EMBO/soda-nel huggingface dataset:
Sinhcaf-dependent histone deacetylation is essential for primordial germ cell specification (2022)
Figure 4

Figure 4: Loss of Sinhcaf leads to an increase in histone 3 acetylation of FG follicles.

(A, B and C) Co-immunoprecipitation of zebrafish Sinhcaf-GFP with HA-Hdac1 (A), HA-Sin3a (B) or HA-Sap18 (C) in HEK293Tcells transfected with both Sinhcaf-GFP and HA-Hdac1 (A), HA-Sin3a (B) or HA-Sap18 (C) plasmids.

(D) Relative mRNA level of sinhcaf in zebrafish PG, PV, EV, MV and FG follicles was measured using RT-qPCR. Data shown are mean ± SEM (n = 3, biological replicates).

(E and F) Western blots for acetyl-histone H3 (K9) (Ace-H3-K9), histone H3 in WT (E) or MZsinhcaf -/- (F) PG, PV, EV, MV and FG follicles. α-tubulin was used as a loading control.

(G and H) Statistical analysis of the relative levels of Ace-H3-K9 in WT (G) or MZsinhcaf -/- (H) PG, PV, EV, MV and FG follicles. Data shown are mean ± SEM (n = 3, biological replicates). Statistical analysis was performed using one-way ANOVA followed by Tukey's post-hoc test. The different letters (a, b, c) indicate a significant difference between the means, p<0.05.

(I, J and K) Western blots for Ace-H3-K9 and H3 in WT and MZsinhcaf -/- EV (I), MV (J) and FG (K) follicles. α-tubulin was used as a loading control.

(L) Western blots for Ace-H4-K5, Ace-H3-K18 and H3 in WT and MZsinhcaf -/- FG follicles. α-tubulin was used as a loading control.

(M, N and O) Statistical analysis of the relative levels of Ace-H3-K9I n WT and MZsinhcaf -/- EV (M), MV (N) and FG (O) follicles. Data shown are mean ± SEM (n = 3, biological replicates). Statistical analysis was performed using unpaired two-tailed Student's t-test.

(P and Q) Statistical analysis of the relative levels of Ace-H3-K18 (P) and Ace-H4-K5 (Q) in WT and MZsinhcaf -/- FG follicles. Data shown are mean ± SEM (n = 3, biological replicates). Statistical analysis was performed using unpaired two-tailed Student's t-test.

Data information: EV, early vitellogenic stage (early stage III); FG, full-grown stage (late stage III); MV, mid vitellogenic stage (mid stage III); PG, primary growth (stage I); PV, previtellogenic stage (stage II).
Sinhcaf-dependent histone deacetylation is essential for primordial germ cell specification (2022)
Figure 7

Figure 7: The role of Sinhcaf in germ plasm aggregation and subsequent PGC specification is mediated by Kif26ab.

(A) Maximal intensity projection of a confocal z-stack of 24hpf Tg(piwil:egfp), Tg(piwil:egfp) injected with kif26ab MO and Tg(piwil:egfp) injected with control MO embryos.

(B) Quantitative analysis of PGCs in 24hpf Tg(piwil:egfp) (n = 11), Tg(piwil:egfp) injected with kif26ab MO (n = 12) and Tg(piwil:egfp) injected with control MO (n = 13) embryos. Data shown are mean ± SEM (n = the number of embryos analyzed).

(C) Maximal intensity projection of a confocal z-stack of 4-cell stage embryos injected with GFP-Buc mRNA + RFP mRNA, GFP-Buc mRNA + RFP mRNA + Control MO and GFP-Buc mRNA + RFP mRNA + kif26ab MO. White arrowheads point to cleavage furrows of 4-cell embryos.

(D) Relative GFP-Buc density at the cleavage furrow of 4-cell stage embryos injected with GFP-Buc mRNA + RFP mRNA (n = 11), GFP-Buc mRNA + RFP mRNA + Control MO (n = 11) and GFP-Buc mRNA + RFP mRNA + kif26ab MO (n = 10). Data shown are mean ± SEM (n = the number of embryos analyzed).

(E) Statistical analysis of the relative levels of GFP-Buc/a-Tubulin. Data shown are mean ± SEM (n = 3, biological replicates).

(F) Western blots for GFP-Buc expression in 4-cell stage embryos injected with GFP-Buc mRNA + RFP mRNA, GFP-Buc mRNA + RFP mRNA + Control MO and GFP-Buc mRNA + RFP mRNA + kif26ab MO.

(G) Maximal intensity projection of a confocal z-stack of MZsinhcaf -/- or WT 4-cell stage embryos injected with GFP-Buc mRNA + RFP mRNA or GFP-Buc mRNA + RFP mRNA + kif26ab mRNA. White arrowheads point to cleavage furrows of 4-cell embryos.

(H) Relative GFP-Buc density at the cleavage furrow of MZsinhcaf -/- (n = 12) or WT (n = 13) 4-cell stage embryos injected with GFP-Buc mRNA + RFP mRNA or GFP-Buc mRNA + RFP mRNA + kif26ab mRNA. Data shown are mean ± SEM (n = the number of embryos analyzed).

(I) Statistical analysis of the relative levels of GFP-Buc/a-Tubulin. Data shown are mean ± SEM (n = 3, biological replicates).

(J) Western blots for GFP-Buc expression in MZsinhcaf -/- or WT 4-cell stage embryos injected with GFP-Buc mRNA + RFP mRNA, GFP-Buc mRNA + RFP mRNA + Control MO and GFP-Buc mRNA + RFP mRNA + kif26ab MO.

Data information: Statistical analysis was performed using one-way ANOVA followed by Tukey's post-hoc test. Scale bars, 100 μm.
Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy (2019)
Figure 1

Figure 1: IGF2R blockade results in CD20 phosphorylation.

(a) RT-PCR expression and quantification of IGF1, IGF1Rβ and IGF2 levels in untreated C2C12 and shCD20 C2C12 myoblasts and 10 nM IGF1-treated C2C12 and over-expressing CD20 C2C12 myoblasts. Each experiment was replicated independently four times. (b) Immunofluorescence for IGF2 (in green) in untreated and 10 nM IGF1-treated C2C12 myoblasts. Scale bars = 75 μm. Representative CD20 immunoprecipitation using anti-pSer+pThr in (c) C2C12 cells treated with IGF1 for two hours (2 h) or overnight (ON), (d) sh-empty (shCTR)- and shCD20-treated C2C12 cells treated with anti-Flag and 10 nM anti-IGF2R, (e) C2C12 cells treated with IGF1 and IGF2, as indicated. Densitometric analysis of data are expressed as the ratio of CD20/vinculin or pSer+pThr/CD20 and are shown normalized to vinculin in arbitrary units in the lower panels shown in c, d and e. Two-way ANOVA. *p<0.05; **p<0.01; ****p<0.0001. Each experiment was performed in triplicate wells. All values are expressed as the mean ± SEM. (f) Representative WB of IGF2R and phosphorylated IGF2R (pIGF2R) in untreated, shCD20-treated and anti-IGF2R-treated C2C12 cells. Densitometric analysis of data are expressed as the ratio of IGF2R/actin or pIGF2R/IGF2R in arbitrary units in the lower panels. One-way ANOVA. *p<0.05; Each experiment was performed in triplicate wells. All values are expressed as the mean ± SEM. (g) IGF2R immunoprecipitation products were immunoblotted for Gαi2 and WB expression of IGF2R in untreated, IGF2-treated, anti-IGF2R-treated and IGF2+anti-IGF2R treated C2C12 cells. Densitometric analysis of data are expressed as the ratio of Gαi2/IGF2R in arbitrary units in the lower panel. (h) Representative WB of IGF1Rβ and IGF1Rβ immunoprecipitation products immunoblotted for p-Tyr in untreated, IGF2-treated and IGF2+anti-IGF2R-treated (1:500, 1:1000 and 1:2000 dilutions of anti-IGF2R) C2C12 cells (cells were treated for 2 and 24 h). Densitometric analysis of data are expressed as the ratio of pTYR/IGF1Rβ in arbitrary units in the lower panel. Each experiment was performed in triplicate wells. All values are expressed as the mean±SEM.
Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy (2019)
Figure 2

Figure 2: IGF2R binding to CD20 is relevant for myogenic differentiation.

Bioinformatic prediction of IGF2R and CD20 cross-reactivity. Cartoon representation of the interaction of IGF2 (yellow) and IGF2R domain 11 complex from Xray structure (PDB code 2v5p). IGF2R domain 11 AB, CD, EF, HI loops and residues (shown in sticks format) involved in the hydrophobic interactions are shown (a). Cartoon representation of the IGF2R domain 11- CD20 (lime) complex obtained from docking simulations and residues involved in hydrophobic interactions are shown in stick format (b). The structure of IGF2R (in red) and IGF2 (green) are shown compared to docking poses of epitope 3. These data show that the helical region (residues 178-184) of the epitope mediates binding to the receptor as well as IGF2. The C-terminal helix of epitope 3 partially overlaps the first α-helix of IGF2. Cluspro-dock IGF2R-CD20 binding epitope poses corresponding to PDB codes 2v5p-2oslP, 2v5p-2oslQ and 2v5p-3pp4P are coloured in yellow, blue and cyan, respectively (c). (d) Over-expressing CD20 in C2C12 myoblasts that co-expressed CD20 (in green) and IGF2R (in red). DAPI-labelled nuclei are shown in blue. Scale bars=75 μm. (e) Representative CD20 and IGF2R immunoprecipitation products immunoblotted for IGF2R and CD20, respectively, in UNTREATED and shCD20-treated C2C12 cell membranes and whole lysates of proteins. The immunoprecipitation output is shown as IP neg. (f) Myotubes immunofluorescence of cells in proliferation medium (P.M.) and after 2, 4 and 6 days of myogenic differentiation in serum free medium. Control (untreated), shCD20-treated C2C12 cells, C2C12 and HSkM myoblasts pre-treated with anti-IGF2R for 24 hours were stained. Desmin-positive myotubes are shown in green. Scale bars=75 μm. (g) Fusion index quantification after 6 days of differentiation. One-way ANOVA. **p<0.01; ***p<0.001; ****p<0.0001. Each experiment was performed in triplicate wells. All values are expressed as mean±SEM. (h) Representative WB of anti-myogenin, anti-Myf5, anti-MyHC and anti-β-actin in total protein lysates obtained from untreated, shCD20-treated C2C12 cells, and C2C12 and HSkM myoblasts pre-treated with anti-IGF2R for 24 hours; cells were collected under P.M. and after 2, 4 and 6 days of myogenic differentiation. Densitometric analysis of WB data expressed as the ratio of the indicated antibody/β-actin in arbitrary units. Two-way ANOVA test. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001 in comparison to the results obtained in untreated cells at the corresponding time point. ##p<0.01; ###p<0.001; ####p<0.0001 in comparison to the results obtained in shCD20-treated C2C12 cells at the corresponding time point. Each experiment was performed in triplicate wells. All values are expressed as the mean±SEM.
Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy (2019)
Figure 3

Figure 3. IGF2R is overexpressed in dystrophic muscle.

Immunofluorescence analysis of CD20 and IGF2R expression (in red) in the TA muscles of three-month-old C57Bl6/J and mdx mice. Scale bars=75μm. (a). IGF2R (green) and sarcomeric actin (magenta) expression in the VM muscles of two DMD patients. Scale bars=25μm. (b). Representative WB analysis of CD20 and β-actin (c) and pSer+pThr and GADPH (d) expression in the TA and VM muscles of C57Bl6/J and mdx mice (n=10 mice per group). Densitometry analysis of WB data are expressed as the CD20/β-actin ratio, and pSer+pThr/CD20 ratio in arbitrary units in the lower panels of c and d. One-way ANOVA. ****p<0.0001. (e) Representative WB analysis of IGF2R and GAPDH expression in total protein lysates of the human (healthy and DMD) and murine (C57Bl6/J and mdx) TA and VM muscles (n=2 per healthy and DMD; n=10 mice per group); quantifications are expressed as the IGF2R/GAPDH ratio in arbitrary units. (f) Representative WB analysis of IGF2R expression in sarcolemma protein lysates of the TA and VM muscles of healthy, DMD, C57Bl6/J and mdx (n=2 per healthy and DMD; n=10 mice per group). Densitometry analysis of IGF2R expression, results are shown as a fold increase, as indicated in the lower panel in arbitrary units. One-way ANOVA. *p<0.05; ****p<0.0001. All values are expressed as the mean±SEM.
Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy (2019)
Figure 4

Figure 4. Blockade of IGF2R ameliorates muscular dystrophy.

Quantification of the relative frequency of myofibre cross-sectional areas (CSAs) expressed as a frequency distribution in the TA and VM muscles of untreated mdx mice or mdx mice treated with a low or high dose of anti-IGF2R for 4 and 9 weeks (a). (b) Quantification of regenerating myofibres as a proportion of centrally nucleated fibres in the total number of fibres after anti-IGF2R administration. The quantification of regenerating and M-cadherin-positive satellite cells per section is shown. One-way ANOVA test, **p<0.01; ***p<0.001; ****p<0.0001. (c) Representative WB analysis of neonatal myosin heavy chain (neonatal MHC) expression and quantification expressed as the ratio of neonatal MHC/β-actin in untreated mdx mice and mdx mice treated with low and high doses of anti-IGF2R for 4 and 9 weeks. One-way ANOVA. *p<0.05; ***p<0.0001 (n=10 mice per group). (d) Specific force (maximal tetanic force normalized to cross-sectional area, Po/CSA) from TA and VM muscles and SERCA activity quantified as ATPase hydrolysis activity vs. min (e) in C57Bl6/J mice, untreated mdx mice, and mdx mice treated with low and high doses of anti-IGF2R for 4 and 9 weeks. One-way ANOVA. *p<0.05; **p<0.01; ***p<0.001 (n=10 mice per group).
Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy (2019)
Figure 5

Figure 5. Calcineurin signalling activation in muscles of mdx mice treated with anti IGF2R. (a) Representative WB analysis of active 48 kDa and inactive 60 kDa calcineurin A and β-actin expression in the TA and VM muscles in untreated C57BL mice, untreated mdx mice and mdx treated with low and high doses of anti-IGF2R for 4 and 9 weeks. Densitometry analysis of data expressed as ratios of 48 kDa and 60 kDa calcineurin A/β-actin in arbitrary units. Two-way ANOVA. *p<0.05; **p<0.01; ****p<0.0001 (n=10 mice per group). (b) Representative WB analysis of CAMKII, p-CAMKII and GAPDH expression quantified and shown as CAMKII/GAPDH and p-CAMKII/CAMKII ratios in arbitrary units. Two-way ANOVA. *p<0.05; **p<0.01 (n=10 mice per group). (c) Representative WB analysis of PKCα, p-PKCα and β-actin expression levels quantified and expressed as the PKCα/β-actin and p-PKCα/PKCα ratios in arbitrary units (n=10 mice per group). All values are expressed as the mean±SEM.
Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy (2019)
Figure 6

Figure 6. IGF2R blockade results in CD20 and IGF1Rβ phosphorylation in mdx mice.

WB analysis performed in TA and VM muscles of mdx mice treated with anti-IGF2R using antibodies against CD20, pSer+pThr and vinculin. Densitometry analysis of data expressed as the pSer+pThr/CD20 and CD20/vinculin ratio in arbitrary units. One-way ANOVA *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001 for the comparison with untreated mice (n=10 mice per group)(a). Representative WB analysis performed on TA and VM muscles of mdx mice treated with anti-IGF2R using anti-IGF1R, anti-pospho-IGF1R and GAPDH. Densitometry analysis of data expressed as the pIGF1R/IGF1R ratio normalized to GAPDH content in arbitrary units. One-way ANOVA. ****p<0.0001 for the comparison with untreated mice (n=10 mice per group) (b). All values are expressed as the mean±SEM.
Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy (2019)
Figure 7

Figure 7. IGF2R blockade induces normalization of capillary architecture in mdx mice.

Corrosion casting results showing the linearization of capillaries in IGF2R-treated VM mdx muscles. Pericytes around capillaries are shown in the insert box (a). Scale bars=100μm. Representative immunofluorescence results (b) and quantification (c) of longitudinal sections of VM mdx muscles for the CD146- and NG2-positive capillaries are shown. Scale bars=75μm. One-way ANOVA. ****p<0.0001 for the comparison with untreated mice (n=10 mice per group). All values are expressed as the mean± SEM. (d) Representative WB showing CD146 and NG2 expression in total protein lysates of TA and VM muscles of mdx mice treated with anti-IGF2R. Densitometry analysis of data expressed as the ratio of the indicated antibody/β-actin in arbitrary units. One-way ANOVA. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001 for the comparison with untreated mice (n=10 mice per group). All values are expressed as the mean ± SEM. (e) Representative Ve-cadherin (red) and CD31 (green) expression in small vessels of VM muscles from untreated mdx and mdx treated with low dose of anti-IGF2R for 4 weeks. Scale bars=25 μm.

Expanded Figure Legends
The Arabidopsis lncRNA ASCO modulates the transcriptome through interaction with splicing factors (2020)
Figure 1

Figure 1. ASCO modulates steady-state levels of transcripts involved in plant immune responses affecting the sensitivity to flg22 peptide

A Number of differentially up- and down-regulated genes (DEG) in RNAi-ASCO1 seedlings as compared to wild type (WT) according to the RNA-seq data (FDR < 0.01, |log2FC| >= 0.75).

B Fraction of DEG found in each transcript class as defined in the Araport11 gene annotation. AS stands for antisense.

C Enriched Gene Ontology (GO) of DEG in RNAi-ASCO1 seedlings as compared to WT, x-axis represent the -log10 FDR for the enrichment of each GO category over genome frequency.

D Representative picture of 14-day-old plants grown 9 days in liquid 1/2MS supplemented with 1 µM flg22. The scale bar representing 0.6cm is included in the picture.

E Lateral root density of WT and two independent RNAi-ASCO lines 9 days after transfer in 1/2MS supplemented with 0.1 µM or 1 µM flg22.

F Representative picture of root apical meristems after cell wall staining, in response to flg22. TZ: Transition Zone; QC: Quiescent Center.

G Root apical meristem size of WT and RNAi-ASCO1 (eg distance from QC to TZ in µm).

Data information: Error bars indicate the standard error. The asterisk (*) indicates a significant difference as determined by Mann Whitney's U test (p < 0.05, n = 18 biological replicates).
The Arabidopsis lncRNA ASCO modulates the transcriptome through interaction with splicing factors (2020)
Figure 2

Figure 2. ASCO modulates alternative splicing

A Comparison of differentially processed transcripts (RNAprof) and differential AS genes (AtRTD2-SUPPA) with Differentially Expressed Genes (DEG).

B Number of genes containing at least one differential RNA processing event (as defined by RNAprof p.adj < 0.001) in CDS, introns, 5'UTR and 3'UTR between RNAi-ASCO1 and WT. Up and Down fractions correspond to increase or decrease, respectively, of RNA-seq coverage in RNAi-ASCO1 for each specified gene feature.

C Proportion of DAS events identified by AtRTD2-SUPPA in RNAi-ASCO1 compared to WT; Alternative 3' site (A3'), Alternative 5' site (A5'), Intron Retention (IR), Exon skipping (ES).

D Comparison of differentially processed transcripts (RNAprof) and differentially AS genes (AtRTD2-SUPPA) with genes showing significant isoform switch events (Isoswitch).

E Summary of the predicted consequence of the isoform switch events as shown by the feature acquired by the upregulated isoform. ncRNA stands for noncoding RNA, and NMD stands for nonsense mediated decay.

F Comparison of differentially processed transcripts (RNAprof), differentially AS genes (AtRTD2-SUPPA) and differentially expressed genes (DEG) with genes significantly upregulated in the upf1upf3 mutant [56], indicating genes potentially regulated by NMD.
The Arabidopsis lncRNA ASCO modulates the transcriptome through interaction with splicing factors (2020)
Figure 3

Figure 3. RNAi-ASCO and 35S:ASCO lines share common and distinct subsets of DEG and DAS targets

A Overlap between differentially expressed (DEG) and spliced (DAS) genes in RNAi-ASCO and 35S:ASCO as compared to WT.

B Scatter plot showing the respective gene expression fold change in RNAi-ASCO and 35S:ASCO lines as compared to WT. Genes showing significant changes in RNAi-ASCO and 35S:ASCO are highlighted as yellow and blue dots, respectively.

C Scatter plots showing the respective Percent Spliced In difference (dPSI) in RNAi-ASCO and 35S:ASCO lines as compared to WT. Genes showing significant changes in RNAi-ASCO, 35S:ASCO or in both lines are highlighted as red, green or blue dot, respectively. Gray dots represent all AS events.
The Arabidopsis lncRNA ASCO modulates the transcriptome through interaction with splicing factors (2020)
Figure 4

Figure 4. Validation of AS events in RNAi-ASCO lines

A-F (A, D) Differential RNA processing events of SR34 (AT1G02840) and NUDT7 (AT4G12720) transcripts detected by RNAprof from the comparison of RNA-seq libraries of 14-day-old WT (red) and RNAi-ASCO1 (blue) plants. Three biological replicates were used. Vertical purple lines and p-values indicate significant differential processing events. Structure of SR34 (A) and NUDT7 (D) RNA isoforms. Large black boxes indicate exons, narrow black boxes indicate UTRs, black lines indicate introns. Colored boxes indicate protein domains affected by an AS event. RS domain: Arg/Ser-rich domain. Red arrows indicate probes used for gel electrophoresis. Protein domains were retrieved from Uniprot database (https://www.uniprot.org). (B, E) Analyses of RT-PCR products of SR34 (B) and NUDT7 (E) transcripts on 8% acrylamide gel.(C, F) Quantification of the ratio of SR34 (C) and NUDT7 (D) isoforms detected in the gel in (B) and (E) respectively. RNAs were extracted from WT and RNAi-ASCO1 14-day-old plants. The asterisk (*) indicates a significant difference as determined by Student's T test (p < 0.05, n = 3 biological replicates). Error bars show mean +/- standard deviation.
The Arabidopsis lncRNA ASCO modulates the transcriptome through interaction with splicing factors (2020)
Figure 5

Figure 5. ASCO modulates the activity of the spliceosome component PRP8a

A Analysis of ASCO enrichment by ChIRP using two sets of independent biotinylated probes ODD and EVEN compared to negative control with probes designed against the LacZ RNA. The fold enrichment was calculated between ODD or EVEN samples against LacZ. These samples were used for protein precipitation and Mass Spectrometry analyses (ChIRP-MS), from which PRP8a was identified as a potential ASCO partner.

B Validation of PRP8a-ASCO interaction by PRP8a-RIP. U5 RNA was used as a positive control.

C Immunoblot analysis was performed during PRP8a immunoprecipitation (IP). The same volume of input, unbound fraction was loaded as well as 20 % of the eluted IP fraction. α-PRP8a and α-IgG : IP performed with anti-PRP8a or control rabbit IgG antibody, respectively.

D PRP8a recognition of a subset of DAS RNAs is impaired in the 35S:ASCO plants. A housekeeping gene (PP2A, AT1G13320) RNA was used as a negative control.

Data information: In B and D, the results are expressed as a percentage of the Input for PRP8a RIP followed by RT-qPCR, and IgG RIP was used as a negative control. Error bars represent the standard error of 3 biological replicates. N/D stands for non-detectable. The asterisk (*) indicates a significant difference as determined by Student's T test (p < 0.05, n = 3).
The Arabidopsis lncRNA ASCO modulates the transcriptome through interaction with splicing factors (2020)
Figure 6

Figure 6. PRP8a and SmD1b regulate AS of ASCO mRNA targets

A ASCO transcript levels in WT and prp8-7 mutants.

B, C prp8-7 leaky mutant displays similar AS events as observed in RNAi-ASCO. Quantification of SR34 (B) and ESP (C) isoforms splicing index by RT-qPCR.

D ASCO transcript levels in smd1b mutant. In A and D RNAs were extracted from WT, prp8-7 and smd1b 14-day-old plants.

E SmD1b can bind ASCO in vivo. U6 RNA was used as a positive control and a housekeeping gene (HKG2, AT4G26410) RNA as a negative control.

F SmD1b recognizes in vivo the RNAs of 4 genes regulated by ASCO. In E and F the results were expressed as % INPUT in SmD1b-GFP RIP and IgG RIP used as a negative control.

G, H smd1b mutant displays similar AS events as observed in RNAi-ASCO. Quantification of SR34 (G) and ESP (H) isoforms splicing index by RT-qPCR.

Data information: The asterisk (*) indicates a significant difference as determined by Student's T test (p< 0.05, n = 3 biological replicates). Error bars show mean +/- standard error.