Cloning and characterization of MDDX28, a putative dead-box helicase with mitochondrial and nuclear localization.

A cDNA encoding a novel member of the helicase family, MDDX28, has been cloned from a human testis library. This apparently intronless gene was transcribed in all tissues studied. MDDX28 encodes a protein of 540 amino acids, with approximately 30% homology to other helicases over the core region, containing all the conserved DEAD-box helicase motifs. No homologue is known. MDDX28 has RNA and Mg(2+)-dependent ATPase activity. Subcellular localization studies of MDDX28 using oligoclonal antibodies raised against the protein as well as its enhanced green fluorescence protein (EGFP) demonstrated that the protein is localized in the mitochondria and the nucleus. To our knowledge, MDDX28 is the first member of the RNA helicase described with this dual location. The nuclear localization of MDDX28 depended on active RNA polymerase II transcription, suggesting that the protein could be transported to and from the nucleus. This was confirmed further in an interspecies heterokaryon assay, in which MDDX28 was seen to translocate to the nucleus and mitochondria. The mitochondrial uptake of the MDDX28-EGFP-N1 fusion protein was inhibited by carbonyl cyanide p-(trichloromethoxy)phenylhydrazone. Our results indicate that MDDX28 can be transported between the mitochondria and the nucleus.