a large proportion of Nesca is in tight association with both actin and microtubule cytoskeletal proteins. Nesca binds to F‐actin, microtubules, βIII and acetylated α‐tubulin, but not neurofilaments or the actin‐binding protein drebrin, in in vitro‐binding assays. Nesca co‐immunoprecipitates with kinesin heavy chain (KIF5B) and kinesin light‐chain motors as well as with the synaptic membrane precursor protein, syntaxin‐1, and is a constituent of the post‐synaptic density. Moreover, in vitro‐binding assays indicate that Nesca directly binds KIF5B, kinesin light‐chain and syntaxin‐1. In contrast, Nesca does not co‐immunoprecipitate with the kinesin motors KIF1B, KIF3A nor does it bind syntaxin‐4 or the synaptosome‐associated protein 25 kDa (SNAP‐25) in vitro. Nesca expression in neurons is highly punctuate, co‐stains with syntaxin‐1, and is found in fractions containing markers of early endosomes and Golgi suggesting that it is involved in vesicular transport. Collectively, these data suggest that Nesca functions as an adapter involved in neuronal vesicular transport including vesicles containing soluble N‐ethylmaleimide sensitive factor attachment protein receptors that are essential to exocytosis.