Immunoprecipitation and mass spectrometry define TET1 interactome during oligodendrocyte differentiation

Abstract

Ten-eleven translocation (TET) proteins, encoding dioxygenase for DNA hydroxymethylation, are important players in nervous system development and disease. In addition to their proverbial enzymatic role, TET proteins also possess non-enzymatic activity and function in multiple protein–protein interaction networks, which remains largely unknown during oligodendrocyte differentiation. To identify partners of TET1 in the myelinating cells, we performed proteome-wide analysis using co-immunoprecipitation coupled to mass spectrometry (IP-MS) in purified oligodendrocyte precursor cells (OPCs) and mature oligodendrocytes (mOLs), respectively. Following a stringent selection of MS data based on identification reliability and protein enrichment, we identified a core set of 1211 partners that specifically interact with TET1 within OPCs and OLs. Analysis of the biological process and pathways associated with TET1-interacting proteins indicates a significant enrichment of proteins involved in regulation of cellular protein localization, cofactor metabolic process and regulation of catabolic process, et al. We further validated TET1 interactions with selected partners. Overall, this comprehensive analysis of the endogenous TET1 interactome during oligodendrocyte differentiation suggest its novel mechanism in regulating oligodendrocyte homeostasis and provide comprehensive insight into the molecular pathways associated with TET1.

The non-apoptotic function of Caspase-8 in negatively regulating the CDK9-mediated Ser2 phosphorylation of RNA polymerase II in cervical cancer

“…To identify Caspase-8 interacting proteins, the interactome analysis was performed as described earlier [ 48 ]. Essentially, Caspase-8 was immunoprecipitated (IP) from non-synchronized (NS) and S/G2-phase synchronized (synch.)…”

Oligodendroglial Epigenetics, from Lineage Specification to Activity-Dependent Myelination

“…As NSCs differentiate into OPCs, EZH2 and EED expression remain high, while their levels decrease in cells transitioning to neuronal and astrocyte lineages [ 90 , 92 ]. Moreover, the ChIP-sequencing of H3K27me3 in postnatal-day-1 rat OPCs highlights its genomic regulation of genes involved in the “global alternative lineage choice” [ 93 ]. Indeed, conditional ablation of Eed favors the differentiation into astroglial cells, to the detriment of the oligodendroglial lineage [ 92 ].…”

“…OPC differentiation is also dependent on marks that are generally associated with gene repression, such as histone methylation, deacetylation, and citrullination. Indeed, the ablation or inhibition of Prmt5 , Ezh2 , Eed , Hdac1/2 , or Padi2 in OPCs results in defective differentiation and myelination [ 55 , 92 , 93 , 121 , 132 , 135 , 137 , 143 , 155 , 156 , 157 , 159 ]. These marks mainly regulate the downregulation of OPC-specific inhibitors of differentiation (such as Id2/Id4 ) or cell cycle (such as Cdk4/6 , Cxcl2/5/10/14 ) genes, and therefore, are often essential for OPC cell cycle exits and early OL differentiation, but less involved in myelin maintenance [ 55 , 92 , 93 , 155 ].…”

“…Indeed, the ablation or inhibition of Prmt5 , Ezh2 , Eed , Hdac1/2 , or Padi2 in OPCs results in defective differentiation and myelination [ 55 , 92 , 93 , 121 , 132 , 135 , 137 , 143 , 155 , 156 , 157 , 159 ]. These marks mainly regulate the downregulation of OPC-specific inhibitors of differentiation (such as Id2/Id4 ) or cell cycle (such as Cdk4/6 , Cxcl2/5/10/14 ) genes, and therefore, are often essential for OPC cell cycle exits and early OL differentiation, but less involved in myelin maintenance [ 55 , 92 , 93 , 155 ]. These enzymes, such as HDACs, PRMT5, and PADI2, can also modify non-histone targets, altering the function or localization of oligodendroglial proteins or transcription factors (i.e., OLIG1, alpha-tubulin, PDGFRα, or myelin proteins) [ 55 , 154 , 158 , 160 , 170 ].…”

Nanoporous Substrates with Molecular-Level Perfluoroalkyl/Alkylamide Surface for Laser Desorption/Ionization Mass Spectrometry of Small Proteins

“…), a similar concept to liquid biopsy in the field of tumor pathology, , has been desired for reducing the physical load of patients and real-time monitoring of the states of diseases. Biological solution samples contain a large number of components with a wide range of molecular weights; therefore, detection of target compounds generally requires separation, concentration, and/or amplification processes, using liquid chromatography, immunoprecipitation, or polymerase chain reaction techniques. − Although these methods are highly effective for analysis of biomolecules, they are laborious, time-consuming, costly, and unsuitable for analyzing various kinds of compounds simultaneously.…”