A cytoplasmic role for histone methyltransferase (MET-2) in Caenorhabditis elegans
Date
2022
Authors
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Journal ISSN
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Publisher
University of Delaware
Abstract
Histone methyltransferases (HMTs) are known to affect chromatin condensation by methylating histones in the nucleus. Our collaborators from the Mango lab identified tropomyosin, which acts in the cytoplasm to regulate muscle contraction and stabilize actin filaments, as a potential target of the histone methyltransferase MET-2 through mass spectroscopy. We discovered that met-2 mutant animals, lev-11 knockdown animals, and animals expressing MET-2 with a nuclear localization sequence (NLS), all exhibited hypersensitivity to the acetylcholine receptor agonist levamisole. These results demonstrated that loss of met-2 and tropomyosin cause the same phenotype, and suggest a cytoplasmic role for MET-2 since sequestration of this histone methyltransferase in the nucleus resulted in a phenotype indistinguishable from the mutant. C. elegans body wall muscles send out actin-rich projections called muscle arms which extend from the body-wall muscles to the nerve cord, allowing the muscle cell to be innervated. Tropomyosin mutants have fewer and wider muscle arms compared to wild-type. To investigate if the loss of met-2 in the cytoplasm would also disrupt muscle arm morphology, I performed extensive imaging of muscle arms in wildtype, met-2 mutants, and NLS::MET-2 animals using LSM880 confocal microscopy. I observed that met-2 mutants exhibited significantly wider and more branched muscle arms compared to wild type but had no significant difference in the number of arms per muscle. This indicates that MET-2 function is necessary for proper muscle arm morphology but not for muscle arm formation. The NLS::MET-2 animals displayed muscle arm width and branching indistinguishable from wild-type animals, which suggests that MET-2 may affect the transcription of genes required to regulate the width of muscle arms. To determine if MET-2 localizes to the cytoplasm of the body wall muscles, I used the Andor Dragonfly spinning disk and super-resolution microscope to visualize MET-2::GFP and observed cytoplasmic signal in a striated pattern in the body wall muscles. I am currently imaging a strain that expresses both MET-2::GFP and TOMM-20::DsRed2 to determine if the MET-2::GFP colocalizes with TOMM-20::DsRed2, a fluorescent protein used to visualize muscle mitochondria. In conclusion, I determined that loss of met-2 leads to defects in muscle arm morphology. While NLS::MET-2 animals exhibit wild-type muscle arm morphology, it remains unclear whether this is because MET-2 nuclear function impacts muscle arms or if NLS::MET-2 is not actually sequestered in the nucleus. However, my MET-2::GFP images showing cytoplasmic localization of a histone methyltransferase for the first time are consistent with a role for MET-2 in the cytoplasm.
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Keywords
Caenorhabditis elegans, Histone methyltransferase, Cytoplasma