Rabbit Anti-Human MEF2A (Phospho-Ser408) polyclonal antibody

Protein methyltransferases mediate posttranslational modifications of both histone and nonhistone proteins. Whereas histone methylation is well-known to regulate gene expression, the biological significance of nonhistone methylation is poorly understood. Methyltransferase-like 21c (Mettl21c) is a newly classified nonhistone lysine methyltransferase whose in vivo function has remained elusive. Using a Mettl21c(LacZ) knockin mouse model, we show here that Mettl21c expression is absent during myogenesis and restricted to mature type I (slow) myofibers in the muscle. Using co-immunoprecipitation, MS, and methylation assays, we demonstrate that Mettl21c trimethylates heat shock protein 8 (Hspa8) at Lys-561 to enhance its stability. As such, Mettl21c knockout reduced Hspa8 trimethylation and protein levels in slow muscles, and Mettl21c overexpression in myoblasts increased Hspa8 trimethylation and protein levels. We further show that Mettl21c-mediated stabilization of Hspa8 enhances its function in chaperone-mediated autophagy, leading to degradation of client proteins such as the transcription factors myocyte enhancer factor 2A (Mef2A) and Mef2D. In contrast, Mettl21c knockout increased Mef2 protein levels in slow muscles. These results identify Hspa8 as a Mettl21c substrate and reveal that nonhistone methylation has a physiological function in protein stabilization.

Isolated chicken myoblasts had previously been utilized in many studies aiming at understanding the emergence and regulation of the adult myogenic precursors (satellite cells). However, in recent years only a small number of chicken satellite cell studies have been published compared to the increasing number of studies with rodent satellite cells. In large part this is due to the lack of markers for tracing avian myogenic cells before they become terminally differentiated and express muscle-specific structural proteins. We previously demonstrated that myoblasts isolated from fetal and adult chicken muscle display distinct schedules of myosin heavy-chain isoform expression in culture. We further showed that myoblasts isolated from newly hatched and young chickens already possess the adult myoblast phenotype. in this article, we report on the use of polyclonal antibodies against the chicken myogenic regulatory factor proteins MyoD and myogenin for monitoring fetal and adult chicken myoblasts as they progress from proliferation to differentiation in culture. Fetal-type myoblasts were isolated from Ii-day-old embryos and adult-type myoblasts were isolated from 3-week-old chickens. We conclude that fetal myoblasts express both MyoD and myogenin within the first day in culture and rapidly transit into the differentiated myosin-expressing state. In contrast, adult myoblasts are essentially negative for MyoD and myogenin by culture Day 1 and subsequently express first MyoD and then myogenin before expressing sarcomeric myosin. The delayed MyoD-to-myogenin transition in adult myoblasts is accompanied by a lag in the fusion into myotubes, compared to fetal myoblasts. We also report on the use of a commercial antibody against the myocyte enhancer factor ZA (MEF2A) to detect terminally differentiated chicken myoblasts by their MEF2+ nuclei. Collectively, the results support the hypothesis that fetal and adult myoblasts represent different phenotypic populations. The fetal myoblasts may already be destined for terminal differentiation at the time of their isolation, and the adult myoblasts may represent progenitors that reside in an earlier compartment of the myogenic lineage.