Insights into DNA hydroxymethylation in the honeybee from in-depth analyses of TET dioxygenase

In mammals, a family of TET enzymes producing oxidized forms of 5-methylcytosine (5mC) plays an important role in modulating DNA demethylation dynamics. In contrast, nothing is known about the function of a single TET orthologue present in invertebrates. Here, we show that the honeybee TET (AmTET) catalytic domain has dioxygenase activity and converts 5mC to 5-hydroxymethylcytosine (5hmC) in a HEK293T cell assay. In vivo, the levels of 5hmC are condition-dependent and relatively low, but in testes and ovaries 5hmC is present at approximately 7–10% of the total level of 5mC, which is comparable to that reported for certain mammalian cells types. AmTET is alternatively spliced and highly expressed throughout development and in adult tissues with the highest expression found in adult brains. Our findings reveal an additional level of flexible genomic modifications in the honeybee that may be important for the selection of multiple pathways controlling contrasting phenotypic outcomes in this species. In a broader context, our study extends the current, mammalian-centred attention to TET-driven DNA hydroxymethylation to an easily manageable organism with attractive and unique biology.

[1]  R. Maleszka.  Elucidating the Path from Genotype to Behavior in Honey Bees: Insights from Epigenomics , 2012 .

[2]  G. L. Miklos,et al.  Epigenomic communication systems in humans and honey bees: From molecules to behavior , 2011, Hormones and Behavior.

[3]  Wael Tadros,et al.  Regulation of maternal transcript destabilization during egg activation in Drosophila. , 2003, Genetics.

[4]  G. Robinson,et al.  DNA methylation dynamics, metabolic fluxes, gene splicing, and alternative phenotypes in honey bees , 2012, Proceedings of the National Academy of Sciences.

[5]  F. Ceccherini‐Silberstein,et al.  Emerging patterns and implications of HIV-1 integrase inhibitor resistance , 2012, Current opinion in infectious diseases.

[6]  William A. Pastor,et al.  TETonic shift: biological roles of TET proteins in DNA demethylation and transcription , 2013, Nature Reviews Molecular Cell Biology.

[7]  T. Carell,et al.  5-Hydroxymethylcytosine, the sixth base of the genome. , 2011, Angewandte Chemie.

[8]  M. Fraga,et al.  Epigenetics and the environment: emerging patterns and implications , 2012, Nature Reviews Genetics.

[9]  S. Forêt,et al.  RNAi-induced phenotypes suggest a novel role for a chemosensory protein CSP5 in the development of embryonic integument in the honeybee (Apis mellifera) , 2007, Development Genes and Evolution.

[10]  N. Strausfeld.  Organization of the honey bee mushroom body: Representation of the calyx within the vertical and gamma lobes , 2002, The Journal of comparative neurology.

[11]  Harrison W. Gabel,et al.  The Maturing Brain Methylome , 2013, Science.

[12]  The Honeybee Genome Sequencing Consortium,et al.  Erratum: Insights into social insects from the genome of the honeybee Apis mellifera , 2006, Nature.

[13]  Matthew D. Schultz,et al.  Global Epigenomic Reconfiguration During Mammalian Brain Development , 2013, Science.

[14]  J. Jui,et al.  Dynamics of 5-hydroxymethylcytosine and chromatin marks in Mammalian neurogenesis. , 2013, Cell reports.

[15]  R. Maleszka,et al.  Insects as innovative models for functional studies of DNA methylation. , 2011, Trends in genetics : TIG.

[16]  R. A. Drewell,et al.  Kin conflict in insect societies: a new epigenetic perspective. , 2012, Trends in ecology & evolution.

[17]  Xiaodong Cheng,et al.  Recognition and potential mechanisms for replication and erasure of cytosine hydroxymethylation , 2012, Nucleic acids research.

[18]  Michael Weber,et al.  Mechanisms of DNA methylation and demethylation in mammals. , 2012, Biochimie.

[19]  David R. Liu,et al.  Conversion of 5-Methylcytosine to 5- Hydroxymethylcytosine in Mammalian DNA by the MLL Partner TET1 , 2009 .

[20]  A. H. Smits,et al.  Dynamic Readers for 5-(Hydroxy)Methylcytosine and Its Oxidized Derivatives , 2013, Cell.

[21]  L. Aravind,et al.  Impaired hydroxylation of 5-methylcytosine in myeloid cancers with mutant TET2 , 2010, Nature.

[22]  Heinrich Leonhardt,et al.  Intrinsic and Extrinsic Connections of Tet3 Dioxygenase with CXXC Zinc Finger Modules , 2013, PloS one.

[23]  Zachary D. Smith,et al.  DNA methylation: roles in mammalian development , 2013, Nature Reviews Genetics.

[24]  H. Leonhardt,et al.  Sensitive enzymatic quantification of 5-hydroxymethylcytosine in genomic DNA , 2010, Nucleic acids research.

[25]  D. Kronauer.  Genomic imprinting and kinship in the social Hymenoptera: what are the predictions? , 2008, Journal of theoretical biology.

[26]  Andrew B. Barron,et al.  Epigenomics and the concept of degeneracy in biological systems , 2013, Briefings in functional genomics.

[27]  Yuting Liu,et al.  Role of Tet1 in erasure of genomic imprinting , 2013, Nature.

[28]  E. Whitelaw,et al.  Epigenetic reprogramming: Enforcer or enabler of developmental fate? , 2010, Development, growth & differentiation.

[29]  Philipp Kapranov,et al.  Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells , 2011, Nature.

[30]  Li-Huei Tsai,et al.  Tet1 Is Critical for Neuronal Activity-Regulated Gene Expression and Memory Extinction , 2013, Neuron.

[31]  Julie A. Law,et al.  Establishing, maintaining and modifying DNA methylation patterns in plants and animals , 2010, Nature Reviews Genetics.

[32]  N. Heintz,et al.  The Nuclear DNA Base 5-Hydroxymethylcytosine Is Present in Purkinje Neurons and the Brain , 2009, Science.

[33]  H. Blau,et al.  DNA Demethylation Dynamics , 2011, Cell.

[34]  R. Kucharski,et al.  Extensive histone post-translational modification in honey bees. , 2013, Insect biochemistry and molecular biology.

[35]  Z. Deng,et al.  The role of Tet3 DNA dioxygenase in epigenetic reprogramming by oocytes , 2011, Nature.

[36]  Xiaoyu Zhang,et al.  Methylation of tRNAAsp by the DNA Methyltransferase Homolog Dnmt2 , 2006, Science.

[37]  S. Forêt,et al.  The Honey Bee Epigenomes: Differential Methylation of Brain DNA in Queens and Workers , 2010, PLoS biology.

[38]  X. Shirley Liu,et al.  Tet3 CXXC Domain and Dioxygenase Activity Cooperatively Regulate Key Genes for Xenopus Eye and Neural Development , 2012, Cell.

[39]  R. Chahwan,et al.  The multidimensional nature of epigenetic information and its role in disease. , 2011, Discovery medicine.