Non-catalytic roles for TET1 protein negatively regulating neuronal differentiation through srGAP3 in neuroblastoma cells

Abstract

The methylcytosine dioxygenases TET proteins (TET1, TET2, and TET3) play important regulatory roles in neural function. In this study, we investigated the role of TET proteins in neuronal differentiation using Neuro2a cells as a model. We observed that knockdown of TET1, TET2 or TET3 promoted neuronal differentiation of Neuro2a cells, and their overexpression inhibited VPA (valproic acid)-induced neuronal differentiation, suggesting all three TET proteins negatively regulate neuronal differentiation of Neuro2a cells. Interestingly, the inducing activity of TET protein is independent of its enzymatic activity. Our previous studies have demonstrated that srGAP3 can negatively regulate neuronal differentiation of Neuro2a cells. Furthermore, we revealed that TET1 could positively regulate srGAP3 expression independent of its catalytic activity, and srGAP3 is required for TET-mediated neuronal differentiation of Neuro2a cells. The results presented here may facilitate better understanding of the role of TET proteins in neuronal differentiation, and provide a possible therapy target for neuroblastoma.

Keywords

methylcytosine dioxygenase / TET1 / srGAP3 / neuronal differentiation / neuroblastoma cells

References

[1] Abdel-Wahab O, Mullally A, Hedvat C, Garcia-Manero G, Patel J, Wadleigh M, Malinge S, Yao J, Kilpivaara O, Bhat R (2009) Genetic characterization of TET1, TET2, and TET3 alterations in myeloid malignancies. Blood 114(1):144–147

[2] Brodeur GM, Bagatell R (2014) Mechanisms of neuroblastoma regression. Nat Rev Clin Oncol 11(12):704–713

[3] Chen K, Mi Y-J, Ma Y, Fu H-L, Jin W-L (2011) The mental retardation associated protein, srGAP3 negatively regulates VPA-induced neuronal differentiation of Neuro2A cells. Cell Mol Neurobiol 31 (5):675–686

[4] Cimmino L, Dawlaty MM, Ndiaye-Lobry D, Yap YS, Bakogianni S, Yu Y, Bhattacharyya S, Shaknovich R, Geng H, Lobry C(2015) TET1 is a tumor suppressor of hematopoietic malignancy. Nat Immunol 16(6):653–662

[5] Dawlaty MM, Breiling A, Le T, Raddatz G, Barrasa MI, Cheng AW, Gao Q, Powell BE, Li Z, Xu M (2013) Combined deficiency of Tet1 and Tet2 causes epigenetic abnormalities but is compatible with postnatal development. Dev Cell 24(3):310–323

[6] Ficz G, Branco MR, Seisenberger S, Santos F, Krueger F, Hore TA, Marques CJ, Andrews S, Reik W (2011) Dynamic regulation of 5-hydroxymethylcytosine in mouse ES cells and during differentiation. Nature 473(7347):398–402

[7] Fu H-L, Ma Y, Lu L-G, Hou P, Li B-J, Jin W-L, Cui D-X (2014) TET1 exerts its tumor suppressor function by interacting with p53-EZH2 pathway in gastric cancer. J Biomed Nanotechnol 10(7):1217–1230

[8] Göttlicher M, Minucci S, Zhu P, Krämer OH, Schimpf A, Giavara S, Sleeman JP, Coco FL, Nervi C, Pelicci PG (2001) Valproic acid defines a novel class of HDAC inhibitors inducing differentiation of transformed cells. EMBO J 20(24):6969–6978

[9] Hahn MA, Qiu R, Wu X, Li AX, Zhang H, Wang J, Jui J, Jin S-G, Jiang Y, Pfeifer GP (2013) Dynamics of 5-hydroxymethylcytosine and chromatin marks in Mammalian neurogenesis. Cell reports 3 (2):291–300

[10] Hsu C-H, Peng K-L, Kang M-L, Chen Y-R, Yang Y-C, Tsai C-H, Chu C-S, Jeng Y-M, Chen Y-T, Lin F-M (2012) TET1 suppresses cancer invasion by activating the tissue inhibitors of metalloproteinases. Cell Rep 2(3):568–579

[11] Ito S, Shen L, Dai Q, Wu SC, Collins LB, Swenberg JA, He C, Zhang Y (2011) Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine. Science 333(6047):1300–1303

[12] Jin C, Lu Y, Jelinek J, Liang S, Estecio MR, Barton MC, Issa J-PJ (2014) TET1 is a maintenance DNA demethylase that prevents methylation spreading in differentiated cells. Nucleic Acids Res 42(11):6956–6971

[13] Kaas GA, Zhong C, Eason DE, Ross DL, Vachhani RV, Ming G-L, King JR, Song H, Sweatt JD (2013) TET1 controls CNS 5-methylcytosine hydroxylation, active DNA demethylation, gene transcription, and memory formation. Neuron 79(6):1086–1093

[14] Ko M, An J, Bandukwala HS, Chavez L, Äijö T, Pastor WA, Segal MF, Li H, Koh KP, Lähdesmäki H (2013) Modulation of TET2 expression and 5-methylcytosine oxidation by the CXXC domain protein IDAX. Nature 497(7447):122–126

[15] Kriaucionis S, Heintz N (2009) The nuclear DNA base 5-hydroxymethylcytosine is present in Purkinje neurons and the brain. Science 324(5929):929–930

[16] Li T, Yang D, Li J, Tang Y, Yang J, Le W (2015) Critical role of Tet3 in neural progenitor cell maintenance and terminal differentiation. Mol Neurobiol 51(1):142–154

[17] Ma Y, Mi Y-J, Dai Y-K, Fu H-L, Cui D-X, Jin W-L (2013) The inverse F-BAR domain protein srGAP2 acts through srGAP3 to modulate neuronal differentiation and neurite outgrowth of mouse neuroblastoma cells. PLoS One 8(3):e57865

[18] Mariani CJ, Vasanthakumar A, Madzo J, Yesilkanal A, Bhagat T, Yu Y, Bhattacharyya S, Wenger RH, Cohn SL, Nanduri J (2014) TET1-mediated hydroxymethylation facilitates hypoxic gene induction in neuroblastoma. Cell Rep 7(5):1343–1352

[19] Mi Y, Gao X, Dai J, Ma Y, Xu L, Jin W-L (2015) A Novel Function of TET2 in CNS: Sustaining Neuronal Survival. Int J Mol Sci 16 (9):21846–21857

[20] Nakamura T, Liu Y-J, Nakashima H, Umehara H, Inoue K, Matoba S, Tachibana M, Ogura A, Shinkai Y, Nakano T (2012) PGC7 binds histone H3K9me2 to protect against conversion of 5mC to 5hmC in early embryos. Nature 486(7403):415–419

[21] Neri F, Dettori D, Incarnato D, Krepelova A, Rapelli S, Maldotti M, Parlato C, Paliogiannis P, Oliviero S (2015) TET1 is a tumour suppressor that inhibits colon cancer growth by derepressing inhibitors of the WNT pathway. Oncogene 34(32):4168–4176

[22] Pastor WA, Aravind L, Rao A (2013) TETonic shift: biological roles of TET proteins in DNA demethylation and transcription. Nat Rev Mol Cell Biol 14(6):341–356

[23] Perera A,Eisen D, Wagner M, Laube Silvia K, Künzel Andrea F, Koch S, Steinbacher J, Schulze E, Splith V, Mittermeier N, Müller M, Biel M, Carell T, Michalakis S (2015) TET3 is recruited by REST for context-specific hydroxymethylation and induction of gene expression. Cell Rep 11(2):283–294

[24] Qiao Y, Wang X, Wang R, Li Y, Yu F, Yang X, Song L, Xu G, Chin YE, Jing N (2015) AF9 promotes hESC neural differentiation through recruiting TET2 to neurodevelopmental gene loci for methylcytosine hydroxylation. Cell Discov 1:15017

[25] Rudenko A, Dawlaty MM, Seo J, Cheng AW, Meng J, Le T, Faull KF, Jaenisch R, Tsai L-H (2013) Tet1 is critical for neuronal activityregulated gene expression and memory extinction. Neuron 79 (6):1109–1122

[26] Scourzic L, Mouly E, Bernard OA (2015) TET proteins and the control of cytosine demethylation in cancer. Genome Med 7(1):9

[27] Tahiliani M, Koh KP, Shen Y, Pastor WA, Bandukwala H, Brudno Y, Agarwal S, Iyer LM, Liu DR, Aravind L (2009) Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1. Science 324(5929):930–935

[28] Tan L, Shi YG (2012) Tet family proteins and 5-hydroxymethylcytosine in development and disease. Development 139(11):1895–1902

[29] Wang Y, Xiao M, Chen X, Chen L, Xu Y, Lv L,Wang P, Yang H, Ma S, Lin H, Jiao B, Ren R, Ye D, Guan KL, Xiong Y (2015) WT1 recruits TET2 to regulate its target gene expression and suppress leukemia cell proliferation. Mol Cell 57(4):662–673

[30] Williams K, Christensen J, Pedersen MT, Johansen JV, Cloos PA, Rappsilber J, Helin K (2011) TET1 and hydroxymethylcytosine in transcription and DNA methylation fidelity. Nature 473(7347): 343–348

[31] Wu H, D’Alessio AC, Ito S, Xia K, Wang Z, Cui K, Zhao K, Sun YE, Zhang Y (2011) Dual functions of Tet1 in transcriptional regulation in mouse embryonic stem cells. Nature 473(7347):389–393

[32] Xu Y, Xu C, Kato A, Tempel W, Abreu JG, Bian C, Hu Y, Hu D, Zhao B, Cerovina T(2012) Tet3 CXXC domain and dioxygenase activity cooperatively regulate key genes for Xenopus eye and neural development. Cell 151(6):1200–1213

[33] Yao B, Jin P (2014) Unlocking epigenetic codes in neurogenesis. Genes Dev 28(12):1253–1271

[34] Zhang R-R, Cui Q-Y, Murai K, Lim YC, Smith ZD, Jin S, Ye P, Rosa L, Lee YK, Wu H-P (2013) Tet1 regulates adult hippocampal neurogenesis and cognition. Cell Stem Cell 13(2):237–245

[35] Zhang Q, Zhao K, Shen Q, Han Y, Gu Y, Li X, Zhao D, Liu Y, Wang C, Zhang X (2015) Tet2 is required to resolve inflammation by recruiting Hdac2 to specifically repress IL-6. Nature 525 (7569):389–393

[36] Zhao X, Dai J, Ma Y, Mi Y, Cui D, Ju G, Macklin WB, Jin W-L (2014) Dynamics of ten-eleven translocation hydroxylase family proteins and 5-hydroxymethylcytosine in oligodendrocyte differentiation. Glia 62(6):914–926

RIGHTS & PERMISSIONS

2016 The Author(s) 2016. This article is published with open access at Springerlink.com and journal.hep.com.cn