U4 snRNA nucleolar localization requires the NHPX/15.5-kD protein binding site but not Sm protein or U6 snRNA association.
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.
Gerbi
Susan A
SA
Division of Biology and Medicine, Brown University, Providence, RI 02912, USA.
Borovjagin
Anton V
AV
Odreman
Federico E
FE
Lange
Thilo Sascha
TS
D000818:Animals; D001324:Autoantigens; D001665:Binding Sites; D002466:Cell Nucleolus / Q000378:metabolism*; D020541:Coiled Bodies / Q000737:chemistry; D006801:Humans; D008845:Microinjections; D009690:Nucleic Acid Conformation; D009865:Oocytes / Q000502:physiology; D012315:RNA Caps / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism*; D014982:Xenopus laevis; D055517:snRNP Core Proteins
D001324:Autoantigens; D012315:RNA Caps; D012342:RNA, Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear; C402291:Snu13 protein, human; C405482:U4 small nuclear RNA; C405485:U5 small nuclear RNA; C405486:U6 small nuclear RNA; D055517:snRNP Core Proteins