Affinage

WDR33

pre-mRNA 3' end processing protein WDR33 · UniProt Q9C0J8

Length
1336 aa
Mass
145.9 kDa
Annotated
2026-06-11
27 papers in source corpus 13 papers cited in narrative 14 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

WDR33 is a core subunit of the mammalian polyadenylation specificity factor (mPSF) that directly recognizes the AAUAAA polyadenylation signal and is essential for pre-mRNA 3'-end cleavage and polyadenylation (PMID:25301781). Within a minimal CPSF subcomplex consisting of CPSF160, CPSF30, hFip1, and WDR33—sufficient to reconstitute AAUAAA-dependent polyadenylation—WDR33 makes direct, sequence-specific contacts with the signal RNA both in vitro and in vivo (PMID:25301781). Structural work showed that CPSF160 acts as a scaffold preorganizing WDR33 and CPSF30 around the RNA, with the U3 and A6 bases of AAUAAA forming an intramolecular Hoogsteen base pair that is read out directly by WDR33, while CPSF30 zinc fingers 2 and 3 recognize the flanking adenosines (PMID:29208711, PMID:25301780); an N-terminal lysine/arginine-rich (KR-rich) motif in WDR33 is the critical determinant of specific signal recognition (PMID:29274231). WDR33 and CPSF30 act cooperatively, and only the ternary CPSF160–WDR33–CPSF30 complex achieves high (~3 nM) affinity for AAUAAA, with binary complexes binding far more weakly (PMID:31462423). WDR33 also interacts with RBBP6, which is recruited in an RNA-dependent manner to activate CPSF endonuclease cleavage (PMID:35177536), and the same machinery polyadenylates Pol III-derived SINE transcripts in an AAUAAA-dependent fashion (PMID:39707854). Distinct from its canonical role, non-canonical alternatively polyadenylated WDR33 isoforms (V2/V3) generated within promoter-proximal introns localize to the endoplasmic reticulum and modulate STING-dependent innate immune signaling and autophagy, with isoform choice governed by CFIm25 and intronic splicing efficiency (PMID:38430516, PMID:39327832).

Mechanistic history

Synthesis pass · year-by-year structured walk · 9 steps
  1. 2014 High

    Established that WDR33 is not merely a CPSF accessory subunit but a direct sequence-specific reader of the AAUAAA polyadenylation signal, answering how CPSF recognizes the canonical PAS.

    Evidence in vitro reconstitution of a CPSF subcomplex, UV cross-linking, and transcriptome-wide PAR-CLIP

    PMID:25301781

    Open questions at the time
    • Did not resolve the atomic contacts between WDR33 and individual signal bases
    • Did not define which WDR33 region mediates RNA contact
  2. 2014 High

    Defined the cooperative RNA-contacting partners of WDR33 by showing CPSF30 zinc fingers 2 and 3 also contact AAUAAA and are essential for 3' processing, also explaining how influenza NS1A subverts host polyadenylation.

    Evidence in vitro/in vivo UV cross-linking, zinc-finger mutagenesis, iCLIP, and 3' processing assays

    PMID:25301780

    Open questions at the time
    • Stoichiometry and geometry of joint WDR33/CPSF30 RNA engagement unresolved
    • Did not establish affinity contributions of each subunit
  3. 2014 High

    Reduced the AAUAAA-recognition machinery to a minimal functional module by showing only CPSF160, CPSF30, hFip1 and WDR33 are necessary and sufficient for AAUAAA-dependent polyadenylation, separating signal recognition from the cleavage subunits.

    Evidence reconstitution of recombinant CPSF subcomplex and AAUAAA-dependent polyadenylation assay

    PMID:25301781

    Open questions at the time
    • Did not address how this subcomplex couples to the catalytic cleavage module in vivo
  4. 2017 High

    Provided the structural mechanism of PAS recognition, showing CPSF160 scaffolds WDR33 and CPSF30 and that U3/A6 form a Hoogsteen base pair read by WDR33, and identified the WDR33 KR-rich motif as the specificity determinant.

    Evidence 3.4 Å cryo-EM of CPSF160–WDR33–CPSF30–AAUAAA, plus crystal structure, XL-MS and fluorescence anisotropy binding assays

    PMID:29208711 PMID:29274231

    Open questions at the time
    • Did not quantify the absolute affinity contribution of each subunit
    • Did not capture the assembled cleavage-competent holo-complex
  5. 2019 High

    Quantified the cooperativity established structurally, showing the ternary CPSF160–WDR33–CPSF30 complex binds AAUAAA at ~3 nM whereas binary complexes are much weaker, demonstrating that both WDR33 and CPSF30 are required for high-affinity recognition.

    Evidence fluorescence polarization binding assays with purified proteins and systematic mutagenesis

    PMID:31462423

    Open questions at the time
    • Did not address kinetics or signal-variant discrimination in vivo
  6. 2022 Medium

    Connected the WDR33 recognition module to catalysis by showing RBBP6 binds WDR33 and CPSF73 and activates CPSF endonuclease activity through RNA-dependent recruitment.

    Evidence biochemical reconstitution of endonuclease activity and mutational analysis of RBBP6 interactions

    PMID:35177536

    Open questions at the time
    • WDR33–RBBP6 interaction inferred from mutational/sequence analysis rather than direct structure
    • Structural basis of endonuclease activation not resolved
  7. 2022 High

    Explained how WDR33-containing mPSF accommodates the variant AUUAAA signal, showing CPSF30 ZF2 adapts via U2 hydrogen bonds while WDR33 contacts remain unchanged.

    Evidence cryo-EM of human mPSF (CPSF160–WDR33–CPSF30–Fip1) with AUUAAA

    PMID:36130077

    Open questions at the time
    • Did not survey the full spectrum of non-canonical signals
    • Functional consequences for variant-signal genes not assessed
  8. 2024 Medium

    Extended WDR33 substrate scope beyond Pol II mRNAs by showing it and CPSF30 polyadenylate Pol III-derived SINE transcripts in an AAUAAA-dependent manner without the cleavage subcomplex.

    Evidence siRNA knockdown of CPSF components and Northern hybridization in HeLa cells

    PMID:39707854

    Open questions at the time
    • Mechanism of mPSF recruitment to Pol III transcripts unclear
    • Biological role of SINE polyadenylation not defined
  9. 2024 Medium

    Revealed a polyadenylation-independent function by showing alternatively polyadenylated WDR33 isoforms V2/V3 localize to the ER, bind STING, and modulate interferon-β induction and autophagy, with isoform production controlled by CFIm25 and intronic splicing.

    Evidence subcellular fractionation, Co-IP, isoform-specific knockdown/overexpression, IFN-β reporter and autophagy assays, plus CFIm25 manipulation and splicing/PA-strength assays

    PMID:38430516 PMID:39327832

    Open questions at the time
    • Single-lab functional study without independent replication
    • Direct structural basis of WDR33–STING interaction unknown
    • Physiological relevance in vivo not established

Open questions

Synthesis pass · forward-looking unresolved questions
  • How WDR33's canonical 3'-processing role is integrated with its non-canonical ER/STING isoform functions, and what governs its testis- and stimulation-dependent expression in physiology, remains unresolved.
  • No in vivo genetic model linking WDR33 to organismal phenotype in the corpus
  • Relationship between nuclear polyadenylation and immune isoform functions uncharacterized

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0003723 RNA binding 6 GO:0140098 catalytic activity, acting on RNA 3
Localization
GO:0005634 nucleus 2 GO:0005783 endoplasmic reticulum 1
Pathway
R-HSA-8953854 Metabolism of RNA 4 R-HSA-74160 Gene expression (Transcription) 2
Partners
CPSF160CPSF30CPSF73FIP1L1RBBP6STING1
Complex memberships
CPSFmPSF (CPSF160–WDR33–CPSF30–Fip1)

Evidence

Reading pass · 14 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2014 WDR33 directly contacts the AAUAAA polyadenylation signal RNA in vitro and in vivo, and is required for CPSF binding to AAUAAA-containing RNA; WDR33 can be specifically UV cross-linked to AAUAAA-containing RNAs, and PAR-CLIP showed WDR33 binds in and very close to the AAUAAA signal in vivo with high specificity. In vitro reconstitution of CPSF subcomplex, UV cross-linking, PAR-CLIP, polyadenylation assay Genes & development High 25301781
2014 CPSF30 (CPSF4) and WDR33 directly contact the AAUAAA polyadenylation signal; the CPSF30-RNA interaction is primarily mediated by zinc fingers 2 and 3 and is essential for mRNA 3' processing; the influenza NS1A protein targets these zinc fingers to suppress host mRNA 3' processing. In vitro and in vivo UV cross-linking, mutagenesis of zinc finger domains, iCLIP, mRNA 3' processing assays Genes & development High 25301780
2014 Only four subunits of CPSF—CPSF160, CPSF30, hFip1, and WDR33—are necessary and sufficient to reconstitute a CPSF subcomplex active in AAUAAA-dependent polyadenylation; CPSF100, CPSF73, and symplekin are dispensable for this minimal activity. Reconstitution of recombinant CPSF subcomplex with purified proteins, AAUAAA-dependent polyadenylation assay Genes & development High 25301781
2017 Cryo-EM structure at 3.4 Å of the quaternary complex CPSF-160–WDR33–CPSF-30–AAUAAA RNA revealed that U3 and A6 bases of AAUAAA form an intramolecular Hoogsteen base pair and directly contact WDR33, while A1/A2 are recognized by ZF2 and A4/A5 by ZF3 of CPSF-30; CPSF-160 functions as a scaffold to preorganize CPSF-30 and WDR33 for high-affinity AAUAAA binding. Cryo-electron microscopy structure determination at 3.4 Å resolution Proceedings of the National Academy of Sciences of the United States of America High 29208711
2017 Crystal structure of the CPSF160–WDR33 subcomplex and cross-linking/mass spectrometry defined molecular architecture of the core CPSF complex; an N-terminal lysine/arginine-rich (KR-rich) motif in WDR33 was identified as a critical determinant of specific AAUAAA motif recognition by quantitative RNA-binding assays. Crystal structure determination, cross-linking coupled mass spectrometry (XL-MS), fluorescence anisotropy RNA-binding assays, mutagenesis eLife High 29274231
2019 The CPSF-160–WDR33–CPSF-30 ternary complex binds AAUAAA with ~3 nM affinity; CPSF-30 and WDR33 are both required for high-affinity PAS binding, as their binary complexes with CPSF-160 alone have much lower affinity; mutations of CPSF-30 residues with van der Waals contacts to AAUAAA bases substantially reduce affinity. Fluorescence polarization binding assays with purified recombinant proteins, systematic sequence and mutagenesis analysis RNA (New York, N.Y.) High 31462423
2022 RBBP6 activates human CPSF endonuclease activity and interacts with the WDR33 and CPSF73 subunits of CPSF; unlike its yeast homolog Mpe1, RBBP6 does not co-purify stably with CPSF but is recruited in an RNA-dependent manner. Biochemical reconstitution of endonuclease activity with purified proteins, sequence and mutational analysis of RBBP6-WDR33 and RBBP6-CPSF73 interactions Genes & development Medium 35177536
2022 Cryo-EM structure of human mPSF (CPSF160–WDR33–CPSF30–Fip1) with AUUAAA PAS revealed conformational differences in A1 and U2 nucleotides compared to AAUAAA, with U2 base making two hydrogen bonds with ZF2 of CPSF30, while the WDR33 and remaining nucleotide contact modes are essentially identical to those for AAUAAA. Cryo-electron microscopy structure determination RNA (New York, N.Y.) High 36130077
2024 Non-canonical WDR33 isoforms V2 and V3 (generated by alternative polyadenylation within promoter-proximal introns) localize to the endoplasmic reticulum and interact with STING; V2 suppresses interferon-β induction by preventing STING disulfide oligomerization and promotes autophagy by recruiting WIPI2 isoforms; V3 increases STING protein levels. Subcellular fractionation/localization, co-immunoprecipitation, isoform-specific knockdown/overexpression, interferon-β reporter assays, autophagy assays Cell reports Medium 38430516
2024 WDR33 alternative polyadenylation producing V2 and V3 isoforms is regulated by CFIm25 levels; V2 production is enabled by inefficient splicing of intron 6 allowing weak PA sites to be used; V3 PA site usage is limited by highly efficient splicing of intron 7 and dependency on an alternative 3' splice site. CFIm25 knockdown/overexpression, newly developed splicing and PA site strength assays, isoform quantification RNA biology Medium 39327832
2001 WDR33 (WDC146) protein is localized to the nucleus and its mRNA is most highly expressed in testis, specifically in pachytene spermatocytes as shown by in situ hybridization. Northern blot, in situ hybridization, subcellular localization by immunostaining Biochemical and biophysical research communications Medium 11162572
2019 WDR33 and CPSF4 polyadenylation factors translocate to the nucleus upon macrophage stimulation, and knockdown of WDR33 prevents nuclear localization of NFκB in stimulated macrophages, indicating WDR33 is required for the inflammatory response in macrophages. siRNA knockdown, immunofluorescence localization, NFκB nuclear translocation assay in macrophages Scientific reports Low 30886197
2024 SNORD51 can competitively bind to WDR33 with the 3'UTR of ZBED6 pre-mRNA, thereby inhibiting 3'-end processing of ZBED6 pre-mRNA and reducing ZBED6 mRNA expression in glioblastoma cells. RNA immunoprecipitation, competitive binding assay, knockdown experiments, gene expression analysis Cell death & disease Low 39516455
2024 WDR33 and CPSF30 (mPSF subunits) contribute to polyadenylation of SINE transcripts generated by RNA polymerase III in an AAUAAA-dependent manner, while CPSF100, CPSF73, and symplekin (mCF subcomplex) are not required for this activity. siRNA knockdown of individual CPSF components in HeLa cells, Northern hybridization of SINE transcript polyadenylation Molekuliarnaia biologiia Medium 39707854

Source papers

Stage 0 corpus · 27 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2014 CPSF30 and Wdr33 directly bind to AAUAAA in mammalian mRNA 3' processing. Genes & development 186 25301780
2014 Reconstitution of CPSF active in polyadenylation: recognition of the polyadenylation signal by WDR33. Genes & development 172 25301781
2017 Molecular basis for the recognition of the human AAUAAA polyadenylation signal. Proceedings of the National Academy of Sciences of the United States of America 119 29208711
2021 R-loop resolution promotes co-transcriptional chromatin silencing. Nature communications 96 33741984
2017 Structural insights into the assembly and polyA signal recognition mechanism of the human CPSF complex. eLife 70 29274231
2019 Circulating essential metals and lung cancer: Risk assessment and potential molecular effects. Environment international 57 30991224
2015 Integrated miRNA and mRNA expression profiling of tension force-induced bone formation in periodontal ligament cells. In vitro cellular & developmental biology. Animal 55 26091625
2020 The 3' processing of antisense RNAs physically links to chromatin-based transcriptional control. Proceedings of the National Academy of Sciences of the United States of America 48 32541063
2022 RBBP6 activates the pre-mRNA 3' end processing machinery in humans. Genes & development 47 35177536
2013 3'-End processing of histone pre-mRNAs in Drosophila: U7 snRNP is associated with FLASH and polyadenylation factors. RNA (New York, N.Y.) 45 24145821
2019 Transcriptome Analyses of FY Mutants Reveal Its Role in mRNA Alternative Polyadenylation. The Plant cell 43 31427469
2019 The polyadenylation inhibitor cordycepin reduces pain, inflammation and joint pathology in rodent models of osteoarthritis. Scientific reports 42 30886197
2019 Nuclear eIF4E Stimulates 3'-End Cleavage of Target RNAs. Cell reports 37 31042468
2022 Causal and Candidate Gene Variants in a Large Cohort of Women With Primary Ovarian Insufficiency. The Journal of clinical endocrinology and metabolism 28 34718612
2001 A novel WD40 repeat protein, WDC146, highly expressed during spermatogenesis in a stage-specific manner. Biochemical and biophysical research communications 25 11162572
2019 The role of the protein-RNA recognition code in neurodegeneration. Cellular and molecular life sciences : CMLS 21 30980111
2023 A splicing transcriptome-wide association study identifies novel altered splicing for Alzheimer's disease susceptibility. Neurobiology of disease 17 37354922
2019 Biophysical characterizations of the recognition of the AAUAAA polyadenylation signal. RNA (New York, N.Y.) 16 31462423
2024 Non-canonical isoforms of the mRNA polyadenylation factor WDR33 regulate STING-mediated immune responses. Cell reports 11 38430516
2022 Molecular basis for the recognition of the AUUAAA polyadenylation signal by mPSF. RNA (New York, N.Y.) 5 36130077
2020 Proteomic analysis of Trypanosoma cruzi spliceosome complex. Journal of proteomics 4 32422275
2024 KHDRBS1 regulates the pentose phosphate pathway and malignancy of GBM through SNORD51-mediated polyadenylation of ZBED6 pre-mRNA. Cell death & disease 3 39516455
2024 In vitro evolution and whole genome analysis to study chemotherapy drug resistance in haploid human cells. Scientific reports 2 38886371
2024 WDR33 alternative polyadenylation is dependent on stochastic poly(a) site usage and splicing efficiencies. RNA biology 2 39327832
2024 [Participation of Proteins of the CPSF Complex in Polyadenylation of Transcripts Read by RNA Polymerase III from SINEs]. Molekuliarnaia biologiia 2 39707854
2025 De Novo Missense Variant in Bovine WDR33 Associated With a Complex Syndromic Form of Cleft Palate With Pentalogy of Fallot and Internal Hydrocephalus. Journal of veterinary internal medicine 1 40525707
2024 New biological markers in diagnosis and follow-up of brucellosis cases. Diagnostic microbiology and infectious disease 1 39550977

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