EMBO/soda-nel huggingface dataset:
Midbody accumulation through evasion of autophagy contributes to cellular reprogramming and tumorigenicity (2011)
figf3

(a) A histological section through mouse seminiferous tubules labelled for MKLP1 shows several MKLP1+ puncta in cells of the basal layer where stem cells reside. Scale bar, 20 μm. Inset: Enlargement of the cell marked by the arrow. (b,c) Electron micrographs of mitotic midbody (b, arrow) and multiple midbody-like structures in interphase cells with similar shapes and sizes in a juxtanuclear position (c, arrows) in basal cells of mouse seminiferous tubules. N, nucleus. Scale bars, 1 μm. (d) Representative planes of a neural progenitor cell in the ventricular zone (Sox2+, bottom left panel) of an E13.5 mouse brain show that an intracellular MBd (asterisk) is associated with the ventricle-facing daughter in the asymmetrically dividing cell (top row). The bottom row emphasizes the position of paired chromosomes in a dividing anaphase cell. CD133, midbody/MBd marker (green); Na/K-ATPase, cell-border marker (red); DRAQ5, DNA (blue); DAPI, DNA. v, ventricle. Scale bar, 5 μm. Note that abscission occurs apically in these cells. (e) A histological section through a hair follicle (left, phase-contrast microscopy) stained for the stem-cell marker keratin 15 (K15) to identify the bulge region (dotted box), the stem-cell niche. DNA stain (DAPI) and the phase-contrast microscopy image show full follicle architecture. (f) Upper panels show MBd-accumulating cells in the bulge region (boxed) co-labelled with K15 and MKLP1. Enlargements (lower panels) of the boxed region highlight a cell with four MBds (asterisks). N, nucleus. Scale bar, 5 μm. (g–i) Quantitative analysis and representative images show a decrease in MBd-accumulating cells on the differentiation of pluripotent stem cells (g) to fibroblast-like cells (h), and an increase in MBd-accumulating cells after reprogramming differentiated cells (h) to induced pluripotent stem cells (i). Numbers refer to mean±s.d.,
n=3. MKLP1, MBds; ZO-1, tight junctions; α-tubulin, microtubules; Aurora B, midbodies. Scale bar, 10 μm.
Midbody accumulation through evasion of autophagy contributes to cellular reprogramming and tumorigenicity (2011)
figf5

(a) Depiction of FPP assay. Digitonin selectively permeabilizes the plasma membrane but not internal membranes. Proteinase K degrades cytoplasmic components but membranous compartments remain intact. Under these conditions, MKLP1–GFP-labelled MBds (blue ellipse) in the cytoplasm will be degraded whereas those inside membrane-bound compartments will not. (b) MBds in MBd-poor hRPE-1 cells are largely protected (∼90% in membranous compartments, 10 cells analysed), whereas most MBds in HeLa cells are not (∼27%, 11 cells analysed), and are thus degraded in cytoplasm. Scale bar, 5 μm. (c) Presence of MBds in lysosomes on chloroquine or E64d–pepstatin A (E64d–PepA) inhibition in hRPE-1 and HeLa cells, but not in MCF-7 and H9 hESCs. Chloroquine treatment of H9 hESCs is not included as it caused differentiation and cell death. A representative image of hRPE-1 cells inhibited by chloroquine is shown, depicting two MBds inside lysosomes. MKLP1 and LAMP2 are used as MBd (red) and lysosome (green) markers, respectively. DAPI, DNA (blue).
n=100 MBds per treatment in each of the biological triplicates. Scale bar, 5 μm. (d) Percentage of MBd+ cells (MBd levels), percentage of MBds within lysosomes and percentage of cells exiting cytokinesis following synchronization. MKLP1 and LAMP2 are used as markers as in c. Note that MBds are transferred into only one of the two nascent daughter cells after abscission (Fig. 2d), so a 50% maximum will be expected for MBd+ cells. The peak of MBds transferred to cells is 3 h after plating followed by a peak of MBds entering lysosomes at 7 h. (e) Both chloroquine and E64d–PepA treatments increase the percentage of MBd+ cells in hRPE-1 cells and HeLa cells (chloroquine,
P=0.0021 and
P=0.0187, respectively; E64d–PepA,
P=0.0022 and
P=0.0043, respectively;
n=3 for all experiments). In contrast, lysosomal inhibition has no detectable effect on hESCs (H1, H9) or MCF-7 cancer cells. DMSO, dimethylsulphoxide. Data are presented as mean±s.d. (c–e), except mean±s.e.m. in hESCs (e).
Midbody accumulation through evasion of autophagy contributes to cellular reprogramming and tumorigenicity (2011)
figf6

(a) Single-plane confocal microscopy images of MBds within LC3-positive autophagosomes in MEFs expressing GFP–LC3 (left) and in hRPE-1 cells stained for endogenous LC3 (right). MBd markers: Cep55, MKLP1 or MgcRacGAP. Autophagosomes: GFP–LC3 or LC3. Note that MKLP1 (blue) and MgcRacGAP (red) are co-localized (magenta) in the autophagosome (green), indicating that MBds are sorted into autophagosomes. Scale bars, 2 μm. (b) Decreasing autophagy levels by deletion of the Atg5 gene (left) or depletion of ATG7 by siRNA (right) significantly increases the percentage of MBd+ cells (P=0.0019 and
P=0.021, respectively,
n=3). Immunoblots confirm loss of the Atg5–Atg12 conjugation in mutant cells and depletion of ATG7 (asterisk). GAPDH, glyceraldehyde 3-phosphate dehydrogenase. (c) Rapamycin (Rapa) and LiCl co-treatment induces autophagy and decreases the percentage of MBd+ cells (left, HeLa;
P=0.0056,
n=3). Immunoblots showing increased LC3-II levels confirm autophagy induction. Induction of autophagy by overexpression of Flag-tagged BECN1 reduces the percentage of MBd+ cells (right, MCF-7;
P=0.0008,
n=4). Ctrl, control. (d) Representative immunoblots showing high autophagy levels in normal cells and low levels in stem cells and cancer cells. Autophagic flux (autophagic activity) was measured by changes in the levels of LC3-II, in the presence or absence of lysosomal inhibitors E64d–PepA. U, uninhibited. I, inhibited. Below, the average of the percentage change in LC3-II levels after lysosomal inhibition from three experiments. α-tubulin, loading control. (e) Quantification of autophagic flux from three experiments in different cell lines. Normal dividing cells (MBd poor) typically have high autophagic flux, whereas stem and cancer cells (MBd rich) have low autophagic flux. The data are presented as mean±s.d. (b–e). Uncropped images of blots are shown in Supplementary Fig. S6.
Midbody accumulation through evasion of autophagy contributes to cellular reprogramming and tumorigenicity (2011)
figf7

(a) Single-plane confocal microscopy images showing co-localization of the MBds and the autophagic receptor NBR1 in U2OS cells and
p62-deleted MEFs. MBd markers: MKLP1 or Cep55. Scale bar, 2 μm. (b) The percentage of MBd+ cells is significantly increased following the depletion of NBR1 (P=0.022,
n=3), but not another autophagic receptor, p62. Co-depletion of NBR1 and p62 does not further increase MBd levels over NBR1 depletion alone. (c) Deletion of the
p62 gene does not affect the percentage of MBd+ cells. For b and c, immunoblots verify protein loss. (d) Co-immunoprecipitation (IP) reveals that CEP55 and NBR1 form a complex. Precipitated proteins and 5% of the input material (Input) were analysed by immunoblotting (IB) with antibodies against NBR1 or CEP55. (e–g) Overexpression of CEP55–eGFP (enhanced green fluorescent protein) increases the percentage of MBd+ cells (e;
P=0.0007,
n=3) and the percentage of NBR1-negative MBds (f;
P=0.0568,
n=3), presumably by sequestering NBR1 (red) away from MBds in cells expressing CEP55–eGFP (green) as shown in g, and consequently preventing MBd degradation. The dotted box in g is enlarged (top right panel), and the labelling of NBR1 and CEP55–eGFP (middle and bottom right panels) are also presented. DAPI, DNA (blue). Scale bar, 5 μm. The data are presented as mean±s.d. (b,c,e,f). Uncropped images of blots are shown in Supplementary Fig. S6.
Midbody accumulation through evasion of autophagy contributes to cellular reprogramming and tumorigenicity (2011)
figf8

(a–c) Reprogramming is more efficient after MBd enrichment. Differentiated cells (dH1f) and embryonic fibroblasts (IMR90) are reprogrammed after stable expression of either NBR1 shRNA or shNT. Emerging iPSC colonies are scored on the basis of TRA-1-60 expression. Cells depleted of NBR1 to increase MBd levels show an increase in iPSC colony formation (a,b, dH1f, 3.1±0.5-fold,
n=15,
P=0.00035; IMR90, 3.4±0.8-fold,
n=3,
P=0.02; data are mean±s.e.m.) but insignificant changes in autophagic activity (c) over shNT control. (b) Representative plates with TRA-1-60-immunostained iPSC colonies. The immunoblot (c, top) and densitometry (c, bottom; percentage of autophagic flux) show representative results (n=3); α-tubulin, loading control. (d) MCF-7 side-population (SP) cells have a significantly higher percentage of MBd+ cells over the non-side-population (MP;
P=0.0015,
n=3; data are mean±s.d.). (e,f) MBd enrichment in cancer cells leads to increased anchorage-independent growth. MKLP1–GFP-expressing HeLa cells are separated into ‘MBd-high’ and ‘MBd-low’ subpopulations. An increase in the ‘MBd high’ over ‘MBd low’ ratio is associated with an increase in soft-agar colony formation (e). No significant difference was observed when the enrichment of the MBd-high subpopulation was less than threefold. More soft-agar colonies are formed when MBds are enriched by NBR1 depletion (NBR1 shRNA) in HeLa (f, left;
P=0.0012,
n=3) and mouse 134-4 cells (f, right;
P=0.0086,
n=3); control, shNT. Data are mean±s.d., and the colony number (e,f) is the sum of INT-violet-stained colonies from ten random fields. (g) Model for MBd fate in cells. The newly formed MBd is preferentially inherited by the daughter cell with the older centrosome (top panel). The inherited MBd (black ring) is recognized by binding of the NBR1 autophagic receptor (grey circle) with the midbody protein CEP55 (magenta). The MBd is then encapsulated by the autophagosome (yellow circle), and degraded after fusion of autophagosome and lysosome (red circle) in differentiated cells. This pathway prevents MBd accumulation. In contrast, stem cells efficiently accumulate MBds through successive divisions and evasion of NBR1-mediated autophagy. Furthermore, differentiated and stem cells possess overall high and low autophagic activity, respectively. Uncropped images of blots are shown in Supplementary Fig. S6.
Transneuronal Dpr12/DIP-δ facilitate compartmentalized dopaminergic innervation of Drosophila mushroom body axons (2021)
Figure 2

Figure 2. Dpr12 is cell autonomously required for γ-axon regrowth into the γ4/5 zones.

(A-L) Confocal z-projections of MARCM neuroblast (NB, A-F) and single-cell (SC, G-L) clones labeled with membrane bound GFP (mCD8-GFP; CD8) driven by the γ specific Gal4 driver GMR71G10-Gal4 (γ-Gal4). At L3, NB and SC clones expressing dpr12 RNAi are similar to equivalent WT clones (A; n=20/20, D; n=15/15, G; n=15/15 and J; n=17/17). At 48hr APF and adult stage, WT NB (B; n=15/15, C; n=10/10) and SC (H; n=16/16, I; n=13/13) clones extend their axons to form the full adult lobe. In contrast, clones expressing dpr12 RNAi (E; n=14/14, F; n=22/2, K; n=18/24, L; n=19/27) fail to extend their axons to the distal part of the medial lobe (asterisks). I' and L' are traces of multiple single-cell clones depicting each cell in a different color.

(M) Top: Schematic representation of WT (orange) and dpr12 RNAi-expressing (green) single γ-KC axons. Bottom: Measurements of the relative location to which WT (I) and dpr12 RNAi (L) axons grow across the entire length of the adult γ lobe, alongside the relative position of the proximal border of the γ4 zone (see O, as well as Figure EV2).

(N) A table depicting the percentage of dpr12 RNAi-expressing single cell clones (SCCs) which stop at the γ3-γ4 border, at 48hr APF compared to the adult stage.

(O) Confocal z-projection of MBONγ4>γ1γ2 labeled by GMR18H09-Gal4 driving the expression of mCD8-GFP (CD8) shown in cyan.

Data information: yellow arrowheads demarcate single cell bodies. Green, white and cyan represent mCD8-GFP. Magenta represents FasII. Scale bar is 20 µm.
Transneuronal Dpr12/DIP-δ facilitate compartmentalized dopaminergic innervation of Drosophila mushroom body axons (2021)
Figure 3

Figure 3. Both Dpr12 and its interacting protein DIP-δ localize to the γ4/5 zones.

(A-H) Confocal z-projections of brains expressing MiMIC mediated Dpr12GFSTF (Dpr12-GFP; A-D) and DIP-δGFSTF (DIP-δ-GFP; E-H) fusion proteins at the indicated time points. See Figures EV 1 and 4 for more details on the fusion protein structure. (A-D) Dpr12-GFP is localized to the distal part of the γ-lobe at L3 (A; n=10/10), 48hr APF (C; n=12/12) and the adult stage (D; n=20/20), where it colocalizes with the γ4>γ1γ2 MBON (γ4; labeled by GMR18H09-Gal4 driving the expression of CD4-tdT). At 24hr APF (B; n=10/10), Dpr12-GFP appears diffuse. (E-H) DIP-δ-GFP is localized to the distal part of the γ-lobe throughout development: L3 (E; n=16/16), 24hr APF (F; n=10/10), 48hr APF (G; n=12/12) and adult (H; n=24/24). At the adult stage, DIP-δ-GFP is colocalized with the γ4>γ1γ2 MBON (γ4).

Data information: White dashed lines depict the γ-lobe. Green is GFP, cyan is CD4-tdT, magenta is FasII. Grayscale in the right panels of D and H represent single channels, as labelled. Insets in C, G represent greyscale magnifications of the GFP channel within the white boxes. In orange are transverse sections through the γ4 zone at the indicated locations; the γ-lobe is outlined in white and the β-lobe in yellow. Scale bar is 20 µm.
Transneuronal Dpr12/DIP-δ facilitate compartmentalized dopaminergic innervation of Drosophila mushroom body axons (2021)
Figure 4

Figure 4. DIP-δ is required for γ-axon regrowth into the γ4/5 zones

(A-H) Confocal z-projections DIP-δ hetero- and homozygous brains in which γ-KCs were labeled by expressing membrane bound tandem tomato (mtdT-HA) driven by the γ specific QF2 driver R71G10-QF2 (γ-QF2). Larval (L3) γ-axons grow normally in DIP-δT2A-Gal4 heterozygotes (A; n=16/16) and homozygotes (E; n=24/24). In contrast, at 48hr APF and adult, γ-axons within DIP-δT2A-Gal4 homozygotes do not enter the distal part of the lobe (asterisks; F; n=8/8, G; n=20/20), while they grow normally in heterozygotes (B; n=12/12, C; n=12/12). Overexpression of a DIP-δ transgene driven by the Gal4 activity of DIP-δT2A-Gal4 (see also Figure S4) does not affect normal growth (D; n=10/10) and rescues mutant phenotypes (H; n=20/20). Note that while the R71G10 driver is consistently expressed in γ-KCs, it is also expressed in α/β-KCs in a stochastic manner. The adult γ-lobe and α/β lobes are outlined in C-D in yellow and orange, respectively, for clarity.

(I) Ranking of regrowth for C, G, H, and Figure EV3D-E. Regrowth defect severity and statistics were calculated as in Fig. 1; Wilcoxon-Mann-Whitney test; ***p<0.001.

(J-M) Confocal z-projections of brains expressing DIP-δ-RNAi driven by the indicated Gal4. γ-KCs are labeled by mtdT-HA driven by R71G10-QF2 (γ-QF2). Expression of DIP-δ-RNAi in all glia (Repo-Gal4, J; n=28/28) or all KCs (OK107-Gal4, K; n=12/12) did not affect γ-neuron regrowth. In contrast, expression of DIP-δ-RNAi in all postmitotic neurons (C155-Gal4, L; n=9/12) or DIP-δ expressing neurons (DIP-δT2A-Gal4, M; n=21/22) induced a defect in γ4/5 innervation by γ-axons.

Data information: Asterisks demarcate distal part of the lobe. Green and white are mtdT-HA, magenta is FasII, Scale bar is 20 µm.
Transneuronal Dpr12/DIP-δ facilitate compartmentalized dopaminergic innervation of Drosophila mushroom body axons (2021)
Figure 5

Figure 5. PAM-DANs are the source of DIP-δ in the γ4/5 zones.

(A-D) Confocal z-projections of brains expressing DIP-δGFSTF (DIP-δ-GFP) together with the indicated Gal4s and transgenes. Expressing diphtheria toxin (DTi) and the membrane bound RFP (mCD8-RFP; CD8) driven by the γ4>γ1γ2 MBON driver MB294B-Gal4 (MBONγ4>γ1γ2-Gal4) did not affect DIP-δ-GFP expression (A, n=16/16; B, n=14/14). In contrast, similar expression of DTi and membrane bound Tomato (CD4-tdT; CD4) in PAM-DANs (using the GMR58E02-Gal4; PAM-DAN-Gal4) abolished the normal DIP-δ-GFP expression in the γ4/5 zone (compare D, n=18/18, to C, n=16/16) and within the PAM-DAN cell bodies (compare D' to C').

Magenta is CD8-RFP (A-B), CD4-mtdT (C-D). Green is GFP, grayscale depict individual channels as labeled. Scale bar is 20 µm. Yellow dashed line demarcates the γ-lobe based on FasII staining (not shown).

(E-G) Confocal z-projections of MARCM clones labeled by DIP-δT2A-Gal4 (DIP-δ Gal4) driving the expression of membrane bound GFP (mCD8-GFP; CD8) and heat shocked at 24hr after egg laying. Clones innervate the γ4/5 zones at 24hr APF (E; n=8), 48hr APF (F; n=8) and adult (G; n=16). Clones become tyrosine hydroxylase (TH) positive only at 48hr APF onwards (F,G).

Magenta is FasII, green is mCD8-GFP, cyan is TH antibody staining, grayscale single channels are shown as indicated. White dashed line demarcates the γ-lobe. Scale bar is 20 µm.
Transneuronal Dpr12/DIP-δ facilitate compartmentalized dopaminergic innervation of Drosophila mushroom body axons (2021)
Figure 6

Figure 6. DIP-δ is required and sufficient for Dpr12 localization

(A-J) Confocal z-projections of brains expressing MiMIC mediated Dpr12GFSTF (Dpr12-GFP) and DIP-δGFSTF (DIP-δ-GFP) fusion proteins of the indicated genotypes and time points. (A-D) Dpr12-GFP expression is diffuse in DIP-δT2A-Gal4 homozygotes mutant brains at L3 (A; n=20/20), 24hr APF (B; n=14/14), 48hr APF (C; n=28/28) and adult (D; n=26/26). (E-H) DIP-δ-GFP expression in dpr12∆50-81 homozygotes mutant brains remains localized to the distal part of the γ-lobe at L3 (E; n=16/16), 24hr APF (F; n=16/16), and 48hr APF (G; n=10/10) but cannot be identified in adult brains (H; n=16/16). (I-J) Dpr12-GFP expression in WT animals (I, n=8/8) or in those ectopically expressing DIP-δ in PAM-DANs that innervate the γ3 zone (J, n=14/14) driven by MB441B-Gal4 (PAM-DAN-γ3-Gal4). DIP-δ expression in PAM-DAN-γ3 resulted in Dpr12-GFP localization within the γ3 zone (arrow), in addition to its normal γ4/γ5 localization.

Data information: Arrowheads demarcate DIP-δ expression at the distal part of the lobe. Asterisks demarcate the distal part of the lobe. Dashed line depicts the medial γ-lobe, as determined by FasII staining. See legend of Figure EV1D for an explanation regarding the β-lobe morphological defects observed in E, J. Green is GFP, magenta is FasII. Grayscale panels represent single channels, as indicated. Scale bar is 20 µm.
Transneuronal Dpr12/DIP-δ facilitate compartmentalized dopaminergic innervation of Drosophila mushroom body axons (2021)
Figure 7

Figure 7. Dpr12-DIP-δ interaction mediates circuit assembly

(A-F) Top: Confocal z-projections of dpr12∆50-81 heterozygous (A, n=15; C, n=10; E, n=10) and homozygous brains (B, n=8; D, n=18; F, n=24) expressing mCD8-GFP (CD8) driven by: (A, B) R10G03-Gal4 (PAM-DAN-γ4-Gal4); (C, D) R48H11-Gal4 (PAM-DAN-γ5-Gal4), or (E, F) R18H09-Gal4 (MBONγ4>γ1γ2 -Gal4). The greyscale channels are sub-z-projections comprised of slices restricted to the γ-lobe region. White dashed line demarcates the γ-lobe. Bottom: Cartoons schematizing MB lobe structure and innervation by specific PAM-DANs or MBON.

(G-H) Single confocal slices of WT (G, n=5/5) and dpr12∆50-81 homozygous brains (H, n=5/5) stained with anti-Brp. Dashed lines demarcate neuropil boundaries, schematic shown below. Cre, crepine, a neuropil that surrounds the medial MB lobes; AL, antenna lobe.

Data information: Magenta is FasII, green and cyan are mCD8-GFP, grayscale depicts single channels as indicated. Scale bar is 20 µm.
Transneuronal Dpr12/DIP-δ facilitate compartmentalized dopaminergic innervation of Drosophila mushroom body axons (2021)
Figure 8

Figure 8. γ4/5 zone formation depends on matching Dpr-DIP-pairing between γ-KCs and PAM-DANs

(A, B) Confocal z-projections of adult DIP-δT2A-Gal4/1-119 trans-heterozygous mutant brains, in which γ-KCs are labeled by membrane bound tandem tomato (mtdT-HA; green) driven by R71G10-QF2 (γ-QF2), and DIP-δ-Gal4 (expressed in DIP-δ+ neurons) either drives expression or UAS-DIP-α (B) or not (A). Grey boxes below the images describe the relevant components within PAM-DANs and γ-KCs.

(C, D) Confocal z-projections of adult DIP-δT2A-Gal4/1-119 trans-heterozygous mutant (D) or DIP-δ1-119/+ heterozygous (C) brains, which express MiMIC mediated Dpr12GFSTF (Dpr12-GFP; green), in which DIP-δ-Gal4 either drives expression or UAS-DIP-α (D) or not (C). Grayscale panels represent single channels, as indicated. The γ-lobe is outlined in white.

(E, F) Confocal z-projections of adult dpr12Δ50-81 homozygous mutant brains, in which DIP-δ-Gal4 drives expression of either UAS-mtdT (E) or UAS-DIP-α-T2A-tdT (F; expected to induce expression of DIP-α as well as tdT encoded by a polycistronic message). In orange are longitudinal sections across the γ-lobe at the indicated location. The γ-lobe is outlined in white, as determined by FasII staining (grey in E'-F'). Cyan is tdT within DIP-δ+ cells. Grey boxes below the images describe the relevant components within PAM-DANs and γ-KCs.

Data information: In all images, magenta is FasII. Asterisks demarcate the distal part of the lobe. Scale bar is 20 µm.

(G) Ranking of regrowth for A, B, E, F and Figure EV8A-B. Regrowth defect severity and statistics were calculated as in Fig. 1; Wilcoxon-Mann-Whitney test; ***p<0.001; n,s, not significant.

Expanded View Figure Legends
Identification of integrin-linked kinase (ILK) as a critical regulator of VEGFR3 signalling and lymphatic vascular growth (2018)
Figure 2

Figure 2

ILK controls tyrosine phosphorylation of VEGFR3 in mouse embryos.

A-D LSM images of proximity ligation assay (PLA) dots composed of VEGFR3 and phosphorylated tyrosine (p-Tyr) on cross-sections through the jugular lymph sac / primordial thoracic duct (jls/pTD) of E13.5 control and ILK K.O. embryos. Scale bars: 10 µm.

E-H LSM images of PLA dots composed of VEGFR2 and phosphorylated tyrosine (p-Tyr) on cross-sections through the jls/pTD of E13.5 control and ILK K.O. embryos. Scale bars: 10 µm.

I Quantification of the PLA dots indicating VEGFR3 with phosphorylated tyrosine (p-Tyr) per LEC of E13.5 control or ILK K.O. embryos (n = 9 embryos per genotype), *P = 0.022.

J Quantification of the PLA dots indicating VEGFR2 with phosphorylated tyrosine (p-Tyr) per LEC of E13.5 control or ILK K.O. embryos (n = 3 embryos per genotype).

K Quantification of the total number of PLA dots indicating both VEGFR2 (left) and VEGFR3 (right) with phosphorylated tyrosine (p-Tyr) per jls/pTD section of E13.5 control or ILK K.O. embryos (n = 3 embryos per genotype), *P = 0.005 (VEGFR2/p-Tyr in ILK K.O. versus VEGFR3/p-Tyr in ILK K.O.), *P = 0.013 (VEGFR3/p-Tyr in control versus VEGFR3/p-Tyr in ILK K.O.).

Data information: Data are presented as means ± SEM and in (I, J) data are shown as percentage of control embryos; unpaired two-tailed Student's t-test (I, J) or two-way ANOVA with Tukey's multiple comparisons test (K). Note that not all significances are shown in (K).
Identification of integrin-linked kinase (ILK) as a critical regulator of VEGFR3 signalling and lymphatic vascular growth (2018)
Figure 3

Figure 3

ILK controls VEGFR3-β1 integrin interactions in mouse embryos.

A-D LSM images of cross-sections through the jugular lymph sac / primordial thoracic duct (jls/pTD) of an E13.5 control embryo stained for surface VEGFR3 and β1 integrin. Scale bar: 10 µm.

E-H LSM images of proximity ligation assay (PLA) dots (arrows) composed of VEGFR3 and β1 integrin on cross-sections through the jls/pTD of E13.5 control and ILK K.O. embryos. Scale bars: 10 µm.

I Quantification of VEGFR3/β1 integrin PLA dots normalised to Lyve1-positive area of control or ILK K.O. embryos (n = 6 embryos per genotype), *P = 0.005.

Data information: Data are presented as means ± SEM, shown as percentage of control embryos, unpaired two-tailed Student's t-test.