U4 snRNA Nucleolar Localization Requires the NHPX/15.5-kD Protein Binding Site but Not Sm Protein or U6 snRNA Association

• Published in: The Journal of cell biology 2003-09, Vol.162 (5), p.821-832
• Main Authors: Gerbi, Susan A., Borovjagin, Anton V., Odreman, Federico E., Lange, Thilo Sascha
• Format: Article
• Language: English
• Subjects: Animals; Autoantigens; Binding Sites; Cell nucleolus; Cell Nucleolus - metabolism; Coiled Bodies - chemistry; Gene loci; Humans; Microinjections; Nucleic Acid Conformation; Nucleotides; Oocytes; Oocytes - physiology; Perceptual localization; Proteins; Ribonucleic acid; Ribonucleoproteins, Small Nuclear - metabolism; RNA; RNA Caps - metabolism; RNA, Small Nuclear - chemistry; RNA, Small Nuclear - genetics; RNA, Small Nuclear - metabolism; Small nuclear ribonucleoproteins; Small nuclear RNA; Small nucleolar RNA; snRNP Core Proteins; Splicing; Xenopus laevis
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.200301071

All small nuclear RNAs (snRNAs) of the [U4/U6.U5] tri-snRNP localize transiently to nucleoli, as visualized by microscopy after injection of fluorescein-labeled transcripts into Xenopus laevis oocyte nuclei. Here, we demonstrate that these RNAs traffic to nucleoli independently of one another, because U4 snRNA deleted in the U6 base-pairing region still localizes to nucleoli. Furthermore, depletion of endogenous U6 snRNA does not affect nucleolar localization of injected U4 or U5. The wild-type U4 transcripts used here are functional: they exhibit normal nucleocytoplasmic traffic, associate with Sm proteins, form the [U4/U6] di-snRNP, and localize to nucleoli and Cajal bodies. The nucleolar localization element (NoLE) of U4 snRNA was mapped by mutagenesis. Neither the 5′-cap nor the 3′-region of U4, which includes the Sm protein binding site, are essential for nucleolar localization. The only region in U4 snRNA required for nucleolar localization is the 5′-proximal stem loop, which contains the binding site for the NHPX/15.5-kD protein. Even mutation of just five nucleotides, essential for binding this protein, impaired U4 nucleolar localization. Intriguingly, the NHPX/15.5-kD protein also binds the nucleolar localization element of box C/D small nucleolar RNAs, suggesting that this protein might mediate nucleolar localization of several small RNAs.