Large Deletions in Class III Gas Vesicle-Deficient Mutants of Halobacterium halobium

Mutants of Halobacterium halobium deficient in production of gas vesicles (Vac^- and Vac^δ-) arise spontaneously at an extremely high frequency, approximately 1%. In this study, we characterized one class of mutants (class III) with a stable and completely Vac^- phenotype. Restriction mapping and Southern analysis of four class III mutants, SD109, SD112A, SD116, and SD118A, demonstrated that the mutants were deleted for the gas vesicle gene cluster and flanking sequences in the 200-kb gas vesicle plasmid, pNRC 100. Deletions resulted from recombination between pNRC 100 fragments HindIII-C and HindIII-D″ (for SD109, SD112A, and SD116) or HindIII-C and an unidentified sequence (for SD118A). In at least three cases, the deletion junctions mapped precisely to the termini of IS elements, ISH2 (for SD112A) and ISH8 (for SD109 and SD118A). These results suggest that deletion formation is a consequence of intramolecular transposition of IS elements, although a multistep mechanism involving recombination between two elements is also possible. Interestingly, a Vac⁺ revenant (SD142R) of a class III mutant (SD142) was isolated from a sectored colony. Southern analysis indicated that SD142 had a deletion of the gvp gene cluster and that SD142R had regained the gas vesicle gene cluster by an unknown mechanism.