Fig. 2: CLDN11 regulates ZO1 localization in seminiferous tubules.
a Whole-mount immunofluorescence staining of seminiferous tubules from Cldn11+/− and Cldn11−/− mice using anti-ZO1 and anti-WT1 antibodies. White dotted squares indicate the regions shown in high magnification images (High mag.). b Immunofluorescence staining of parental L cells and L cells stably expressing mouse CLDN11 using anti-CLDN11 and anti-ZO1 antibodies. Yellow arrowheads show colocalization of CLDN11 with ZO1. c Schematic diagram of the domain structure of the mouse ZO1 protein (1,745 amino acids). The N-terminal portion of ZO1 contains three PDZ domains, an SH3 domain, a guanylate kinase (GUK) domain, and an acidic domain (AD). Distinct portions of the ZO1 protein were fused to maltose-binding protein (MBP); shown by three black lines with amino acid numbers. Glutathione S-transferase (GST)-fused proteins of aa 179–207 and aa 179–205 of mouse CLDN11 were designated as GST-CL11 and GST-CL11ΔHV, respectively. MBP-fused proteins of PDZ1 (aa 19–113), PDZ2 (aa 181–292), and PDZ3 domain (aa 423–503) of ZO1 were designated as MBP-zPDZ1, MBP-zPDZ2, and MBP-zPDZ3, respectively. Input: purified MBP-fused proteins. GST pull-down assays were performed using GST-CL11, MBP-zPDZ1, MBP-zPDZ2, and MBP-zPDZ3 (d) or GST-CL11, GST-CL11ΔHV, and MBP-zPDZ1 (e). The resulting samples were separated by SDS–PAGE followed by Coomassie Brilliant Blue (CBB) staining. Scale bars: 30 μm (a) and 10 μm (b).
