Analysis of the interplay between MeCP2 and histone H1 during in vitro differentiation of human ReNCell neural progenitor cells
Siqueira, E (Institut Germans Trias i Pujol. Institut de Recerca contra la Leucèmia Josep Carreras)
Kim, Bo-Hyun (Department of Biochemistry and Microbiology. University of Victoria)
Reser, Larry (Department of Chemistry. University of Virginia)
Chow, Robert (Department of Biology. University of Victoria)
Delaney, Kerry (Department of Biology. University of Victoria)
Esteller, M (Universitat de Barcelona. Departament de Ciències Fisiològiques)
Ross, Mark M. (Department of Chemistry. University of Virginia)
Shabanowitz, Jeffrey (Department of Chemistry. University of Virginia)
Hunt, Donald F. (Department of Pathology. University of Virginia)
Guil, Sonia (Institut Germans Trias i Pujol. Institut de Recerca contra la Leucèmia Josep Carreras)
Ausiö, Juan (Department of Biochemistry and Microbiology. University of Victoria)

Data:	2023
Resum:	An immortalized neural cell line derived from the human ventral mesencephalon, called ReNCell, and its MeCP2 knock out were used. With it, we characterized the chromatin compositional transitions undergone during differentiation, with special emphasis on linker histones. While the WT cells displayed the development of dendrites and axons the KO cells did not, despite undergoing differentiation as monitored by NeuN. ReNCell expressed minimal amounts of histone H1. 0 and their linker histone complement consisted mainly of histone H1. 2, H1. 4 and H1. 5. The overall level of histone H1 exhibited a trend to increase during the differentiation of MeCP2 KO cells. The phosphorylation levels of histone H1 proteins decreased dramatically during ReNCell's cell differentiation independently of the presence of MeCP2. Immunofluorescence analysis showed that MeCP2 exhibits an extensive co-localization with linker histones. Interestingly, the average size of the nucleus decreased during differentiation but in the MeCP2 KO cells, the smaller size of the nuclei at the start of differentiation increased by almost 40% after differentiation by 8 days (8 DIV). In summary, our data provide a compelling perspective on the dynamic changes of H1 histones during neural differentiation, coupled with the intricate interplay between H1 variants and MeCP2. Abbreviations: ACN, acetonitrile; A, absorbance at 230 nm; bFGF, basic fibroblast growth factor; CM, chicken erythrocyte histone marker; CNS, central nervous system; CRISPR, clustered regulated interspaced short palindromic repeatsDAPI, 4,'6-diaminidino-2-phenylindole; DIV, days in vitro (days after differentiation is induced); DMEM, Dulbecco's modified Eagle medium; EGF, epidermal growth factor; ESC, embryonic stem cell; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GFAP, glial fibrillary acidic proteinHPLC, high-performance liquid chromatography; IF, immunofluorescence; iPSCs, induced pluripotent stem cells; MAP2, microtubule-associated protein 2; MBD, methyl-binding domain; MeCP2, methyl-CpG binding protein 2; MS, mass spectrometry; NCP, nucleosome core particle; NeuN, neuron nuclear antigen; NPC, neural progenitor cellPAGE, polyacrylamide gel electrophoresis; PBS, phosphate buffered saline; PFA, paraformaldehyde; PTM, posttranslational modification; RP-HPLC, reversed phase HPLC; ReNCells, ReNCells VM; RPLP0, ribosomal protein lateral stalk subunit P0; RT-qPCR, reverse transcription quantitative polymerase-chain reaction; RTT, Rett Syndrome; SDS, sodium dodecyl sulphate; TAD, topologically associating domain; Triple KO, triple knockout.
Nota:	The work was supported by the National Institutes of Health [GM037537]; Ontario Rett Syndrome Association Catalan and Spanish Associations for Rett Syndrome. This work was supported by a grant of the Ontario Rett Syndrome Association (ORSA) to JA and NIH grant GM037537 to D.F.H. We thank Lissa C. Anderson of the National High Magnetic Field Laboratory (NHMFL) for advice on mass spectrometry experiments and data analysis, and Greg Blakney of NHMFL for Predator Manual Validation Helper v2.8. We thank Dina L. Bai of the University of Virginia for Fragment Calculator.
Nota:	This work was supported by a grant of the Ontario Rett Syndrome Association (ORSA) to JA and NIH grant GM037537 to D.F.H. We thank Lissa C. Anderson of the National High Magnetic Field Laboratory (NHMFL) for advice on mass spectrometry experiments and data analysis, and Greg Blakney of NHMFL for Predator Manual Validation Helper v2.8. We thank Dina L. Bai of the University of Virginia for Fragment Calculator.
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, sempre que no sigui amb finalitats comercials, i sempre que es reconegui l'autoria de l'obra original.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	Histone H1 ; Chromatin and ReNCe
Publicat a:	Epigenetics, Vol. 18 Núm. 1 (2023) , p. 2276425, ISSN 1559-2308

DOI: 10.1080/15592294.2023.2276425
PMID: 37976174

6 p, 909.0 KB

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