1. Fig. 2. Binding of recombinant full-length Lm-411 to muscle receptors, myotubes, and self-polymerization is enhanced by mag and αLNNd.
Binding of recombinant full-length Lm-411 to muscle receptors, myotubes, and self-polymerization is enhanced by mag and αLNNd. (A) Schematic of Lm-111, Lm-211, and Lm-411 and the linker molecules mag and αLNNd. Binding interactions of mag, αLNNd, laminins, integrins, and dystroglycan are indicated. (B) Binding curves of integrin α7X2β1 to Lm-111 and Lm-411. (C) Binding curves of Lm-111 and Lm-411 to purified αDG. (D) Binding curves of Lm-411 and Lm-411 plus an equimolar concentration of mag to purified αDG. Data in (B) to (D) are means ± SEM with n = 3 per condition. Curve fitting used the equation Y = Bmax*X/(Kd + X). (E) Polymerization assay for Lm-111, Lm-411, and an equimolar mixture of Lm-411 and αLNNd. Polymer formation was monitored by SDS–polyacrylamide gel electrophoresis (PAGE) and Coomassie blue staining after centrifugation. r2 = (Lm-111), r2 = (Lm-411 and αLNNd), and r2 = (Lm-411) using linear regression. (F) Confluent layer of cultured C2C12 myotubes after incubation with no Lm, recombinant Lm-211, Lm-411, or Lm-411 in the presence of an equimolar concentration of αLNNd and/or mag. C2C12-bound Lm-211 or Lm-411 was visualized by staining with antibodies to laminin-γ1. Representative immunofluorescence images (left) and quantification (right) of laminin-γ1 immunofluorescence (IF). Scale bar, 200 μm. n = 9 cultures (Lm-411 and mag), n = 6 cultures (Lm-411, Lm αLNNd, Lm αLNNd + mag, and Lm-211). Data are means ± SEM. ***P < 0.001; n.s., P > 0.05 (for exact P values, see table S3), one-way analysis of variance (ANOVA) with Bonferroni post hoc test.
Judith R. Reinhard et al., Sci Transl Med 2017;9:eaal4649
Published by AAAS