title: Phosphorylation and subcellular redistribution of high mobility group proteins 14 and 17, analyzed by mass spectrometry.
abstract: High mobility group (HMG) proteins 14 and 17 are nonhistone nuclear proteins that have been implicated in control of transcription and chromatin structure. To examine the posttranslational modifications of HMG-14 and -17 in vivo, HMG proteins were prepared from nuclear vs. cytosolic fractions of human K562 cells treated with 12-O-tetradecanoylphorbol 13-acetate (TPA) or okadaic acid (OA) and examined by electrospray mass spectrometry. Analysis of full-length masses demonstrated mono-, di-, and triphosphorylation of HMG-14 and mono- and diphosphorylation of HMG-17 from OA treated cells, whereas HMG-14 and -17 from TPA treated cells were monophosphorylated. Peptide mass and sequence analysis showed major and minor phosphorylation sites, respectively, at Ser24 and Ser28 in HMG-17, and Ser20 and Ser24 in HMG-14. These sites were found in the consensus sequence RRSARLSAK, within the nucleosomal binding domain of each protein. A third phosphorylation site in HMG-14 was located at either Ser6 or Ser7. Interestingly, the proportion of HMG-14 and -17 found in cytosolic pools increased significantly after 1 h of treatment compared to control cells and showed preferential phosphorylation compared with proteins from nuclear fractions. These results suggest that phosphorylation of HMG-14 and -7 interferes with nuclear localization mechanisms in a manner favoring release from nuclei.
authors.affiliation: Howard Hughes Medical Institute, University of Colorado, Boulder 80309, USA.
authors.forename: D F
authors.initials: DF
authors.lastname: Louie
authors.forename: K K
authors.initials: KK
authors.lastname: Gloor
authors.forename: S C
authors.initials: SC
authors.lastname: Galasinski
authors.forename: K A
authors.initials: KA
authors.lastname: Resing
authors.forename: N G
authors.initials: NG
authors.lastname: Ahn
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003600:Cytosol / Q000378:metabolism; D006609:High Mobility Group Proteins / Q000378:metabolism*; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D013347:Subcellular Fractions / Q000378:metabolism*
chemical_list: D006609:High Mobility Group Proteins