The DEAD box proteins encoded by the genes ddx5 (p68) and ddx17 (isoforms p72 and p82) are more closely related to each other than to any other member of their family. We found that p68 negatively controls p72/p82 gene expression but not vice versa. Knocking down of either gene does not affect cell proliferation, in case of p68 suppression, however, only on condition that p72/p82 overexpression was granted. In contrast, co-silencing of both genes causes perturbation of nucleolar structure and cell death. In mutant studies, the apparently redundant role(s) of p68 and p72/p82 correspond to their ability to catalyze RNA rearrangement rather than RNA unwinding reactions. In search for possible physiological targets of this RNA rearrangement activity it is shown that the nucleolytic cleavage of 32S pre-rRNA is reduced after p68 subfamily knock-down, most probably due to a failure in the structural rearrangement process within the pre-60S ribosomal subunit preceding the processing of 32S pre-rRNA.