The human nuclear antigen p68 cross reacts with a monoclonal antibody to SV40 large‐T antigen. Its deduced amino acid sequence contains short motifs which place it in a large superfamily of proteins of known or putative helicase activity. Recently, a p68 subfamily (DEAD box proteins) which share more extensive regions of homology has been identified in mouse, Drosophila, Saccharomyces cerevisiae and Escherichia coli. These proteins are involved in translation, ribosome assembly, mitochondrial splicing, spermatogenesis and embryogenesis. We show here that immunopurified human p68 has RNA dependent ATPase activity. In addition, we show that the protein undergoes dramatic changes in cellular location during the cell cycle.

The human p68 protein is an RNA-dependent ATPase and RNA helicase which was first identified because of its immunological cross-reaction with a viral RNA helicase, simian virus 40 large T antigen. It belongs to a recently discovered family of proteins (DEAD box proteins) that share extensive regions of amino acid sequence homology, are ubiquitous in living organisms, and are involved in many aspects of RNA metabolism, including splicing, translation, and ribosome assembly. We have shown by immunofluorescent microscopy that mammalian p68, which is excluded from the nucleoli during interphase, translocates to prenucleolar bodies during telophase. We have cloned 55% identical genes from both Schizosaccharomyces pombe and Saccharomyces cerevisiae and shown that they are essential in both yeasts. The human and yeast genes contain a large intron whose position has been precisely conserved. In S. cerevisiae, the intron is unusual both because of its size and because of its location near the 3' end of the gene. We discuss possible functional roles for such an unusual intron in an RNA helicase gene.Manipulation of RNA secondary structure is essential for the proper execution of a large number of processes in the cell. The importance that the cell attaches to this problem has only recently come to light with the discovery of a large family of proteins with putative ATP-dependent RNA helicase activity. These proteins are readily identified because they all possess a core region of highly conserved motifs in their primary amino acid sequences. One of the most conserved motifs is the DEAD box (LDEADxxL), which gives the family its name (34). DEAD box proteins form a subset of a more loosely defined family, identified by Hodgman (27) and Gorbalenya et al. (21), of proteins with known or putative helicase activity.RNA helicase activity has been demonstrated in vitro for only two DEAD box proteins: p68 (26), in which the DEAD motifs were first identified (18), and eIF4A (44), a translation initiation factor. Although the biological processes in which the other members are involved may prove very diverse, it seems reasonable to expect that all of them will show RNA helicase activity, at least in strand displacement assays in vitro. The family appears to be very large, since DEAD box proteins have been found in all prokaryotes and eukaryotes examined, and even the relatively small genome of Saccharomyces cerevisiae contains genes encoding at least 12 different members We are interested in p68 because of its specific immunological cross-reaction with the simian virus 40 large T antigen (31). We have taken two approaches to determine the function of p68. First, we have used immunofluorescent microscopy and antibody microinjection to study the cell biology of p68 in mammalian cells. Using antibodies to * Corresponding author. t Present address:

The human p68 protein, an SV40 large T related antigen, is an RNA dependent ATPase and RNA helicase. It belongs to a new large and highly conserved gene family, the DEAD box proteins, whose members are involved in a variety of processes requiring manipulation of RNA secondary structure such as translation and splicing. Multiple DEAD box genes are present in S.cerevisiae, but only one has previously been described in E.coli. Low stringency screening of an E.coli genomic library with a p68 cDNA probe led to the identification of dbpA, a new E.coli DEAD box gene located at 29.6 minutes on the W3110 chromosome. We report here the nucleotide and deduced amino acid sequences of the gene. We have overexpressed dbpA from its own promoter on a high copy number plasmid and identified the gene product as a approximately 50 kD protein by immunoblotting with an anti-DEAD antibody.

The human p68 protein is an RNA-dependent ATPase and RNA helicase which was first identified because of its immunological cross-reaction with a viral RNA helicase, simian virus 40 large T antigen. It belongs to a recently discovered family of proteins (DEAD box proteins) that share extensive regions of amino acid sequence homology, are ubiquitous in living organisms, and are involved in many aspects of RNA metabolism, including splicing, translation, and ribosome assembly. We have shown by immunofluorescent microscopy that mammalian p68, which is excluded from the nucleoli during interphase, translocates to prenucleolar bodies during telophase. We have cloned 55% identical genes from both Schizosaccharomyces pombe and Saccharomyces cerevisiae and shown that they are essential in both yeasts. The human and yeast genes contain a large intron whose position has been precisely conserved. In S. cerevisiae, the intron is unusual both because of its size and because of its location near the 3' end of the gene. We discuss possible functional roles for such an unusual intron in an RNA helicase gene.