Exocytotic mutants can be obtained in Paramecium thataffect the organization of the fusion machinery, visible byelectron microscopy. The site of action of the genes in theplasma membrane, cytosol or secretory compartmentcan easily be determined in such mutants. Functionalcomplementation cloning of exocytotic mutants specific-ally affected in the secretory compartment, nd2-1 andnd169-1, reported here, and the previously studied nd7-1,led to the discovery of a set of novel proteins that displayPSI and EGF domains, normally found in extracellularmatrix proteins and involved in transmembrane signal-ing. The structure of one of these proteins, Nd2p, and ofthe product of a paralog found in the genome Nd22p,corresponds to that of type I membrane receptors,generally involved in protein and vesicle sorting. Ourcharacterization suggests that the proteins we haveidentified are required to indicate the presence of amature secretory vesicle to the plasma membrane, toprepare the machinery for fusion. We propose to namethis novel subclass of receptors VEMIF, for VesicularExtracellular-Matrix-like proteins Involved in preparingmembrane Fusion.