Transient Secretory Enzyme Expression in Leaf Protoplasts to Characterize SNARE Functional Classes in Conventional and Unconventional Secretion

Abstract

Despite a long case history, the use of protoplasts in cell biology research still divides scientists but their weaknesses can be exploited as strengths. Transient expression in protoplasts can saturate protein-protein interactions very efficiently, inhibiting the process of interest more efficiently than other approaches at gene expression level. The method described here consists of an assay providing a functional characterization of SNARE proteins in a heterogeneous population of cells, by the comparison of native and dominant negative mutant forms. In particular, it allows for discriminating between t-SNARE and i-SNARE functional classes.

Keywords: Dominant negative mutant; SNARE; Transient expression.