| 2008 |
Crystal structure of MBNL1 zinc fingers 3 and 4 (ZnF3/4) bound to r(CGCUGU) revealed that both ZnF3 and ZnF4 target GC steps, with site-specific recognition mediated primarily by main-chain hydrogen bonds. The relative alignment of the two ZnF domains is dictated by the interdomain linker topology, resulting in an antiparallel orientation of bound GC elements and a chain-reversal loop trajectory for bound pre-mRNA targets. |
X-ray crystallography (crystal structure of ZnF3/4–RNA complex) |
Nature structural & molecular biology |
High |
19043415
|
| 2006 |
MBNL1 protein translocates from a predominantly cytoplasmic to nuclear distribution during a post-natal window (days 2–20 in mice), and this nuclear localization is required for a synchronized set of developmentally regulated splicing switches in skeletal muscle. In MBNL1 knockout mice these physiological splicing transitions fail, reproducing the splicing defects of DM1. Deficiency of MBNL2 does not reproduce this phenotype, establishing MBNL1 as the non-redundant factor for these post-natal transitions. |
Transgenic mouse model (CUG-repeat expression), MBNL1 knockout mice, RT-PCR splicing assays, immunofluorescence for subcellular localization |
Human molecular genetics |
High |
16717059
|
| 2004 |
MBNL1 is the primary determinant of DM1 nuclear focus integrity and aberrant insulin receptor (IR) exon 11 splicing. siRNA-mediated knockdown of MBNL1 in normal myoblasts recapitulates aberrant IR splicing; rescue experiments in DM1 myoblasts showed that restoring MBNL1 function (but not suppressing CUG-BP) is the key event for correcting splicing. MBNL1 facilitates IR exon 11 inclusion in a dose-dependent manner, antagonized by CUG-BP. |
siRNA knockdown, overexpression rescue experiments, RT-PCR splicing assays in DM1 myoblasts |
The Journal of biological chemistry |
High |
15546872
|
| 2004 |
MBNL1 specifically binds CHHG (where H = A, U, or C) and CHG repeat RNA motifs, including CUG and CCUG repeats, as demonstrated by yeast three-hybrid assays with synthetic RNAs. MBNL1 does not bind canonical double-stranded CUG/CAG RNA, indicating preference for bulge-containing double-stranded RNA. Deletion analysis showed differences in RNA-binding ability among splice variants of MBNL1. |
Yeast three-hybrid assay, RNA-binding assays with synthetic RNAs, deletion mutagenesis |
Human molecular genetics |
High |
14722159
|
| 2010 |
SELEX identified the YGCY (Y = pyrimidine) motif as the preferred RNA-binding element for MBNL1. Insertion of multiple YGCY motifs into an MBNL1-independent splicing reporter was sufficient to confer MBNL1-dependent regulation. MBNL1 regulates ATP2A1 exon 22 splicing through YGCY motifs. YGCY motifs are enriched in positions predicted to cause exon skipping or inclusion consistent with DM1 mis-splicing patterns. |
Doped SELEX, minigene splicing reporter assays, RT-PCR |
Nucleic acids research |
High |
20071745
|
| 2009 |
MBNL1 controls splicing of cardiac troponin T (cTNT) exon 5 by directly competing with the essential splicing factor U2AF65 for binding at the 3' end of intron 4. MBNL1 and U2AF65 bind mutually exclusive RNA secondary structures: MBNL1 binds a stem-loop form while U2AF65 binds the same region in a single-stranded form. When U2AF65 is displaced, U2 snRNP cannot be recruited and the downstream exon is skipped. |
In vitro RNA-binding competition assays, RNA secondary structure probing, splicing assays, mutagenesis |
Proceedings of the National Academy of Sciences of the United States of America |
High |
19470458
|
| 2010 |
MBNL1 promotes insulin receptor exon 11 inclusion by binding directly to a downstream intronic splicing enhancer within intron 11. RNA affinity chromatography identified MBNL1 as the binding factor; RNP immunoprecipitation confirmed in-cell binding to INSR RNA; overexpression or knockdown of Mbnl1 altered exon 11 inclusion levels; deletion of the intronic enhancer abolished MBNL1-mediated regulation. |
RNA affinity chromatography, RNP immunoprecipitation, minigene deletion mutagenesis, overexpression/knockdown |
The Journal of biological chemistry |
High |
20519504
|
| 2011 |
MBNL1 autoregulates its own pre-mRNA by binding within the 3' end of intron 4 to suppress inclusion of exon 5. Structure probing and footprinting showed the MBNL1 response element is primarily unstructured. The branch point is 141 nucleotides from the 3' splice site (non-canonical). Deletion of the MBNL1 response element abolished regulation and led to constitutive exon 5 inclusion. |
Minigene splicing assay, RNA structure probing, MBNL1 footprinting, branch-point mapping, deletion mutagenesis |
The Journal of biological chemistry |
High |
21832083
|
| 2012 |
MBNL1 binds to 3' UTRs genome-wide and promotes accelerated mRNA decay for a broad set of transcripts, a function distinct from and in addition to its role in alternative splicing regulation. Position of MBNL1 binding on pre-mRNA influences whether it promotes exon inclusion or skipping. |
CLIP-seq (in vivo RNA-binding sites), exon array, RT-PCR, mRNA stability analysis |
Scientific reports |
High |
22355723
|
| 2010 |
A systematic analysis of MBNL1 binding to single-stranded RNAs showed that a single GC dinucleotide in a poly-U context is sufficient for MBNL1 binding; a second GC dinucleotide confers higher affinity; additional GC dinucleotides do not further enhance binding. The distance between the two GC dinucleotides can vary from 1 to 17 nucleotides for high-affinity binding, indicating conformational flexibility. Flanking sequence preference: U > C > A > G. |
Fluorescence-based RNA-binding assays, systematic RNA sequence variation |
BMC molecular biology |
Medium |
21548961
|
| 2010 |
Combinatorial mutagenesis of MBNL1 zinc finger domains showed that the two ZnF pairs have differential RNA-binding affinity and distinct splicing activities. Splicing activity profiles of ZnF mutants vary across different pre-mRNA substrates, revealing two distinct classes of MBNL1-regulated splicing events. For some transcripts, robust splicing activity is maintained even in the absence of detectable RNA binding, indicating RNA-binding-independent mechanisms. |
Combinatorial ZnF mutagenesis, splicing reporter assays, RNA-binding assays |
Molecular and cellular biology |
High |
22890842
|
| 2010 |
Functional domains of MBNL1 required for splicing activation and repression are separable from the zinc-finger RNA-binding domains. An 80-amino-acid segment downstream of the N-terminal zinc-finger pair contains core regulatory regions for both splicing activation (IR exon 11) and repression (cTNT exon 5). The MBNL1 response element for IR exon 11 consists of a cluster of three downstream binding sequences. Deletions of these regulatory regions abolished splicing regulation without preventing RNA binding. |
Sequential deletion mapping, minigene splicing reporter assays, RNA-binding assays |
Nucleic acids research |
High |
21109529
|
| 2011 |
MBNL1 colocalizes with YB-1 and DDX1 in cytoplasmic stress granules upon cellular stress. GST pulldown identified YB-1 and DDX1 as MBNL1-binding proteins; MBNL1 forms an RNP complex with these proteins. This provides evidence for a cytoplasmic role of MBNL1 in mRNA metabolism. |
GST pulldown, immunofluorescence colocalization, RNP complex assays |
Journal of neuroscience research |
Medium |
18335541
|
| 2011 |
The RNA helicase p68/DDX5 forms complexes on in vitro-transcribed CUG repeats, colocalizes with nuclear RNA foci in DM1 cells, and increases MBNL1 binding to both pathological CUG repeats and the cTNT pre-mRNA regulatory stem-loop. Mutations in the helicase core of p68 prevented both the stimulatory effect on MBNL1 binding and colocalization with CUG foci, suggesting that p68 remodels RNA secondary structure to facilitate MBNL1 binding. |
In vitro RNA-protein complex assembly, immunofluorescence colocalization, MBNL1 binding assays, helicase-core mutagenesis, splicing reporter assays |
Nucleic acids research |
High |
22156369
|
| 2013 |
MBNL1 binds UGC/CUG clusters in Tpm1 (alpha-tropomyosin) pre-mRNA and cooperates with PTB to repress exon 3 splicing in smooth muscle cells. The N-terminal region of MBNL1 containing all four CCCH zinc fingers is sufficient for repression. MBNL1 makes a direct protein-protein interaction with PTB, and RNA binding by MBNL1 promotes this interaction by inducing a conformational change. Single-molecule analysis showed MBNL1 binding sites increase PTB occupancy at its own sites. |
Minigene splicing reporter, pulldown (protein-protein interaction), single-molecule FRET/TIRF, RNA footprinting |
Nucleic acids research |
High |
23511971
|
| 2014 |
MBNL1 nuclear localization is controlled by two classes of nuclear localization signals (NLS): a classical bipartite NLS and a novel conformational NLS. Alternative splicing of exon 7 acts as a switch between these NLS types and couples MBNL1 splicing activity to intracellular localization. Nuclear MBNL1 promotes nuclear retention of mutant CUG/CAG-repeat-containing RNA and represses expression of homopolymeric proteins produced through RAN translation from these retained RNAs. |
NLS domain mapping, exon 7 splicing manipulation, immunofluorescence/subcellular fractionation, RAN translation reporter assays |
Human molecular genetics |
High |
25274774
|
| 2017 |
MBNL1 exon 1 autoregulation: CLIP-seq revealed MBNL1 binds its own exon 1 (encoding the 5'UTR major portion and amino-terminal region). MBNLs induce skipping of exon 1 from precursor MBNL1 mRNA. Exon 1 exclusion impacts polysome association and translation. The exon 1-deficient protein isoform (lacking the first two zinc fingers) is highly unstable and has severely compromised splicing activity. MBNL1 can be transcribed from three different promoters, and transcription initiation site determines the mode of exon 1 regulation. |
CLIP-seq, minigene/splicing assays, polysome profiling, protein stability assays, promoter analysis |
Nucleic acids research |
High |
27903900
|
| 2018 |
K63-linked polyubiquitination of MBNL1 is required for its cytoplasmic localization and function in promoting neurite outgrowth. Expanded CUG RNA induces deubiquitination of cytoplasmic MBNL1, causing nuclear translocation and morphological impairment. Inhibiting K63-linked polyubiquitin chain degradation ameliorated morphological defects in DM1 neurons. The cytoplasmic (but not nuclear) MBNL1 isoform promotes neurite morphogenesis. |
Ubiquitination assays, subcellular fractionation, immunofluorescence, neurite morphology quantification, pharmacological inhibition of deubiquitination |
Cell reports |
High |
29490267
|
| 2015 |
MBNL1 directly binds to and regulates a network of differentiation-specific transcripts including SRF (serum response factor) and calcineurin Aβ 3'UTRs to promote myofibroblast differentiation. CRISPR-Cas9 editing of the MBNL1-binding site within the Srf 3'UTR impaired myofibroblast differentiation. Loss of Mbnl1 in mice abrogated myofibroblast transformation and impaired fibrotic wound healing after myocardial infarction and dermal injury. |
RNA-IP (direct binding), CRISPR-Cas9 mutagenesis of binding site, mouse KO models (myocardial infarction, dermal injury), genome-wide screen |
Nature communications |
High |
26670661
|
| 2015 |
MBNL1 overexpression increases nuclear retention of full-length expanded HTT (expHTT) RNA and decreases cytoplasmic expHTT protein expression. U2AF65 has the opposite effect, decreasing expHTT nuclear retention and increasing cytoplasmic expHTT protein, suggesting MBNL1 and U2AF65 antagonistically regulate nuclear export of expHTT RNA. |
Overexpression studies, subcellular fractionation, immunofluorescence, western blotting |
Scientific reports |
Medium |
26218986
|
| 2013 |
MBNL1 and RBFOX2 cooperatively control a splicing programme involved in late mesoderm differentiation during reprogramming to iPSCs and redifferentiation. MBNL1 and RBFOX2 co-regulate at least 10 conserved alternative splicing events (including PLOD2, ATP2A1, ITGA6, MARK2, and others), and their combined activity is required for differentiated splicing patterns in vertebrates. |
High-throughput RT-PCR during iPSC induction and redifferentiation, knockdown of individual splicing factors |
Nature communications |
Medium |
24048253
|
| 2014 |
MBNL1 and MBNL2 act synergistically as enhancers of Tau exon 2 inclusion. An intronic region 250 nucleotides downstream of Tau exon 2 contains cis-regulatory splicing enhancers that directly bind MBNL1. Interaction of both MBNL1 and MBNL2 is required to fully reverse Tau exon 2 mis-splicing induced by long CUG repeats. |
Tau minigene reporter assays, MBNL1 direct binding assay, siRNA knockdown, overexpression |
Biochimica et biophysica acta |
Medium |
24440524
|
| 2014 |
MBNL1 knockdown in murine fetal liver erythroid progenitors caused a strong block in erythroid terminal differentiation and disrupted developmentally regulated exon skipping of Ndel1 mRNA. MBNL1 directly binds Ndel1 mRNA (confirmed by CLIP). Ndel1 exon skipping regulated by MBNL1 is critical for erythroid terminal proliferation. |
shRNA knockdown, erythroid differentiation assay, CLIP, RT-PCR splicing analysis |
Blood |
Medium |
24869935
|
| 2017 |
Pseudouridine (Ψ) modification of uridines in MBNL1-binding RNAs inhibits MBNL1 binding by reducing RNA flexibility. Ψ modification of minimally structured YGCY-containing RNA more robustly inhibited MBNL1 binding than modification of CCUG repeats (which can only be modified at one pyrimidine position per motif). Molecular dynamics simulations confirmed that reduced RNA flexibility underlies the reduced binding. |
Fluorescence polarization binding assays, pseudouridine-modified RNA synthesis, molecular dynamics simulations |
The Journal of biological chemistry |
Medium |
28130447
|
| 2018 |
rbFOX1 binds expanded CCUG RNA repeats (DM2) but not expanded CUG repeats (DM1), and competes with MBNL1 for binding to CCUG expanded repeats. Overexpression of rbFOX1 partly releases MBNL1 from CCUG RNA foci in DM2 muscle cells and corrects MBNL1-dependent alternative splicing alterations. |
RNA-binding assays (competition), immunofluorescence (foci), alternative splicing RT-PCR, Drosophila genetic rescue model |
Nature communications |
High |
29789616
|
| 2018 |
MBNL1 splicing isoforms containing exon 7 (ex7) are required for MBNL1 homodimerization. Isoforms lacking ex7 (MBNL1 Δex7) act as dominant negative proteins: splice-switching antisense oligonucleotides inducing ex7 exclusion caused DNA damage and inhibited cell viability and migration in prostate cancer cells. Components of the U2 splicing complex (SF3B1, SF3A1, PHF5A) are required for efficient ex7 inclusion. |
Antisense oligonucleotides for splice switching, siRNA, dimerization assays, viability/migration assays, SF3B1/SF3A1/PHF5A knockdown |
Life science alliance |
Medium |
30456384
|
| 2019 |
TRIM71 represses MBNL1 expression in mouse embryonic stem cells by binding hairpin motifs in the MBNL1 3'UTR, predominantly causing MBNL1 mRNA degradation. Through MBNL1 repression, TRIM71 promotes embryonic (fetal) splicing patterns, thereby regulating stem cell differentiation state. |
RNA-IP (TRIM71-MBNL1 mRNA interaction), TRIM71 loss-of-function, alternative splicing analysis, target mRNA stability assays |
Genes & development |
Medium |
31371437
|
| 2020 |
MBNL1 regulates alternative splicing (predominantly intron exclusion) of DOT1L and SETD1A transcripts in MLL-rearranged leukemia cells. Loss of MBNL1 significantly impairs propagation of murine and human MLL-rearranged leukemia in vitro and in vivo. A small molecule MBNL1 inhibitor selectively kills leukemic cells. |
shRNA knockdown, RNA-seq (transcriptomic profiling), in vivo mouse leukemia model, small molecule inhibitor treatment |
Nature communications |
High |
32398749
|
| 2020 |
MBNL1 inhibits autophagy via the mTOR pathway to promote proliferation of skeletal muscle satellite cells. In DM1 satellite cells, reduced cytoplasmic MBNL1 leads to elevated autophagy and reduced mTOR phosphorylation. MBNL1 overexpression increased mTOR phosphorylation and enhanced proliferative capacity; this effect was abolished by rapamycin (mTOR inhibitor), placing MBNL1 upstream of mTOR in the autophagy-regulatory pathway. |
iPSC-derived satellite cells, TALEN gene editing, MBNL1 overexpression, rapamycin inhibition, western blotting for mTOR phosphorylation, autophagy markers |
Cell death & disease |
Medium |
32683410
|
| 2015 |
MBNL1 binds to the terminal loop of C-allelic pre-miR-1307 at a 'UGCUGC' motif and blocks Dicer processing, resulting in downregulation of mature miR-1307 expression. This reduces miR-1307-mediated repression of its target Bcl2. Both C-allelic and T-allelic pre-miR-1307 are bound by MBNL1. |
RNA-protein binding assays, Dicer processing assay, miRNA expression measurement, luciferase reporter for Bcl2 |
Carcinogenesis |
Medium |
25977444
|
| 2021 |
Loss of MBNL1 induces aberrant splicing of the Abi1 gene, specifically promoting the Abi1-Δe10 isoform in vascular smooth muscle cells. Abi1-Δe10 activates Rac1 independently of upstream stimulation, triggering the Rac1-NOX1-ROS pathway and increasing KLF4 transcription factor expression, which drives VSMC macrophage-like transdifferentiation during atherogenesis. |
siRNA knockdown, overexpression, RT-PCR splicing assay, in vivo atherosclerosis tissue analysis, pathway inhibition assays |
Cell proliferation |
Medium |
33759281
|
| 2023 |
Loss of MBNL1 increases inclusion of Mbnl2 exon 6 and exon 9. Mbnl2 exon 6 inclusion increases MBNL2 nuclear translocation. Mbnl2 exon 9 encodes a PEST domain that targets the protein for proteasomal degradation; exon 9 exclusion (caused by loss of MBNL1) removes this PEST domain and stabilizes MBNL2, providing a compensatory mechanism whereby MBNL1 loss upregulates MBNL2 protein levels. |
RT-PCR splicing analysis, subcellular fractionation, protein stability assays, proteasome inhibition, DM1 mouse model validation |
Nucleic acids research |
High |
36617982
|
| 2024 |
MBNL1 is regulated by the MEIS1/calcineurin signaling axis and stabilizes adult myocyte mRNAs (including cell cycle inhibitor transcripts) to maintain cardiomyocyte maturity and suppress proliferation. Early MBNL1 overexpression prematurely transitions cardiomyocytes to hypertrophic growth and hypoplasia; MBNL1 deletion increases cardiomyocyte cell cycle entry and proliferation through altered cell cycle inhibitor transcript stability. MBNL1 dosage tunes the neonatal cardiac regeneration window. |
Cardiac-specific MBNL1 gain/loss-of-function mouse models, multi-omics, mRNA stability assays, histology, surgical heart regeneration models |
Circulation |
High |
38426339
|
| 2022 |
CDK12 phosphorylates MBNL1, and BUD13 stabilizes CDK12 mRNA via m6A methylation. CDK12-mediated MBNL1 phosphorylation regulates vasculogenic mimicry (VM) formation in glioblastoma cells. Knockdown of BUD13 or CDK12 or overexpression of MBNL1 inhibited VM formation. |
Co-IP, western blotting for phosphorylation, siRNA knockdown, in vitro kinase assay (implied by phosphorylation data), xenograft tumor model |
Cell death & disease |
Medium |
36463205
|
| 2020 |
Loss of MBNL1 in the thymus leads to postnatal thymic hyperplasia with thymocyte accumulation in Mbnl1 129S1 knockout mice. MBNL1 is required for normal thymus development and function, with its loss causing extensive mis-splicing events including TCF/LEF family transcription factor transcripts. |
Mbnl1 knockout mouse model, RNA-seq transcriptome analysis, histology |
Nature communications |
Medium |
32332745
|
| 2018 |
In glioblastoma, hypoxia causes export of MBNL1 from the nucleus, resulting in loss of MBNL1 splicing activity and promotion of fetal/stem-like alternative splicing patterns. Forced expression of a constitutively active (nuclear) MBNL1 isoform inhibited glioma stem cell self-renewal and tumor initiation in orthotopic models and dramatically inhibited tumor progression. |
Subcellular fractionation/immunofluorescence under hypoxia, splicing assays, orthotopic xenograft models, forced MBNL1 expression |
Cancer research |
Medium |
32928918
|