5-hmC in the brain is abundant in synaptic genes and shows differences at the exon-intron boundary.

Paradigms and Technologies
Methods Development


The 5-methylcytosine (5-mC) derivative 5-hydroxymethylcytosine (5-hmC) is abundant in the brain for unknown reasons. Here we characterize the genomic distribution of 5-hmC and 5-mC in human and mouse tissues. We assayed 5-hmC by using glucosylation coupled with restriction-enzyme digestion and microarray analysis. We detected 5-hmC enrichment in genes with synapse-related functions in both human and mouse brain. We also identified substantial tissue-specific differential distributions of these DNA modifications at the exon-intron boundary in human and mouse. This boundary change was mainly due to 5-hmC in the brain but due to 5-mC in non-neural contexts. This pattern was replicated in multiple independent data sets and with single-molecule sequencing. Moreover, in human frontal cortex, constitutive exons contained higher levels of 5-hmC relative to alternatively spliced exons. Our study suggests a new role for 5-hmC in RNA splicing and synaptic function in the brain.


Khare, Tarang; Pai, Shraddha; Koncevicius, Karolis; Pal, Mrinal; Kriukiene, Edita; Liutkeviciute, Zita; Irimia, Manuel; Jia, Peixin; Ptak, Carolyn; Xia, Menghang; Tice, Raymond; Tochigi, Mamoru; Moréra, Solange; Nazarians, Anaies; Belsham, Denise; Wong, Albert H C; Blencowe, Benjamin J; Wang, Sun Chong; Kapranov, Philipp; Kustra, Rafal; Labrie, Viviane; Klimasauskas, Saulius; Petronis, Arturas;


  • 5-Methylcytosine/ metabolism
  • Alternative Splicing
  • Animals
  • Brain/ physiology
  • Cell Line
  • Cytosine/ analogs & derivatives
  • Cytosine/ metabolism
  • Glucosyltransferases/ metabolism
  • Humans
  • Introns
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Microarray Analysis
  • Organ Specificity
  • RNA Splicing
  • Reproducibility of Results
  • Synapses/ genetics
  • Synapses/ metabolism

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