Session: 886 APS Skeletal Muscle, Bone and Connective Tissue Poster Session
(886.10) Myonuclei Can Replicate DNA
Tuesday, April 5, 2022
10:15 AM – 12:15 PM
Location: Exhibit/Poster Hall A-B - Pennsylvania Convention Center
Poster Board Number: E319
Agnieszka Borowik (Oklahoma Medical Research Foundation), Frederick Peelor (Oklahoma Medical Research Foundation), Esther Dupont-Versteegden (University of Kentucky, University of Kentucky), John McCarthy (University of Kentucky, University of Kentucky), Benjamin Miller (Oklahoma Medical Research Foundation)
Presenting Author Oklahoma Medical Research Foundation Oklahoma City, Oklahoma
Skeletal muscle is a heterogenous tissue composed of multinucleated muscle fibers and accompanying supporting cells like satellite cells (SCs). Myonuclei are believed to be post-mitotic, arrested in G0 phase of the cell cycle and thus unable to proliferate. Consequently, it is hypothesized that growth of muscle is dependent on the donation of nuclei from SCs. Although some studies suggest that under certain conditions myonuclei can replicate, it has been difficult to address whether this replicative potential exists in vivo. We hypothesized that skeletal muscle resident myonuclei possess the ability to replicate in vivo. To test this hypothesis, we used the HSArtTA;Tet-O-H2B-GFP (HSA-H2B) mouse model, which uses doxycycline (DOX) treatment for temporal labeling (Tet-ON) of residential myonuclei. After removing DOX, and thus turning off GFP-labeling, we provided the stable isotope deuterium oxide (D2O) for 8 weeks to measure DNA synthesis. At the end of D2O labeling, muscles were harvested and GFP-positive myonuclei were sorted using an antibody-independent approach and a restrictive gating strategy. Both GFP+ (myonuclei) and GFP- (non-myonuclei) fractions were then analyzed for D2O incorporation into DNA by GC-QQQ. By our approach, DNA from GFP-positive myonuclei was deuterium-enriched and therefore deemed to be replicating. DNA replication occurred in at least one muscle of the 7 mice studied. We observed deuterium enrichment into DNA in 6 out of 7 plantaris (PLA), 4 out of 7 extensor digitorum longus (EDL), 4 out of 7 tibialis anterior (TA), 6 out of 7 gastrocnemius (GAS) and 7 out of 7 quadriceps (QUAD). The average fractional synthesis rates (FSR, %/day) of replicating myonuclei were: 0.0351 ± 0.017 in PLA, 0.0156 ± 0.015 in EDL, 0.0220 ± 0.018 in TA, 0.0223 ± 0.018 in GAS, and 0.02692 ± 0.043 in QUAD. These data are the first to unambiguously demonstrate that myonuclei replicate DNA in vivo. The proposed project challenges current dogma and might significantly impact skeletal muscle treatment to maintain muscle mass with age and disease.
