DICTYOSTELIUM VAULTS - DISRUPTION OF THE MAJOR PROTEINS REVEALS GROWTH AND MORPHOLOGICAL DEFECTS AND UNCOVERS A NEW ASSOCIATED PROTEIN

Vaults are large cytoplasmic ribonucleoprotein particles that are high ly conserved in both morphology and protein composition. Protein compo nents of vaults isolated from Dictyostelium discoideum migrate on SDS- polyacrylamide gels as two bands, one at 94 kDa (MvpA) and the other a t 92 kDa (MvpB). An MvpB cDNA clone was isolated from a Dictyostelium expression library. MvpB shares 60% identity with MvpA at the amino ac id level. This cDNA has been used to disrupt the single mvpB gene in b oth wild-type and mvpA(-) genetic backgrounds. Although the mvp(-) mut ant lines are viable, they show that loss of MvpA and/or MvpB interfer es with vault function sufficiently to impede growth under conditions of nutritional stress. The resulting mutant cell lines reach stationar y phase in suspension culture at one-third of the density of wild-type cells. Ovoid structures isolated from mvp(-) single mutant lines repr esent what remains of vaults in these cells. Similar ovoid structures isolated from the mvpA(-) mvpB(-) line copurify with a newly identifie d protein of 92 kDa (MvpC), which lacks cross-reactivity with currentl y available anti-vault antibodies. Our results indicate that the major vault proteins are necessary for optimal cell growth in Dictyostelium and reveal an unanticipated complexity in vault composition.