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, MMDX28, has been clo ned from a human testis library. This apparently intronless gene was transc ribed in all tissues studied. MDDX28 encodes a protein of 540 amino acids, with similar to 30% homology to other helicases over the core region, conta ining all the conserved DEAD-box helicase motifs. No homologue is known. MD DX28 has RNA and Mg2+-dependent ATPase activity. Subcellular localization s tudies of MDDX28 using oligo-clonal antibodies raised against the protein a s well as its enhanced green fluorescence protein (EGFP) demonstrated that the protein is localized in the mitochondria and the nucleus. To our knowle dge, MDDX28 is the first member of the RNA helicase described with this dua l location. The nuclear localization of MDDX28 depended on active RNA polym erase II transcription, suggesting that the protein could be transported to and from the nucleus. This was confirmed further in an interspecies hetero karyon assay, in which MDDX28 was seen to translocate to the nucleus and mi tochondria. The mitochondrial uptake of the MDDX28-EGFP-N1 fusion protein w as inhibited by carbonyl cyanide p-(trichloro-methoxy)phenylhydrazone. Our results indicate that MDDX28 can be transported between the mitochondria an d the nucleus.