EMBO/soda-nel huggingface dataset:
CPEB2-dependent translation of long 3'-UTR Ucp1 mRNA promotes thermogenesis in brown adipose tissue (2018)
Figure 3

Figure 3. Adrenergic signaling enhances CPEB2-promoted polyadenylation and translation of Ucp1L RNA.

Northern blotting of Ucp1L and Ucp1S in BAT from CPEB2-WT and -KO male mice at room temperature or after 5-h cold exposure (CE) at 4oC (5 mice per genotype, 4-7 mo old).

RNA immunoprecipitation (RNA-IP). BAT isolated from mice at 10 min after intraperitoneal (i.p.) injection of PBS or CL316243 was used for IP with control (Ctrl) or CPEB2 (CP2) IgGs. RT-qPCR analysis of precipitated substances for level of Ucp1 mRNA with the non-target control, Gapdh RNA, as the reference. Data are mean ± SD (3-4-mo-old male mice, n = 3 per group). **p < 0.01, Student's t test.

PAT assay to monitor the poly(A) tail length of Ucp1L RNA in BAT isolated from 4-mo-old WT and KO mice at 10 min after i.p. injection of PBS or CL316243. As a poly(A)- control, the oligo(dT)-annealed total RNAs were treated with RNaseH to remove poly(A) tails before PAT assay.

Dual luciferase reporter assay. The reporter plasmids, Fluc-mUcp1S, -mUcp1L, -mUcp1LΔHex or -mUcp1LmutCPE (the 2 CPEs were mutated to UUUUGGC and UUUGCC) 3'-UTR, and Rluc were co-transfected with the plasmid expressing EGFP, or myc-tagged full-length (CP2) or RNA-binding domain (RBD) of CPEB2 into differentiated HIB1B adipocytes. Cells were treated ± CL-316243 (CL) before harvesting for luciferase assay. Data are mean ± SD from 3 independent experiments. ap < 0.05 compared with EGFP, bp < 0.05 compared with RBD, cp < 0.05 compared with CP2, one-way ANOVA.

Representative polysome profiles from CPEB2-WT and -KO BAT. RT-qPCR analysis of the polysomal distribution of Ucp1L mRNA with RNA isolated from each fraction.
CPEB2-dependent translation of long 3'-UTR Ucp1 mRNA promotes thermogenesis in brown adipose tissue (2018)
Figure 4

Figure 4. CPEB2-KO mice with decreased protein but not RNA level of UCP1 have enlarged brown adipocytes.

Western blot analysis and RT-qPCR analysis of protein and mRNA levels of UCP1 and GAPDH in WT and KO male BAT (13 mo old, n = 3-4 per group).

Western blot analysis of UCP1 and GAPDH in BAT lysates from WT and KO female mice at 4 and 12 mo old (n = 3 per group). Protein levels were normalized to that of GAPDH.

RT-qPCR analysis of Ucp1 mRNA level in BAT collected from (B) relative to Gapdh mRNA level.

Hematoxylin and eosin sections of BAT from WT and CPEB2-KO female mice at 4 and 12 mo old. Scale bars, 1 mm.

Lipid droplet size and cell number (i.e., number of nuclei) in WT and KO BAT (representative images shown in 4D) were quantified and displayed as arbitrary units. Three tissue sections from each BAT and 3 mice per group were analyzed.

Weight and representative images of WT and KO BAT. Three mice per group were analyzed.

Data information: All data are mean ± SD. *p < 0.05, **p < 0.01, Student's t test.
CPEB2-dependent translation of long 3'-UTR Ucp1 mRNA promotes thermogenesis in brown adipose tissue (2018)
Figure 6

Figure 6. Ectopic expression of CPEB2 in the KO BAT increases UCP1 synthesis and improves thermogenesis.

CPEB2-WT and -KO mice were transduced with adeno-associated virus (AAV) expressing full-length (AAV_CP2) or RNA-binding domain (AAV_RBD) of CPEB2. Immunostaining of CPEB2 and UCP1 in KO BAT transduced with AAV_CP2 for 2 weeks, with Hoechst staining of nuclei. Scale bar, 10 µm. See more images in Fig EV4D.

Western blot analysis of UCP1 in BAT lysates from WT and KO male mice 2 weeks after AAV transduction (n = 6 mice per group). *p < 0.05, Student's t test.

BAT temperature of WT and KO male mice after 2 weeks of AAV transduction measured during 12-h light time (n = 7 mice per group). *p < 0.05, Student's t test.

BAT temperature in WT and KO male mice, and of AAV_CP2-transduced WT and KO male mice (n = 7 mice per group) before and after i.p. injection of CL316243 (CL). * p < 0.05, repeated-measures two-way ANOVA.

CPEB2 activates polyadenylation-induced translation of mouse Ucp1 mRNA with long 3'-UTR via β3AR signaling for adaptive thermogenesis in BAT. Mice lacking the translational regulator CPEB2 show impaired BAT thermogenesis.

Data information: All data are mean ± SEM.
Synaptogenic gene therapy with FGF22 improves circuit plasticity and functional recovery following spinal cord injury (2022)
Figure 1

Fig. 1. Targeted FGF22 overexpression in long propriospinal neurons triggers excitatory synapse formation.

(A) Illustration of axonal plasticity of the hindlimb corticospinal tract (hCST; detour circuit formation) following spinal cord injury.

(B) Schematic illustration of the experiment to overexpress FGF22 into long propriospinal neurons (LPSN) and timeline.

(C) Representative confocal image of long propriospinal neurons expressing FGF22 (magenta) in cervical spinal cord.

(D) Viral distribution of control and FGF22 overexpressing AAVs in long propriospinal neurons (LPSN) with number of transduced neurons (n = 4-5 mice per group).

(E) Schematic illustration of the hCST collateral into the cervical cord and confocal images of hCST collaterals (green), stained for vGlut (red) and vGAT (blue) 21 days following spinal cord injury. Insets represent 3D views generated in Imaris of the deconvolved confocal image.

(F) Quantification of bouton density and characterization of bouton between control and FGF22 overexpressing mice (n = 4-5 mice per group).

(G) Schematic illustration and representative confocal image of LPSN viral labeling (magenta) and CST collaterals (green). Insets represent 3D views generated in Imaris of the deconvolved confocal image.

(H) Quantification of hCST contacts onto LPSNs and characterization of vGlut intensity onto individual LPSNs (n = 4-5 mice per group).

Data information: ns: p>0.05; *P < 0.05 and **P < 0.01. Unpaired t-test for panel D and Mann-Whitney test for panels F, H. The data are presented as means ± SEM. Scale bar for C equals 300 µm; E and G equals 10µm. Inset is magnified ~2 times in C and insets are magnified 4 times in E and G. Arrowheads in C represent labeled LPSNs. Asterisk in E represent individual hCST boutons positive for vGlut staining. Arrowheads in G represent putative contacts between hCST and LPSN.
Synaptogenic gene therapy with FGF22 improves circuit plasticity and functional recovery following spinal cord injury (2022)
Figure 2

Fig. 2. Targeted FGF22 overexpression in cervical excitatory neurons enhances hCST connectivity.

(A) Schematic illustration of the experiment to overexpress FGF22 into excitatory (vGlut2+) cervical neurons and timeline.

(B) Viral distribution of control and FGF22 overexpressing AAVs in excitatory interneurons with number of transduced neurons (n = 5-4 mice per group).

(C) Schematic illustration and representative confocal image for vGlut2-Cre neuronal labeling (cyan) and hCST collaterals (white). Insets represent 3D views generated in Imaris of the confocal image. Quantification of hCST contacts onto vGlut neurons (n = 4-6 mice per group).

Data information: ns: p>0.05; *P < 0.05. Unpaired t-test for panel B and Mann-Whitney test for panel C. Scale bar for C equals 50µm. Insets are magnified ~4 times in C. The data are presented as means ± SEM.
Synaptogenic gene therapy with FGF22 improves circuit plasticity and functional recovery following spinal cord injury (2022)
Figure 3

Fig. 3. Non-selective FGF22 gene therapy promotes widespread circuit rewiring.

(A) Schematic illustration and timeline of the experiment, in which we administered FGF22 before the spinal cord lesion.

(B) Schematic of the location of fluorogold-labeled LPSN in control and FGF22 overexpressing mice. Quantification of the number of fluorogold-labeled LPSN. Each dot represents individual animal (n = 6-7 animals per group).

(C) Schematic illustration of the hCST collateral exiting into the cervical cord and confocal images showing exiting hCST collaterals in control (left panel) and FGF22 treated (right panel) mice and quantification of the number of hCST collaterals that enter the cervical grey matter 3 weeks following spinal cord injury. (n = 6 mice per group).

(D) Quantification of branching points on newly formed cervical hCST collaterals in control and FGF22 treated mice (n = 6 mice per group).

(E) Schematic illustration of the boutons on cervical hCST collateral and confocal images showing putative synaptic boutons (arrows) on newly formed cervical hCST collaterals at 3 weeks following spinal cord injury in a FGF22 (right) and a control (left) mice. Quantification of bouton density in control and FGF22 treated mice. (n = 6 mice per group

(F) Schematic illustration of the hCST contacts onto relay neurons in the cervical cord and confocal images showing contacts between CST collaterals (red) and cervical interneurons (cyan). Insets represent 3D views generated in Imaris of the confocal image. Quantification of the contacts between the hCST collaterals and cervical interneurons neurons. (n = 6 per group).

(G) Confocal images showing contacts between CST collaterals (red) and long propriospinal neurons (magenta) transduced with FGF22 (cyan). Quantification of the contacts between the hCST collaterals and long propriospinal transduced with FGF22 or control virus (DP refers to double positive cells). Insets on the right are magnification of the boxed area on the left. (n = 6 mice per group).

Data information: ns: p>0.05; *P < 0.05 and **P < 0.01. Unpaired t-test for panels E, F and G and Mann-Whitney test for panel D. The data are presented as means ± SEM. Scale bar equals 50µm in panel C; 10µm in panel F, G. Images in E are magnified 5 times from C. Arrowheads in C represent exiting CST collaterals. Arrowheads in D represent bouton on hCST collaterals. Arrowhead in G represents putative contact between hCST collateral and FGF22 overexpressing LPSN.
Synaptogenic gene therapy with FGF22 improves circuit plasticity and functional recovery following spinal cord injury (2022)
Figure 4

Fig. 4. Post-injury therapeutic treatment with AAV-CMV-FGF22-ires-GFP triggers functional recovery in the ladder rung test.

(A) Schematic illustration of the experimental design for therapeutic post-injury treatment with FGF22 following spinal cord injury.

(B) Illustration of regular (left) and irregular (right) ladder rung used to test functional recovery following spinal cord injury.

(C) Timeline of the experiment with FGF22 injection right after the onset of the spinal cord lesion (top). Quantification of the functional recovery in the regular (middle) and irregular (bottom) ladder rung test in controls (grey) and FGF22 (orange) treated mice at baseline, at 3 days post injury ("3dpi"), 14dpi and 21dpi after spinal cord injury. (n = 9-10 mice per group).

(D) Timeline of the experiment with FGF22 injection 24hrs after the onset of the spinal cord lesion (top). Quantification of the functional recovery in the regular (middle) and irregular (bottom) ladder rung test in control (grey) and FGF22 (orange) treated mice at baseline, at 3 days post injury ("3dpi" or 2 days "2dpi for the late treatment), 14dpi and 21dpi after spinal cord injury. (n = 9-10 per group).

(E) Timeline of the experiment with FGF22 injection 5 days after the onset of the spinal cord lesion (top). Quantification of the functional recovery in the regular (middle) and irregular (bottom) ladder rung test in control (grey) and FGF22 (orange) treated mice at baseline, at 3 days post injury ("3dpi"), 14dpi and 21dpi after spinal cord injury. (n = 9-10 per group).).

(F) Representative confocal image of CST boutons (green) and synapsin staining (red) and quantification of the percentage of bouton synapsin positive (n = 6-7 mice per group).

(G) Representative confocal images of CST bouton (green) and bassoon staining (magenta) and quantification of the percentage of boutons that are bassoon positive (n = 5-6 mice per group).

Data information: Insets in F and G represent 3D views generated in Imaris of the confocal images. *P < 0.05 and **P < 0.01. ANOVA with the Šidak's post hoc test in C-E and t-test in F, G. The data are presented as means ± SEM. Scale bars equals 10μm in F, G. Arrowhead in F represents hCST bouton expressing synapsin. Arrowhead in G represents hCST bouton expressing bassoon.
TRIP6 inhibits Hippo signaling in response to tension at adherens junctions (2017)
Figure 1

Figure 1.

TRIP6 promotes YAP activity by inhibiting LATS1/2 (A) Full length (WT), the amino-terminal half (1-277), or the carboxy-terminal half (278-476) of TRIP6 were tested for binding to LATS2 by immunoprecipitation. FLAG-TRIP6 variants were co-expressed with LATS2-GFP in HEK293 cells, anti-FLAG or control (IgG) antibodies were used to isolate immune complexes. Immune complexes and lysates were probed by western blotting for LATS2-GFP and FLAG-TRIP6. Schematic diagram depicts TRIP6 domains (NES: Nuclear Export Signal; LIM: LIM domain; PDZ: PDZ domain binding motif). (B) Lysates from MCF10A cells were subjected to immunoprecipitation using anti-TRIP6 or control (IgG) antibodies, and immune complexes and lysates were probed for TRIP6 and LATS1. (C) FLAG-TRIP6 was tested for binding to various LATS2-GFP deletion mutants as described in part (A). Schematic diagram of LATS2 shows MOB1 binding domain, and the autophosphorylation (S872) and MST1/2 phosphorylation sites (T1041) in the kinase domain. The regions marked in green depict TRIP6 binding sites on LATS2 (D) Lysates from HEK293A cells transfected with control or FLAG-TRIP6 plasmid were analyzed by western blotting using the indicated antibodies (quantification is shown in (F)). (E) Lysates from control (WT) or CRISPR generated TRIP6 null (TRIP6-KO) HEK293A cells were analyzed by western blotting using the indicated antibodies (quantification shown in (G)). (F) The relative levels of LATS1 activating phosphorylation (pLATS1-1079, 909) and YAP S127 inhibitory phosphorylation from (D) were measured relative to LATS1 and YAP levels respectively. (Mean ± SD; n=3; **P≤0.01, ***P≤0.001, T-test) (G) The levels of LATS1 activating phosphorylation and YAP inactivating phosphorylation from part (E) were quantified (Mean ± SD; n=3; *P≤0.05, T-test). (H) GFP-MST2 was expressed with or without FLAG-TRIP6 in HEK293 cells and the levels of MST2, MST2 activating phosphorylation (pMST2-T180), and FLAG-TRIP6 were measured by western blotting with the indicated antibodies. (Mean ± SD; n=3; n.s.≥0.05, T-test). (I) TRIP6 was overexpressed in HEK293A cells and the levels of TRIP6, and YAP target gene expression was analyzed using RT-qPCR. (Mean ± SD; n=3; *P≤0.05, **P≤0.01, ****P≤0.0001, T-test). (J) The levels of YAP target gene expression was analyzed using RT-qPCR in control (WT) and TRIP6-KO HEK293A cells. (Mean ± SD; n=3; ***P≤0.001, T-test) (K) Control (WT) and TRIP6-KO HEK293A cells were stained for YAP and TRIP6. Merged image shows YAP (green), TRIP6 (red), and DNA (blue). Quantification of at least 100 cells is shown (Mean ± SD; n=3; ****P≤0.0001, Fisher's test). Scale bar=20µm.
TRIP6 inhibits Hippo signaling in response to tension at adherens junctions (2017)
Figure 2

Figure 2.

TRIP6 competes with MOB1 for binding to LATS2. (A) LATS2-GFP and Myc-MOB1A were overexpressed in HEK293 cells with or without co-overexpression of full length FLAG-TRIP6 and FLAG-TRIP6 1-277. Myc-MOB1A was immunoprecipitated using anti-Myc antibodies and immune complexes were assayed for Myc-MOB1A and LATS2-GFP levels. Levels of FLAG-TRIP6, FLAG-TRIP6 1-277, Myc-MOB1A, and LATS2-GFP in the lysate are also shown. The levels of LATS2-GFP in immune complexes relative to the level of Myc-MOB1A are shown in the graph (Mean ± SD; n=3; **P≤0.01, T-test) (B) Competitive binding experiments were done using purified recombinant MBP-LATS2, GST-TRIP6, and 6His-MOB1A. MBP-LATS2 bound to maltose beads was incubated with GST-TRIP6 with or without increasing amounts of 6His-MOB1A, and the levels of each protein bound to MBP-LATS2 on the beads at the end of the experiment was determined by western blotting. The levels of input proteins are shown (lysate). The binding of 6His-MOB1A and GST-TRIP6 to MBP alone, and the use BSA as a competitor instead of 6His-MOB1A are shown as controls. The numbers at the bottom are referred to in the text.
TRIP6 inhibits Hippo signaling in response to tension at adherens junctions (2017)
Figure 3

Figure 3.

TRIP6-LATS binding and localization to cell-cell junctions is regulated by tension. (A) MCF10A cells were infected with lentivirus carrying control shRNA (shEGFP), or a mix of two different shRNA against TRIP6 (shTRIP6-1 and shTRIP6-4) and were stained for TRIP6 and LATS1. Merged images show LATS1 (green), TRIP6 (red) and DNA (blue) (quantification of LATS1 localization at cell-cell junctions is shown in Figure EV3D). Scale bar=20µm. (B) LATS1 and LATS2 were knocked down using siLATS1 and siLATS2 SMARTPools in MCF10A cells. MCF10A control cells were treated with control siRNA (siControl) against fire fly luciferase. Cells were stained for TRIP6 and LATS1 as in (A) (quantification of TRIP6 localization at cell-cell junctions is shown in Figure EV3E). Scale bar=20µm. (C) MCF10A cells were grown to high density, and were stained for TRIP6 and LATS1 as in (A). Scale bar=20µm. (D) Cells were grown as in (C), then lysed and anti-TRIP6 antibodies were used to isolate immune complexes. Immune complexes and lysates were probed by western blotting for LATS1 and TRIP6. Quantification is shown. (Mean ± SD; n=3; *P≤0.05, T-test). (E) MCF10A cells grown at high density on PDMS membranes and were stretched (or not) at 17% elongation for 2 hours and lysed while under tension. Anti-TRIP6 antibodies were used to isolate immune complexes. Immune complexes and lysates were probed by western blotting for LATS1 and TRIP6. Quantification is shown. (Mean ± SD; n=3; *P≤0.05, T-test). (F) Cells were treated as in (E), fixed while under tension, and stained for TRIP6 and LATS1 as in (A). Scale bar=20µm. (G) MCF10A cells infected with lentivirus carrying control shRNA (shEGFP), or a mix of two different shRNA against TRIP6 (shTRIP6-1 and shTRIP6-4), were grown at high density on PDMS membranes and were stretched or not (only stretched cells shown) at 17% elongation for 2 hours, fixed while under tension, and were stained for TRIP6 and LATS1 as in (A). Scale bar=20µm. (H) Cells were treated as in (G) and YAP target gene (CTGF and Cyr61) and TRIP6 expression were analyzed using RT-qPCR. (Mean ± SD; n=3; *P≤0.05, **P≤0.01, T-test). (I) TRIP6 expression levels in (H) in control (shEGFP) and TRIP6 (shTRIP6) knockdown cells just prior to stretch were analyzed using RT-qPCR. (Mean ± SD; n=3; *P≤0.05, T-test).
TRIP6 inhibits Hippo signaling in response to tension at adherens junctions (2017)
Figure 4

Figure 4.

Perturbations of junctions and f-actin reduces TRIP6-LATS1 binding and localization to cell-cell junctions. (A) MCF10A cells were either not treated or treated separately with Latrunculin B, Blebbistatin, EGTA, serum starvation, and Y27632, and were stained for TRIP6 and LATS1. Merged images show LATS1 (green), TRIP6 (red) and DNA (blue). Scale bar=20µm. (B) Cells were treated as in (A), then lysed and anti-TRIP6 antibodies were used to isolate immune complexes. Immune complexes and lysates were probed by western blotting for LATS1 and TRIP6. (C) Quantification of (B) is shown. (Mean ± SD; n=3; *P≤0.05, **P≤0.01, T-test).
TRIP6 inhibits Hippo signaling in response to tension at adherens junctions (2017)
Figure 5

Figure 5.

Vinculin interacts with TRIP6, and regulates LATS and YAP activity. (A) MCF10A cells were lysed and anti-TRIP6 or control (IgG) antibodies were used to isolate immune complexes. Immune complexes and lysates were probed by western blotting for vinculin and TRIP6. (B) MCF10A cells were either not treated (control) or treated separately by growth to high density, serum starvation, Latrunculin B, Blebbistatin, EGTA, or Y27632 treatment, and stained using anti-vinculin antibodies by immunofluorescence. Scale bar=20µm. (C) MCF10A cells grown at high density on PDMS membranes and were stretched (or not) at 17% elongation for 2 hours and fixed while under tension and stained for vinculin. Scale bar=20µm. (D) Vinculin was knocked down using two different siRNAs or control siRNA in MCF10A cells and the cell lysates were probed by western blotting for phospho-LATS1 (T1079 and S909), phospho-YAP (S127), LATS1, YAP, vinculin, and tubulin antibodies and the relative levels quantified (Mean ± SD; n=3; **P≤0.01, T-test). (E) Vinculin was knocked down as described in (D) and cells were stained for YAP and vinculin. Merged image shows YAP (green), vinculin (red), and DNA (blue). Quantification of at least 100 cells is shown (Mean ± SD; n=3; ***P≤0.001, Fisher's test). Scale bar=20µm. (F) Vinculin was knocked down as described in (D) and the levels of vinculin and YAP target gene expression was analyzed using RT-qPCR (Mean ± SD; n=3; ***P≤0.001, ****P≤0.0001, T-test). (G) Vinculin was knocked down in MCF10A cells as described in (D), grown at high density on PDMS membranes and cells were stretched (or not) at 17% elongation for 2 hours and the levels of vinculin and YAP target gene expression was analyzed using RT-qPCR (Mean ± SD; n=3; ***P≤0.001, ****P≤0.0001, T-test).
TRIP6 inhibits Hippo signaling in response to tension at adherens junctions (2017)
Figure 6

Figure 6.

Vinculin regulates TRIP6-LATS1 interaction and localization. (A) Vinculin was knocked down as described (Figure 5D) in MCF10A cells and cells were stained for LATS1 and TRIP6. Merged images show LATS1 (green), TRIP6 (red) and DNA (blue). Scale bar=20µm. (B) Vinculin was knocked down as described (Figure 5D) and TRIP6 immune complexes and lysates were probed by western blotting for LATS1, TRIP6, and vinculin and the relative levels quantified (Mean ± SD; n=3; **P≤0.01, T-test). (C) MCF10A cells were subjected to different treatments as described (Figure 4A) and TRIP6 immune complexes and lysates were probed by western blotting for vinculin and TRIP6. (D) Quantification of TRIP6-vinculin binding from part (C) is shown. (Mean ± SD; n=3; *P≤0.05, **P≤0.01, ***P≤0.001, T-test). (E, F) MCF10A cells were treated separately (or not) with Latrunculin B (E) and by serum starvation (F) and LATS1 immune complexes and lysates were probed by western blotting for vinculin, LATS1, and TRIP6. Quantification of LATS1-TRIP6-vinculin binding is shown. (Mean ± SD; n=3; *P≤0.05, **P≤0.01, ***P≤0.001, T-test). (G) Wild type and T12 mutant of vinculin were co-expressed with LATS2 and FLAG-TRIP6 and anti-FLAG antibody was used to isolate immune complexes. Immune complexes and lysates were probed by western blotting for vinculin, LATS2 and FLAG-TRIP6. Quantification of LATS2-TRIP6-vinculin (WT or T12) binding is shown. (Mean ± SD; n=3; *P≤0.05, **P≤0.01, ***P≤0.001, T-test). (H) Vinculin was knocked down as described (Figure 5D) in MCF10A cells grown at high density on PDMS membranes. The membranes were stretched at 17% elongation for 2 hours and fixed while under tension and stained for TRIP6. Merged images show TRIP6 (red) and DNA (blue). Scale bar=20µm.

Expanded View Figure Legends.
Arabidopsis MAKR5 is a positive effector of BAM3-dependent CLE45 4 signaling (2016)
Figure 1

Figure 1 - SKM1 and SKM2 do not play a role in CLE45 perception in the root. (A-B) Expression patterns (blue stain) of GUS reporter genes driven by the SKM1 (A) or SKM2 (B) promoter in roots of 5-day-old Col-0 seedlings. (C) Non-complementation of CLE45 insensitivity of the bam3 mutant by BAM3 expressed under control of the SKM1 promoter (7-day-old seedlings; 2 independent transgenic lines per construct are shown). (D) Full CLE45 sensitivity of skm1 or skm2 single and double mutant roots (7-day-old seedlings). (E) Failure of SKM1 or SKM2 expressed under control of the BAM3 promoter to complement CLE45 insensitivity of the bam3 mutant (7-day-old seedlings; 3 independent transgenic lines per construct are shown). Differences as compared to mock are not statistically significant unless indicated (student’s t-test); * = p < 0.05; ** = p < 0.01; *** = p < 0.001; error bars indicate standard error of the mean;