Nonphosphorylated human La antigen interacts with nucleolin at nucleolar sites involved in rRNA biogenesis.
La is a RNA-binding protein implicated in multiple pathways related to the production of tRNAs, ribosomal proteins, and other components of the translational machinery (D. J. Kenan and J. D. Keene, Nat. Struct. Mol. Biol. 11:303-305, 2004). While most La is phosphorylated and resides in the nucleoplasm, a fraction is in the nucleolus, the site of ribosome production, although the determinants of this localization are incompletely known. In addition to its conserved N-terminal domain, human La harbors a C-terminal domain that contains an atypical RNA recognition motif and a short basic motif (SBM) adjacent to phosphoserine-366. We report that nonphosphorylated La (npLa) is concentrated in nucleolar sites that correspond to the dense fibrillar component that harbors nascent pol I transcripts as well as fibrillarin and nucleolin, which function in early phases of rRNA maturation. Affinity purification and native immunoprecipitation of La and fluorescence resonance energy transfer in the nucleolus reveal close association with nucleolin. Moreover, La lacking the SBM does not localize to nucleoli. Lastly, La exhibits SBM-dependent, phosphorylation-sensitive interaction with nucleolin in a yeast two-hybrid assay. The data suggest that interaction with nucleolin is, at least in part, responsible for nucleolar accumulation of La and that npLa may be involved in ribosome biogenesis.
Intine
Robert V
RV
Laboratory of Molecular Growth Regulation, National Institute of Child Health & Human Development, 6 Center Dr., Rm. 416, Bethesda, MD 20892-2753, USA.
Dundr
Miroslav
M
Vassilev
Alex
A
Schwartz
Elena
E
Zhao
Yingmin
Y
Zhao
Yingxin
Y
Depamphilis
Melvin L
ML
Maraia
Richard J
RJ
D020816:Amino Acid Motifs; D001324:Autoantigens; D002466:Cell Nucleolus / Q000378:metabolism*; D002478:Cells, Cultured; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism*; D010768:Phosphoserine / Q000737:chemistry / Q000378:metabolism; D038761:Photobleaching; D011233:Precipitin Tests; D017434:Protein Structure, Tertiary; D012335:RNA, Ribosomal / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics; D012261:Ribonucleoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques; D000097585:SS-B Antigen; D000097796:Nucleolin
D001324:Autoantigens; D002868:Chromosomal Proteins, Non-Histone; D049452:Green Fluorescent Proteins; D010750:Phosphoproteins; D010768:Phosphoserine; D012335:RNA, Ribosomal; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012261:Ribonucleoproteins; D000097585:SS-B Antigen; D000097796:Nucleolin; C047458:fibrillarin