The function of the transcription regulator ArgRIII in the expression of several genes involved in the metabolism of arginine in yeast has been well studied. It was previously reported that it is also an inositol phosphate multikinase and an important factor of the mRNA export pathway [reviewed by Shears (2000) Bioessays 22, 786–789]. In the present study we report the cloning of a full-length 1248-bp cDNA encoding a human inositol phosphate multikinase (IPMK). This protein has a calculated molecular mass of 47.219kDa. Functionally important motifs [inositol phosphate-binding site, ATP-binding site, catalytically important SSLL (Ser-Ser-Leu-Leu) domain] are conserved between the human IPMK and yeast ArgRIII. Bacterially expressed protein demonstrated an inositol phosphate multikinase activity similar to that of yeast ArgRIII. Ins(1,4,5)P3 is phosphorylated at positions 3 and 6 up to Ins(1,3,4,5,6)P5. The human IPMK fused with a fluorescent protein tag is localized predominantly in the nucleus when transiently expressed in mammalian cells. A basic cluster in the protein's C-terminus is positively involved in nuclear targeting. These findings are consistent with the concept of a nuclear inositol phosphate signalling and phosphorylation pathway in mammalian cells.
- inositol 1,3,4,5,6-pentakisphosphate
- nuclear localization signal
The sequence of HsIPMK has been deposited with the GenBank®, DDBJ, EMBL and GSDB Nucleotide Sequence Databases under the accession no. AF432853.
Abbreviations used: DAPI, 4′,6-diamidino-2-phenylindole dihydrochloride; DTT, dithiothreitol; EGFP, enhanced green fluorescent protein; EST, expressed sequence tag; EYFP, enhanced yellow fluorescent protein; gi, gene info identifier; HGMPRC, Human Genome Mapping Project Resource Centre; IP, inositol phosphate; IP3K, Ins(1,4,5)P3 3-kinase; IP6K, InsP6 kinase; (Hs or Rn)IPMK, (human or rat) inositol phosphate multikinase; LP, long pass; MADS, MCM1, agamous, deficiens and SRF; MDD, metal dye detection; NLS, nuclear localization signal; SSLL, Ser-Ser-Leu-Leu; tR, retention time.
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