Fig 1: Phosphorylated kindlin-2 (S159) levels positively correlate with IKKe expression in colorectal cancer tissues. (A) Immunohistochemical staining for phosphor-kindlin-2-S159 in human colorectal cancer tissues and paired adjacent normal tissues. Scale bar, 200 µm. (B) Phosphor-kindlin-2-S159 protein levels in 161 paired human colorectal cancer tissues and adjacent normal tissues as measured by immunohistochemical analysis. The data represent the means ± S.D. (***P < 0.001). (C) Representative images of paraffin tissue sections stained with IKKe and phosphorylated kindlin-2 (S159) antibody. Scale bar, 200 µm. (D) Correlation analyses of IKKe and phosphorylated kindin-2 (S159) protein levels in human colorectal cancer specimens.
Fig 2: HIF-1a interacts with Kindlin-2 and influences collagen biogenesis by targeting P4HA1 and FAK. (A-C) HIF-1a was overexpressed with HIF-1a plasmid or knocked down with HIF-1a siRNA (A), HIF-1a mRNA expression level was analyzed by real-time PCR (B), cell lysates were analyzed by western blot with the indicated antibodies. The expression levels of Kindlin-2, p-FAK, and P4HA1 were consistent with HIF-1a expression (C). (D-F) Kindlin-2 was overexpressed with Kindlin-2 plasmid or knocked down with Kindlin-2 siRNA (D), Kindlin-2 mRNA expression level was analyzed by real-time PCR (E), and cell lysates were analyzed by Western blot with the indicated antibodies. The expression levels of HIF-1a, p-FAK, and P4HA1 were consistent with Kindlin-2 expression (F)
Fig 3: Depletion of Kindlin-2 in adult smooth muscle leads to apoptosis. (A) Kindlin-2, a-SMA, cleaved caspase-3 immunohistochemical staining of aortic cross-sections (left) and colon (right). WT and iKO mice were treated with tamoxifen injection parallelly, ascending aortas and sigmoid colons were excised and fixed 14 days after injection. Scale bar, 100µm. (B) Immunofluorescence of TUNEL (red), DAPI (blue) in combination with a-SMA (green) on cross-sections of ascending aortas (up) and sigmoid colons (below). Higher magnified views of highlighted regions were shown on the right. Scale bar, 20µm. See Figure S5 for additional information.
Fig 4: IKKε-induced invadopodia formation depends on kindlin-2 S159 phosphorylation. (A) HCT116 cells were transfected with GFP-tagged IKKε and stained with antibodies against kindlin-2, Tks5 and F-actin (phalloidin). Confocal images were acquired to visualize the localization of IKKε, kindlin-2, Tks5 and F-actin. White arrowhead point to the co-localization of IKKε, kindlin-2, Tks5 and F-actin. Scale bar, 10 μm. (B) Co-localization of Tks5 and F-actin in HCT116 cells with kindlin-2 knockout and rescue with wild-type IKKε, wild-type kindlin-2, phosphor-null mutant kindlin-2 (S159A) and phosphor-mimetic mutant (S159D) kindlin-2 (left). Inset shows magnified image of invadopodia in the box. Quantification of cells with invadopodia (right). (C) Gelatin degradation in HCT116 cells with kindlin-2 knockout and rescue with wild-type IKKε, wild-type kindlin-2, phosphor-null mutant kindlin-2 (S159A) and phosphor-mimetic mutant (S159D) kindlin-2 (left). The area of degraded gelatin per cell (right). All data represent the means ± S.D. (N>400 cells/sample, *P<0.05, **P< 0.01, and ***P < 0.001).
Fig 5: Sertoli cell-specific knockout of Kindlin-2 in mice induced destruction of seminiferous tubules and testicular dysplasia.A Gross morphology of testes from WT (refers to Kindlin-2f/f) and KO (refers to Amh-Cre; Kindlin-2f/f) mice at 2–12 W. Compared to the WT mice, the size of KO testes is dramatically decreased. Scale bar, 5 mm. B Quantification of bilateral testes for mice (mean ± S.D. and n = 6). The variance was similar between the groups that are being statistically compared. Statistical analyses were performed using Student’s t-test at indicated time point. The variance was similar between the groups. *p < 0.05, **p < 0.01, ***p < 0.001. C Hematoxylin and eosin-staining (HE) and immunohistochemistry (IHC) of Kindlin-2, WT1 (Sertoli cell nucleus marker) and StAR (Leydig cell marker) in 4 W testes. Scale bar, 50 μm. Red arrows indicate the Sertoli cells, red and blank broken lines indicate the testicular tubular lumina. D Immunofluorescence of α-Tubulin (green) and laminin (red) in 4 W testes of WT and KO mice. The basement membrane of KO mice was thickener than that of WT mice (white arrow heads) Scale bar, 50 μm. E Transmission electron microscopy (TEM) of 8 W testes. SC means Sertoli cell, GC means germinal cell, BM means basement membrane. Scale bar, 2 μm. Red arrows: tight junction in WT testes and a large gap with only a loose cell junction between adjacent SCs in KO testes. Blue arrows: vacuoles located between adjacent SCs in KO testis.
Supplier Page from MilliporeSigma for Anti-Kindlin-2 antibody produced in rabbit