Extended Data Fig. 9: Genetic activation of MKK3b induces hypertrophy through a mechanism that is only partially dependent on mTORC1.
a, Schematic describing how electroporation was used to transfect mouse tibialis anterior (TA) muscles. Specifically, the TA muscles of male and female C57BL6 mice were co-transfected with plasmid DNA encoding tdTomato and LacZ as a control condition, Rheb as a direct activator of mTORC1, or with constitutively active (c.a.) mutant of MKK3b. Following electroporation, the mice were given daily intraperitoneal (IP) injections of the drug rapamycin (1.5 mg/kg) to inhibit signaling through mTORC1 or the solvent vehicle as a control. b, At 7 days post-transfection, the muscles were collected and subjected to immunohistochemistry for laminin to identify the periphery of the transfected (tdTomato positive) vs. non-transfected (tdTomato negative) fibers, scale bars = 50 µm. c, The cross-sectional area (CSA) of randomly selected fibers were measured, and then the values in the transfected fibers were expressed relative to the mean of the values observed in the non-transfected (control) fibers within each sample (n = 60–155 transfected and non-transfected fibers per sample). Values in the graphs are presented as the group mean ± SEM, the number of samples per group is indicated at the bottom of the bars in the graphs (605–889 fibers per group). The data were analyzed with two-way ANOVA. The P-value for each statistically significant pairwise comparison is annotated in the graphs with a * being used when P < 0.0001. Panel a created with BioRender.com.
