Iggo, Richard Derek; (1991) The p68 RNA Helicase. Doctoral thesis (Ph.D), UCL (University College London).

Text The p68 RNA helicase.pdf Download (12MB)

Abstract

The p68 protein is immunologically related to large T antigen, the dominant transforming oncogene of simian virus 40. The deduced amino acid sequence of human p68 contains amino acid sequence motifs characteristic of a large family of putative RNA helicases called "DEAD box" proteins. The family derives its name from the sequence of the most conserved motif: aspartate- glutamate-alanine-aspartate. Human p68 protein was immunoprecipitated from Hela cells and shown to possess RNA-dependent ATPase activity. As expected for an RNA helicase, the ATPase activity is stimulated preferentially by RNA lacking secondary structure. Cell staining shows that the sub-nuclear location of mammalian p68 changes dramatically during the cell cycle. p68 is present in the nucleoplasm and excluded from the nucleoli in interphase. This pattern is reversed in telophase, when p68 appears to be concentrated in prenucleolar bodies. The human p68 cDNA was completed by anchor PCR and the human p68 gene was mapped to chromosome 17q23. Putative p68 homologues were cloned from yeast and named DBP2 in S.cerevisiae and dbp2 in S.pombe. The yeast genes were shown by gene disruption and tetrad analysis to be essential for viability. The p68, DBP2 and dbp2 genes contain an intron at the same site in helicase motif V. The intron is 1.2 kb long in p68, 1 kb long in DBP2, and 700 bp long in dbp2. The unusual length, position and conservation of the intron in DBP2 suggest that it may have a function. The possibility that the intron autoregulates DBP2 expression was examined by Northern and Western blotting in S.cerevisiae.

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