| 2000 |
PHAX (phosphorylated adaptor for RNA export) is an essential factor for U snRNA nuclear export complex assembly in vitro; it directly contacts RNA, the nuclear cap-binding complex (CBC), and the export receptor CRM1/Xpo1, and requires phosphorylation for export complex assembly. Dephosphorylation in the cytoplasm causes export complex disassembly, providing directionality. |
In vitro export complex assembly assay, in vivo knockdown/depletion, identification of phosphorylation state by biochemistry |
Cell |
High |
10786834
|
| 2001 |
The most evolutionarily conserved region of PHAX is a novel RNA-binding domain (RBD) essential for U snRNA export. PHAX also contains two major nuclear localization signals (NLSs) required for nuclear recycling after export, and a CBC interaction domain at least partly distinct from the RBD and NLSs, allowing PHAX to act as a scaffold for U snRNA export complex assembly. |
Systematic mutagenesis/deletion analysis of PHAX domains combined with in vivo export assays and binding assays |
RNA (New York, N.Y.) |
High |
11333016
|
| 2004 |
PHAX binds m7G-capped U3 snoRNA precursors (as well as U8, U13 box C/D snoRNA precursors and telomerase RNA) and is required for transport of U3 snoRNA to Cajal bodies in the first step of intranuclear routing; CRM1 is subsequently required for routing from Cajal bodies to nucleoli. PHAX does not export m7G-capped U3 because its cap becomes hypermethylated in the nucleus. |
Immunoprecipitation of RNA–protein complexes, inactivation of PHAX and CRM1 in vivo, fluorescence microscopy |
Molecular cell |
High |
15574332
|
| 2010 |
The PHAX RNA-binding domain (RNA_GG_bind domain) adopts a novel helical fold and is monomeric in the absence of ligand, only adopting a tertiary structure upon RNA binding; it binds single-stranded RNA with micromolar affinity without sequence specificity. Mutational analysis confirmed that RNA-binding by PHAX-RBD is required for PHAX-mediated nuclear export. |
NMR spectroscopy, X-ray crystallography, mutational analysis of RNA binding |
RNA (New York, N.Y.) |
High |
20430857
|
| 2020 |
PHAX is required for efficient DNA damage response (DDR): PHAX knockdown reduces H2AX mRNA levels by inhibiting both transcription of the H2AX gene and nuclear export of H2AX mRNA (one of the shortest mRNAs), leading to reduced γH2AX and increased cellular sensitivity to DNA damage. |
siRNA knockdown of PHAX, RT-qPCR/Northern blot for H2AX mRNA, γH2AX immunofluorescence, DNA damage sensitivity assays |
RNA (New York, N.Y.) |
Medium |
32759388
|
| 2023 |
The CBC directly interacts with hnRNP C on mRNA, and the tetramer-forming activity and strong RNA-binding activity of hnRNP C together impede PHAX recruitment to longer transcripts, thereby directing them to the mRNA export pathway rather than the U snRNA export pathway. This reveals the molecular mechanism by which length-dependent RNA classification excludes PHAX from long Pol II transcripts. |
Co-immunoprecipitation of CBC–hnRNP C complex on mRNA, RNA-binding and tetramerization mutants of hnRNP C, in vivo export assays |
Nucleic acids research |
Medium |
36620872
|
| 2024 |
The RNA helicase UAP56/DDX39B (and its paralog URH49/DDX39A) stimulates RNA binding of PHAX in an ATP-dependent manner, thereby loading PHAX onto U snRNA and promoting U snRNA export. ALYREF acts as a bridge between PHAX and UAP56/DDX39B. This TREX-component-dependent mechanism for U snRNA export is distinct from the mRNA export mechanism. |
In vitro ATP-dependent factor identification assay, RNA-binding assays, co-immunoprecipitation, in vivo U snRNA export assays |
Nucleic acids research |
High |
39011894
|
| 2024 |
Cryo-EM structure of the complete snRNA export complex (phosphorylated PHAX + CBC + CRM1 + Ran-GTP + capped RNA) reveals that the central region of PHAX bridges CBC-bound capped RNA to CRM1–RanGTP, reinforces cap dinucleotide binding, and contacts CRM1 via a phosphorylated region that engages the basic surface of RanGTP. CBC engagement in this complex is incompatible with ALYREF or NCBP3 interactions, displaces ARS2 from CBC, and synergistic binding of all components is required for export complex formation. |
Cryo-EM structure determination, in vitro mutagenesis, in-cell mutagenesis experiments |
bioRxivpreprint |
High |
bio_10.1101_2024.11.28.625805
|
| 2024 |
PHAX directly binds LIN28B and enhances LIN28B-mediated stabilization of PBX3 mRNA in esophageal cancer cells; PHAX knockdown reduces PBX3 mRNA stability, inhibits cancer cell proliferation, and promotes apoptosis and autophagy in vitro and in vivo. |
Co-immunoprecipitation (PHAX–LIN28B interaction), RNA immunoprecipitation, siRNA knockdown with proliferation/apoptosis/autophagy readouts, mouse xenograft |
Cancer science |
Medium |
39668567
|
| 2024 |
In glioma, HRasV12 activates PHAX, which recruits TRIM24 and Ku-dependent DNA-PKcs to U3 snoRNAs; DNA-PKcs then phosphorylates TRIM24 at S767/768, driving epigenome reprogramming. This places PHAX as a scaffold linking U3 snoRNA to a kinase signaling cascade in oncogenesis. |
Co-immunoprecipitation, RNA immunoprecipitation, phosphorylation site mutagenesis, single-cell RNA-seq, pharmacological inhibition (NU7441) |
Advanced science (Weinheim, Baden-Wurttemberg, Germany) |
Medium |
38828688
|
| 2022 |
In Drosophila, loss of Phax causes widespread alternative splicing changes; genetic suppressor analysis shows that allele-specific alternative splicing (intron retention vs. splicing) of Ribosomal protein S21 (RpS21) can fully suppress larval lethality of Phax mutants, placing RpS21 splicing downstream of Phax function in snRNP biogenesis. |
Genetic epistasis/suppressor analysis in Drosophila, RT-PCR for splicing, transposon mutant characterization |
G3 (Bethesda, Md.) |
Medium |
35920767
|