title: Analysis of DNA Methylation and Hydroxymethylation in the Genome of Crustacean Daphnia pulex.
abstract: The aim of our study was to analyze the presence of 5-methyl-cytosine (5-mC) and 5-hydroxymethyl-cytosine (5-hmC) in the genome of crustacean Daphnia pulex. First, the presence of 5-mC and 5-hmC in genomic DNA was demonstrated by using antibodies specific to either 5-mC or 5-hmC. Then, analysis of 5-mC and 5-hmC using pairs of restriction enzymes with different sensitivity to methylation and hydroxymethylation confirmed the presence of both modifications in selected regions of three genes (Cox4, Cand2 and Ephx1). To get a detailed picture of 5-hmC distribution over the D. pulex genome, we performed 5-hmC enrichment and sequenced the enriched fraction using next generation sequencing and non-enriched library (input) as a control. Comparison of input and enriched libraries showed that 5-hmC in exons is twice as frequent as in introns. Functional analysis indicated that 5-hmC abundance is associated with genes that are involved in the adenylate cyclase-activating G-protein-coupled receptor signaling pathway, molting cycles, morphogenesis and cell fate determination. Genes that lack 5-hmC tend to be involved in the regulation of the transforming growth factor beta receptor signaling pathway and in many mRNA-related processes. Our results suggest that epigenetic modifications are present in the genome of D. pulex and most likely are involved in the regulation of gene expression of this crustacean.
authors.affiliation: Department of Botany and Genetics, Vilnius University, 21 Čiurlionis Str., LT-03101 Vilnius, Lithuania. dovile.strepetkaite@thermofisher.com.|Thermo Fisher Scientific Baltics, Graičiūno g. 8, LT-02241 Vilnius, Lithuania. dovile.strepetkaite@thermofisher.com.
authors.forename: Dovilė
authors.initials: D
authors.lastname: Strepetkaitė
authors.affiliation: Thermo Fisher Scientific Baltics, Graičiūno g. 8, LT-02241 Vilnius, Lithuania. gediminas.alzbutas@thermofisher.com.
authors.forename: Gediminas
authors.initials: G
authors.lastname: Alzbutas
authors.affiliation: Thermo Fisher Scientific Baltics, Graičiūno g. 8, LT-02241 Vilnius, Lithuania. eimantas.astromskas@thermofisher.com.
authors.forename: Eimantas
authors.initials: E
authors.lastname: Astromskas
authors.affiliation: Thermo Fisher Scientific Baltics, Graičiūno g. 8, LT-02241 Vilnius, Lithuania. arunas.lagunavicius@thermofisher.com.
authors.forename: Arūnas
authors.initials: A
authors.lastname: Lagunavičius
authors.affiliation: Thermo Fisher Scientific Baltics, Graičiūno g. 8, LT-02241 Vilnius, Lithuania. rasa.sabaliauskaite@thermofisher.com.
authors.forename: Rasa
authors.initials: R
authors.lastname: Sabaliauskaitė
authors.affiliation: Nature Research Centre, Akademijos str. 2, LT-08412 Vilnius, Lithuania. arbas@ekoi.lt.
authors.forename: Kęstutis
authors.initials: K
authors.lastname: Arbačiauskas
authors.affiliation: Department of Botany and Genetics, Vilnius University, 21 Čiurlionis Str., LT-03101 Vilnius, Lithuania. juozas.lazutka@gf.vu.lt.
authors.forename: Juozas
authors.initials: J
authors.lastname: Lazutka
keywords: 5-hydroxymethyl-cytosine; 5-methyl-cytosine; Cand2; Cox4; Daphnia pulex; Ephx1; epigenetic modifications; whole genome sequencing