A Cytoplasmic Role for a Histone Methyltransferase in C. Elegans Muscle Development
Date
2021-05
Authors
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Publisher
University of Delaware
Abstract
Conventionally, histone methyltransferases (HMTs)act in the nucleus to methylate histones, impacting chromatin compaction. Surprisingly, our collaborators found that C.elegans tropomyosin, which acts in the cytoplasm to regulate muscle contraction and stabilize actin filaments, was methylated by a HMT. In C. elegans, actin rich muscle arms extend from the body-wall muscles to the nerve cord, allowing the muscle cell to be innervated. This led us to hypothesize that cytoplasmic HMT activity regulates actin filament organization and neuromuscular junction structure. A genome wide RNA interference (RNAi)screen showed that altered sensitivity to the acetylcholine receptor agonist levamisoleis indicative of genes required for muscle structure and function. We performed behavioral assays and discovered that the tropomyosin mutant, the HMT mutant and animals expressing the HMT with a nuclear localization sequence(NLS), all exhibited hypersensitivity to levamisole, further suggesting a cytoplasmic role for the HMT. Tropomyosin mutants have fewer and wider muscle arms compared to wildtype. We performed extensive imaging of muscle arms in HMT mutants using LSM880 confocal microscopy and observed that the HMT mutant exhibited significantly wider muscle arms compared to wild type but had no difference in the number of arms per muscle. This indicates that HMT function is necessary for proper muscle arm morphology but not for muscle arm formation, whereas tropomyosin is needed for both.
We then used tropomyosin RNAi to knock down tropomyosin in the HMT mutant to determine if the loss of the HMT and tropomyosin has an additive effect on muscle arm phenotype. After extensive muscle arm imaging and analysis, we discovered that the HMT mutant grown on tropomyosin RNAi had significantly wider and fewer muscle arms per muscle compared to wild type but was indistinguishable from the tropomyosin knockdown animals. This indicates that HMT and tropomyosin function together. We are now crossing a transgene expressing YFP-tagged acetylcholine receptors into the HMT mutant in order to determine if the altered muscle arm width causes a change in postsynapti cacetylcholine receptor (AChR) abundance. In conclusion, our results suggest that a HMT plays a novel cytoplasmic role to control muscle development and function.
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Keywords
Histone methyltransferase, C. elegans, Muscle, Tropomyosin