RBM41 (RNA-binding motif protein 41) is an X-linked nuclear RNA-binding protein of the minor (U12-type) spliceosome, with a conserved, largely helical N-terminal region that lacks annotated domains and a single C-terminal RNA recognition motif (RRM). It is the paralog of U11/U12-65K (RNPC3) within the RBM41/RNPC3 family, and its RRM is homologous to the 65K C-terminal RRM. In human cells, the ortholog's RRM binds the 3'-terminal stem-loops of U12 and U6atac snRNAs, and the protein associates with the U12 mono-snRNP rather than the U11/U12 di-snRNP. It also associates with excised U12-type intron lariats and, through its N-terminal region, with the DEAH-box helicase DHX8, and it concentrates in Cajal bodies. These findings place RBM41 in the post-splicing steps of the minor spliceosome assembly/disassembly cycle, where it is thought to replace 65K on the U12 snRNA 3' stem-loop. Loss of RBM41 is tolerated by cultured cells but shifts 3' splice site choice at a subset of U12-type introns. The donkey protein (432 aa) is a full-length ortholog, 95.8% identical to human RBM41 over paired residues, with a 24-residue insertion in the N-terminal region and an RRM that is 78 of 79 residues identical.
Summary: mRNA splicing, via spliceosome (TreeGrafter transfer from the PANTHER PTHR16105 family node).
Reason: Donkey RBM41 is classified in PANTHER subfamily PTHR16105:SF2 (RNA-BINDING PROTEIN 41), the same subfamily as human RBM41 (Q96IZ5), so the human experimental data transfer to it as an ortholog. The PAINT family node behind this term is seeded by the 65K paralogs (RNPC3, Arabidopsis SNRNP65, fly and fish orthologs), and the term holds for the RBM41 branch on its own evidence. Human RBM41 is a specific component of the minor spliceosome, binds U12 snRNA through its RRM, co-precipitates excised U12-type intron lariats and DHX8, and its knockout alters 3' splice site usage at U12-type introns. The protein therefore does part of the work of spliceosomal splicing, in the post-catalytic steps of the minor spliceosome cycle, rather than only being required for it.
Reason: True, since the RRM binds RNA, but uninformative beside the accepted U12 snRNA binding and the proposed U6atac snRNA binding, which both entail it.
Reason: Correct. The human ortholog's RRM binds RNA directly, and alanine substitutions of the conserved Gln352 and Phe354 in the RNP1 motif abolish its snRNA associations. The specific targets are known, however, and U12 snRNA binding (accepted) and U6atac snRNA binding (proposed) both entail this term.
Reason: Human RBM41, in the same PANTHER subfamily (SF2) as the donkey protein, specifically associates with minor spliceosomal snRNAs (U12, U4atac, U6atac, not U11) and with excised U12-type intron lariats, which places it in U12-type spliceosomal complexes. Its association with the U12 mono-snRNP is proposed separately as U12 snRNP.
Reason: This is the core molecular function. The family-level PAINT node rests on the 65K paralog, and the RBM41 branch now has its own direct evidence. The isolated human RBM41 RRM binds the U12 snRNA 3'-terminal stem-loop in EMSA (Kd about 2 uM) but not a complementary control hairpin. In cells, endogenous and tagged RBM41 co-precipitate U12 snRNA in an RRM-dependent way. The donkey RRM is 78 of 79 residues identical to human, including the RNA-contacting Tyr/Gln/Phe triad positions.
Reason: This term comes from the family-level PAINT node (Arabidopsis SNRNP65 among its donors) and not from RBM41 data. For human RBM41, the characterized RNA contacts are the 3' stem-loops of U12 and U6atac snRNAs, and the RRM did not bind a non-cognate control hairpin. RBM41 does co-precipitate excised U12-type intron lariats, but this was shown by immunoprecipitation from cells and is consistent with indirect association through the post-splicing complex. Direct contact with intronic sequence has not been shown. No evidence against inheritance has been found either, so the row is left undecided rather than removed.
ISS PMID:38499487 Distinct functions for the paralogous RBM41 and U11/U12-65K ...
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Summary: Proposed. The human ortholog's RRM binds the U6atac snRNA 3'-terminal stem-loop.
Reason: The isolated human RBM41 RRM binds the U6atac 3' stem-loop with low-micromolar affinity, about half its affinity for U12. U6atac snRNA co-precipitates with RBM41 in cells, and this depends on the RRM. The donkey protein is an SF2 ortholog with an essentially identical RRM. The authors judge the U12 interaction to be the more significant one in vivo, so this term is an additional activity, not the core one.
ISS PMID:38499487 Distinct functions for the paralogous RBM41 and U11/U12-65K ...
NEW
Summary: Proposed. Human RBM41 cosediments with the U12 mono-snRNP, not the U11/U12 di-snRNP.
Reason: In glycerol gradients of HeLa nuclear extract, RBM41 peaks with the U12 mono-snRNP and cosediments with U11/U12-31K. It co-precipitates U12 snRNA but not U11 snRNA. This separates it from its paralog 65K, which sits in the U11/U12 di-snRNP. The donkey protein is in the same PANTHER subfamily as human RBM41.
ISS PMID:38499487 Distinct functions for the paralogous RBM41 and U11/U12-65K ...
NEW
Summary: Proposed. Endogenous human RBM41 localizes to Cajal bodies.
Reason: Endogenous RBM41 colocalizes with coilin-GFP in HEK293 nuclear bodies, and coilin is a major RBM41 BioID partner. Cajal body localization depends on the RRM, consistent with RBM41 entering Cajal bodies as part of a U12 snRNA-containing particle. This holds for the SF2 ortholog of the donkey protein.
This suggests that RBM41 localizes to Cajal bodies in an RNA binding-dependent manner.
Core Functions
RRM-containing protein of the minor (U12-type) spliceosome. Its C-terminal RRM binds the 3'-terminal stem-loop of U12 snRNA, and it associates with the U12 mono-snRNP and post-splicing complexes containing excised U12-type intron lariats and DHX8. It is concentrated in Cajal bodies and acts in the post-catalytic phase of U12-type intron splicing.
Q: Does RBM41 contact intronic pre-mRNA directly in post-splicing complexes, or does its lariat association go entirely through U12 snRNA and DHX8? The answer decides whether the inherited pre-mRNA intronic binding term holds for the RBM41 branch.
Q: At what step is U11/U12-65K exchanged for RBM41 on the U12 snRNA 3' stem-loop, and does RBM41 recruit or activate DHX8 to release spliced mRNA from minor spliceosomes?
Suggested Experiments
Experiment: Crosslinking immunoprecipitation (CLIP/eCLIP) of RBM41 to map its direct RNA contacts on U12 and U6atac snRNAs and on U12-type intron lariats.
Experiment: In vitro minor-spliceosome disassembly assays with RBM41-depleted extracts, with or without recombinant RBM41 or an RBM41 N-terminal mutant that cannot bind DHX8, to test whether RBM41 is needed for mRNA release or lariat-intron complex disassembly.
External Prediction Reviews
These computational predictions are reviewed separately from the GOA annotation set used for this review. The assessments below are from this project and do not constitute official GO annotations or endorsement by GO/UniProt. They are not included in the existing annotation review above.
Assessment of the single ProtNLM2 GO prediction for donkey RBM41 (A0A9L0IK51), an RRM protein of the RBM41/RNPC3 family (PANTHER PTHR16105:SF2, the same subfamily as human RBM41). RNA binding is correct, supported by direct RRM-snRNA binding experiments on the human ortholog, but less precise than the U12 snRNA binding already in donkey GOA and accepted in the gene review (LSP). ProtNLM2 also emitted a subcellular location of Nucleus (not scored); this is consistent with the human ortholog's association with minor spliceosomal snRNPs and Cajal bodies. No function text was emitted.
LSP β Less precise than existing annotation Review score: 2/2
Prediction method: ProtNLM2 Β· Version: UniProt API snapshot 2026-10-14
Review rationale: RNA binding is directly supported for the human ortholog, which sits in the same PANTHER subfamily (PTHR16105:SF2): the isolated human RBM41 RRM binds the U12 and U6atac snRNA 3' hairpins with low-micromolar affinity, and RNP1 point mutations or RRM deletion abolish its snRNA associations in cells. The donkey RRM is 78 of 79 residues identical to the human RRM, so the transfer is well justified. The exact term is already in donkey GOA, and donkey GOA also carries the more specific U12 snRNA binding (TreeGrafter), which the gene review accepts as the core function while marking RNA binding as too general. Because a supported, more specific annotation exists, the prediction is correct but less precise (LSP).
Supporting Evidence:
PMID:38499487: "the RBM41 RRM has a low micromolar affinity to both U12 (Kd= 2.14 ΞΌM) and U6atac hairpins (Kd= 3.78 ΞΌM; Figure 2D)"
PMID:38499487: "The Q352A and F354A mutations and deletion of the RRM led to dramatic loss of the U12, U4atac and U6atac interactions"