Involvement of NOS3 in RA-Induced neural differentiation of human NT2/D1 cells

Anna Jezierski; Paromita Deb-Rinker; Caroline Sodja; P Roy Walker; Dao Ly; Julie Haukenfrers; Jagdeep K Sandhu; Mahmud Bani-Yaghoub; Marianna Sikorska

doi:10.1002/jnr.23118

Involvement of NOS3 in RA-Induced neural differentiation of human NT2/D1 cells

J Neurosci Res. 2012 Dec;90(12):2362-77. doi: 10.1002/jnr.23118. Epub 2012 Sep 18.

Authors

Anna Jezierski¹, Paromita Deb-Rinker, Caroline Sodja, P Roy Walker, Dao Ly, Julie Haukenfrers, Jagdeep K Sandhu, Mahmud Bani-Yaghoub, Marianna Sikorska

Affiliation

¹ Neurogenesis and Brain Repair, Neurobiology Program, Institute for Biological Sciences, National Research Council Canada, Ottawa, Ontario, Canada.

PMID: 22987726
DOI: 10.1002/jnr.23118

Abstract

Nitric oxide (NO) plays a key role in neurogenesis as a regulator of cell proliferation and differentiation. NO is synthesized from the amino acid L-arginine by nitric oxide synthases (NOS1, NOS2, and NOS3), which are encoded by separate genes and display different tissue distributions. We used an in vitro model of RA-induced neural differentiation of NT2 cells to examine which of the three NO-synthesizing enzymes is involved in this process. The results revealed a transient induction of NOS3 (known as the constitutively expressed endothelial nitric oxide synthase; eNOS) during the time course of the RA treatment. The peak of gene expression and the nuclear presence of NOS3 protein coincided with cell cycle exit of NT2-derived neuronal precursors. The subsequent analysis of cytosine methylation and histone H3 acetylation of the human NOS3 5' regulatory sequences indicated that epigenetic modifications, especially upstream of the proximal promoter (-734 to -989, relative to exon 2 TSS at +1), were also taking place. NOS1 was expressed only in the differentiated neurons (NT2-N), whereas NOS2 was not expressed at all in this cellular model. Thus, a burst of NO production, possibly required to inhibit neural cell proliferation, was generated by the transient expression of NOS3. This pattern of gene expression, in turn, required epigenetic remodeling of its regulatory region.

MeSH terms

5' Untranslated Regions / genetics
Acetylation
Cell Cycle / drug effects
Cell Cycle / physiology
Cell Line, Tumor / cytology
Cell Line, Tumor / drug effects
Cell Nucleus / enzymology
Chromatin Immunoprecipitation
CpG Islands / genetics
DNA Methylation
Enzyme Induction / drug effects
Gene Expression Regulation, Developmental / drug effects
Histones / metabolism
Humans
Nerve Tissue Proteins / antagonists & inhibitors
Nerve Tissue Proteins / biosynthesis
Nerve Tissue Proteins / genetics
Nerve Tissue Proteins / physiology*
Neurogenesis / drug effects*
Neurogenesis / physiology
Neuroglia / cytology
Neurons / cytology
Nitric Oxide / physiology
Nitric Oxide Donors / pharmacology
Nitric Oxide Synthase Type I / biosynthesis
Nitric Oxide Synthase Type I / genetics
Nitric Oxide Synthase Type III / antagonists & inhibitors
Nitric Oxide Synthase Type III / biosynthesis
Nitric Oxide Synthase Type III / genetics
Nitric Oxide Synthase Type III / physiology*
Ornithine / analogs & derivatives
Ornithine / pharmacology
Protein Processing, Post-Translational
RNA, Messenger / biosynthesis
RNA, Messenger / genetics
Teratocarcinoma / pathology
Tretinoin / pharmacology*
Triazenes / pharmacology

Substances

1-hydroxy-2-oxo-3,3-bis(2-aminoethyl)-1-triazene
5' Untranslated Regions
Histones
Nerve Tissue Proteins
Nitric Oxide Donors
RNA, Messenger
Triazenes
Nitric Oxide
N(G)-iminoethylornithine
Tretinoin
Ornithine
NOS1 protein, human
NOS3 protein, human
Nitric Oxide Synthase Type I
Nitric Oxide Synthase Type III