title: Comparative analyses of SUV420H1 isoforms and SUV420H2 reveal differences in their cellular localization and effects on myogenic differentiation.
abstract: BACKGROUND
  Methylation of histone H4 on lysine 20 plays critical roles in chromatin structure and function via mono- (H4K20me1), di- (H4K20me2), and trimethyl (H4K20me3) derivatives. In previous analyses of histone methylation dynamics in mid-gestation mouse embryos, we documented marked changes in H4K20 methylation during cell differentiation. These changes were particularly robust during myogenesis, both in vivo and in cell culture, where we observed a transition from H4K20me1 to H4K20me3. To assess the significance of this change, we used a gain-of-function strategy involving the lysine methyltransferases SUV420H1 and SUV420H2, which catalyze H4K20me2 and H4K20me3. At the same time, we characterized a second isoform of SUV420H1 (designated SUV420H1_i2) and compared the activity of all three SUV420H proteins with regard to localization and H4K20 methylation.
  
  
  RESULTS
  Immunofluorescence revealed that exogenous SUV420H1_i2 was distributed throughout the cell, while a substantial portion of SUV420H1_i1 and SUV420H2 displayed the expected association with constitutive heterochromatin. Moreover, SUV420H1_i2 distribution was unaffected by co-expression of heterochromatin protein-1α, which increased the targeting of SUV420H1_i1 and SUV420H2 to regions of pericentromeric heterochromatin. Consistent with their distributions, SUV420H1_i2 caused an increase in H4K20me3 levels throughout the nucleus, whereas SUV420H1_i1 and SUV420H2 facilitated an increase in pericentric H4K20me3. Striking differences continued when the SUV420H proteins were tested in the C2C12 myogenic model system. Specifically, although SUV420H1_i2 induced precocious appearance of the differentiation marker Myogenin in the presence of mitogens, only SUV420H2 maintained a Myogenin-enriched population over the course of differentiation. Paradoxically, SUV420H1_i1 could not be expressed in C2C12 cells, which suggests it is under post-transcriptional or post-translational control.
  
  
  CONCLUSIONS
  These data indicate that SUV420H proteins differ substantially in their localization and activity. Importantly, SUV420H2 can induce a transition from H4K20me1 to H4K20me3 in regions of constitutive heterochromatin that is sufficient to enhance myogenic differentiation, suggesting it can act an as epigenetic 'switch' in this process.
authors.affiliation: Department of Medical Genetics, School of Human Development, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: Leanna W K
authors.initials: LW
authors.lastname: Tsang
authors.forename: Ninghe
authors.initials: N
authors.lastname: Hu
authors.forename: D Alan
authors.initials: DA
authors.lastname: Underhill
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D002460:Cell Line; D002843:Chromatin / Q000378:metabolism; D044127:Epigenesis, Genetic; D006570:Heterochromatin / Q000378:metabolism; D011495:Histone-Lysine N-Methyltransferase / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008809:Mice, Inbred C3H; D008856:Microscopy, Fluorescence / Q000379:methods; D024510:Muscle Development; D020033:Protein Isoforms
chemical_list: D002843:Chromatin; D006570:Heterochromatin; D020033:Protein Isoforms; D011495:Histone-Lysine N-Methyltransferase; C542918:KMT5C protein, human