No abstract

Neurulation is a complex process of histogenesis involving the precise temporal and spatial organization of gene expression. Genes influencing neurulation include proneural genes determining primary cell fate, neurogenic genes involved in lateral inhibition pathways and genes controlling the frequency of mitotic events. This is reflected in the aetiology and genetics of human and mouse neural tube defects, which are of both multifactorial and multigenic origin. The X-linked gene Nap1l2, specifically expressed in neurons, encodes a protein that is highly similar to the nucleosome assembly (NAP) and SET proteins. We inactivated Nap1l2 in mice by gene targeting, leading to embryonic lethality from mid-gestation onwards. Surviving mutant chimaeric embryos showed extensive surface ectoderm defects as well as the presence of open neural tubes and exposed brains similar to those observed in human spina bifida and anencephaly. These defects correlated with an overproduction of neuronal precursor cells. Protein expression studies showed that the Nap1l2 protein binds to condensing chromatin during S phase and in apoptotic cells, but remained cytoplasmic during G1 phase. Nap1l2 therefore likely represents a class of tissue-specific factors interacting with chromatin to regulate neuronal cell proliferation.

The deletion of the neuronal Nap1l2 (nucleosome assembly protein 1-like 2) gene in mice causes neural tube defects. We demonstrate here that this phenotype correlates with deficiencies in differentiation and increased maintenance of the neural stem cell stage. Nap1l2 associates with chromatin and interacts with histones H3 and H4. Loss of Nap1l2 results in decreased histone acetylation activity, leading to transcriptional changes in differentiating neurons, which include the marked downregulation of the Cdkn1c (cyclin-dependent kinase inhibitor 1c) gene. Cdkn1c expression normally increases during neuronal differentiation, and this correlates with the specific recruitment of the Nap1l2 protein and an increase in acetylated histone H3K9/14 at the site of Cdkn1c transcription. These results lead us to suggest that the Nap1l2 protein plays an important role in regulating transcription in developing neurons via the control of histone acetylation. Our data support the idea that neuronal nucleosome assembly proteins mediate cell-type-specific mechanisms of establishment/modification of a chromatin-permissive state that can affect neurogenesis and neuronal survival.Mammals possess three neuron-specific nucleosome assembly protein (NAP)-encoding genes, NAP1L2 (29), NAP1L3 (39), and NAP1L5 (37), all of which are both intronless and monoallelically expressed. While the role of the ubiquitously expressed NAPs, NAP1 (NAP1L1) (15,17,19,25,35) and NAP2 (NAP1L4) (14,26), in the assembly of nucleosomes and transport of histones has been extensively characterized, little is known about the role and functioning of neuron-specific NAP1-like proteins.We previously showed that targeted deletion of the murine neuron-specific Nap1l2 (nucleosome assembly protein 1-like 2) gene leads to embryonic lethality from mid-gestation phase onwards, with surviving mutant chimeric embryos showing extensive surface ectoderm defects and open neural tubes similar to those observed in humans with spina bifida and anencephaly. These developmental defects were attributed to the overproliferation of neural precursor cells that is thought to be associated with the absence of Nap1l2 activity (27). The present study aimed to understand the cellular and molecular mechanisms underlying this knockout phenotype.Here we show by ex vivo differentiation studies of embryonic stem (ES) cells from which Nap1l2 was deleted that Nap1l2 regulates the kinetics of neuronal differentiation. In the absence of Nap1l2, neural precursors exhibit a diminished capacity for differentiation, an increase in proliferation, and increased levels of apoptosis. These effects of Nap1l2 deletion are associated with a global decrease in cellular levels of histone acetyltransferase (HAT) activity. This finding is supported by observations that the highly acidic Nap1l2 protein colocalizes to the chromatin in the neuronal nucleus, binds to histones H3 and H4 in vitro, and increases HAT activity. Loss of Nap1l2 results in extensive changes in the transcriptional profile of neural precursor cel...