A compartmentalized phosphorylation/dephosphorylation system that regulates U snRNA export from the nucleus.
PHAX (phosphorylated adaptor for RNA export) is the key regulator of U snRNA nuclear export in metazoa. Our previous work revealed that PHAX is phosphorylated in the nucleus and is exported as a component of the U snRNA export complex to the cytoplasm, where it is dephosphorylated (M. Ohno, A. Segref, A. Bachi, M. Wilm, and I. W. Mattaj, Cell 101:187-198, 2000). PHAX phosphorylation is essential for export complex assembly, whereas its dephosphorylation causes export complex disassembly. Thus, PHAX is subject to a compartmentalized phosphorylation/dephosphorylation cycle that contributes to transport directionality. However, neither essential PHAX phosphorylation sites nor the modifying enzymes that contribute to the compartmentalized system have been identified. Here, we identify PHAX phosphorylation sites that are necessary and sufficient for U snRNA export. Mutation of the phosphorylation sites inhibited U snRNA export in a dominant-negative way. We also show, by both biochemical and RNA interference knockdown experiments, that the nuclear kinase and the cytoplasmic phosphatase for PHAX are CK2 kinase and protein phosphatase 2A, respectively. Our results reveal the composition of the compartmentalized phosphorylation/dephosphorylation system that regulates U snRNA export. This finding was surprising in that such a specific system for U snRNA export regulation is composed of two such universal regulators, suggesting that this compartmentalized system is used more broadly for gene expression regulation.
Kitao
Saori
S
Institute for Virus Research, Kyoto University, Shogoin Kawahara-cho, Sakyo-ku, Kyoto 606-8507, Japan.
Segref
Alexandra
A
Kast
Juergen
J
Wilm
Matthias
M
Mattaj
Iain W
IW
Ohno
Mutsuhito
M
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D047390:Casein Kinase II / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D029543:Nucleocytoplasmic Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D012342:RNA, Small Nuclear / Q000378:metabolism*
D029543:Nucleocytoplasmic Transport Proteins; C498688:PHAX protein, human; D010750:Phosphoproteins; D012342:RNA, Small Nuclear; D047390:Casein Kinase II; D010744:Phosphoric Monoester Hydrolases