TET1 DISPLAYS CATALYTIC AND NON-CATALYTIC FUNCTIONS IN THE ADULT MOUSE CORTEX
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dioxygenases
DNA demethylation
DNA hydroxymethylcytosine
Infinium BeadChip array
TET1
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Abstract
TET1/2/3 are epigenetic enzymes that reverse DNA methylation through oxidation, ultimately restoring 5-methylcytosine (5mC) to cytosine (C) via passive dilution during cell division or base excision repair of more highly oxidized forms. These oxidized constituents could be lost via passive dilution during cell division. Alternatively, 5-formylcytosine (5fC) and 5-carboxycytosine (5caC) are targeted by thymine DNA glycosylase (TDG) to restore unmodified C through the base excision repair (BER) pathway. Post-mitotic brain uniquely maintains high levels of 5hmC and at an order of magnitude more than other peripheral tissues or mouse embryonic stem cells. Thus, the role of TET protein functions in brain is of interest. Although TET2 and TET3 are more highly expressed in brain, TET1 is also expressed but has only been studied using deletion alleles, which precludes the disentangling of the catalytic and non-catalytic functions of TET1. Here, we systematically dissect the catalytic and non-catalytic functions of TET1 in the adult cortex by comparing TET1 wildtype (Tet1 WT), TET1 catalytically dead mutant (Tet1 HxD), and TET1 knockout (Tet1 KO) mice. New TET1 mouse models together with a chemoenzymatic sequencing allow us to parse out TET1’s functions. We show that Tet1 HxD and Tet1 KO mutations perturb distinct subsets of CpG sites. TET1 can demethylate 5mC through multiple modes of regulation, including bona fide catalytic activity and non-catalytic function. Gene Ontology (GO) analysis on specific differential 5hmC regions (5hMRs) demonstrates that TET1’s catalytic activity is crucial for neuronal-specific functions. With High-performance Liquid Chromatography Mass-Spectrometry (HPLC-MS) lipidomics, we report the novel phenotype of the Tet1 HxD mouse with accumulation of lysophospholipids lysophosphatidylethanolamine (LPE) and lysophosphatidylcholine (LPC). RNA-Seq further shows that TET1 mutations predominantly impact the genes that are alternatively spliced. In summary, we have shown that Tet1 HxD does not completely phenocopy Tet1 KO, providing evidence that TET1 could modulate distinct cortical functions through its catalytic and non-catalytic roles.