The role of high mobility group of nucleosome binding proteins in stem cell biology and differentiation

The high mobility group of nucleosome binding proteins (HMGNs) are chromatin architectural proteins that bind specifically to nucleosomes and influence chromatin structure and DNA-dependent functions. However, the mechanisms underlying these events remain largely unknown. HMGN1 and HMGN2 are highly expressed by embryonic stem cells and are downregulated as differentiation proceeds. Nevertheless, embryonic and adult neural stem cells retain elevated levels of these proteins. Chromatin plasticity is essential for the pluri- and multipotency of stem cells and it is achieved by maintaining an open and dynamic chromatin conformation. Conversely, developmental potential seems to be restricted by chromatin condensation. The present work shows that loss of HMGN1 or HMGN2 in pluripotent embryonal carcinoma cells leads to increased spontaneous neuronal differentiation, which is accompanied by a reduction in pluripotency markers and higher gene expression of lineage-specific transcription factors. Inhibition of signalling pathways relevant for neurogenesis does not re-establish the phenotype observed in Hmgn2-knockout cells. Withdrawal of the factors sustaining pluripotency in embryonal carcinoma cells results in higher induction of pro-neural factors in cells lacking HMGN1 or HMGN2. Neural stem cells derived from Hmgn-knockout cells also display higher gene expression of pro-neural transcription factors and increased spontaneous neuronal differentiation. Loss of HMGN2 disrupts the active histone modification landscape, and therefore affects the chromatin structure at local and global levels. The proposition is that the local changes directly influence the transcription rates of pluripotency and lineage-specific transcription factors, while the global changes may restrict chromatin plasticity. The present data support a hypothesis whereby HMGNs contribute to the chromatin plasticity of stem cells by promoting an active histone modification landscape and open chromatin conformation, which are essential for preserving the self-renewal and developmental potential of stem cells

Characterising the HMGN Proteins Expression, and their Role, During Neuronal Differentiation of Mouse P19-ECCs in a Defined Adherent Culture System

No abstract available

Knockdown High Mobility Nucleosomal Binding Proteins 2 (HMGN2) Alter the Histone Modification H4K4me3 and H3K27me3 and Regulates Stem Cells Pluripotency

No abstract available

Nucleosome-binding HMGN Proteins Inhibit Stem Cell Differentiation Down the Neuronal Lineage

No abstract available

Maintenance of active chromatin states by HMGN2 is required for stem cell identity in a pluripotent stem cell model

Background: Members of the HMGN protein family modulate chromatin structure and influence epigenetic modifications. HMGN1 and HMGN2 are highly expressed during early development and in the neural stem/progenitor cells of the developing and adult brain. Here, we investigate whether HMGN proteins contribute to the chromatin plasticity and epigenetic regulation that is essential for maintaining pluripotency in stem cells. Results: We show that loss of Hmgn1 or Hmgn2 in pluripotent embryonal carcinoma cells leads to increased levels of spontaneous neuronal differentiation. This is accompanied by the loss of pluripotency markers Nanog and Ssea1, and increased expression of the pro-neural transcription factors Neurog1 and Ascl1. Neural stem cells derived from these Hmgn-knockout lines also show increased spontaneous neuronal differentiation and Neurog1 expression. The loss of HMGN2 leads to a global reduction in H3K9 acetylation, and disrupts the profile of H3K4me3, H3K9ac, H3K27ac and H3K122ac at the Nanog and Oct4 loci. At endodermal/mesodermal genes, Hmgn2-knockout cells show a switch from a bivalent to a repressive chromatin configuration. However, at neuronal lineage genes whose expression is increased, no epigenetic changes are observed and their bivalent states are retained following the loss of HMGN2. Conclusions: We conclude that HMGN1 and HMGN2 maintain the identity of pluripotent embryonal carcinoma cells by optimising the pluripotency transcription factor network and protecting the cells from precocious differentiation. Our evidence suggests that HMGN2 regulates active and bivalent genes by promoting an epigenetic landscape of active histone modifications at promoters and enhancers

The Hmgn Family of Chromatin Binding Proteins Regulates Stem Cells Pluripotency and Neuronal Differentiation

No abstract available

Knockdown High Mobility Nucleosomal Binding Proteins 2 (HMGN2) Alter the Histone Modification H4K4me3 and H3K27me3 and Regulates Stem Cells Pluripotency

No abstract available

Characterising the HMGN Proteins Expression, and their Role, During Neuronal Differentiation of Mouse P19-ECCs in a Defined Adherent Culture System

No abstract available

Nucleosome-binding HMGN Proteins Inhibit Stem Cell Differentiation Down the Neuronal Lineage

No abstract available

Development and Testing of a Low-Cost Inactivation Buffer That Allows for Direct SARS-CoV-2 Detection in Saliva

Massive testing is a cornerstone in efforts to effectively track infections and stop COVID-19 transmission, including places with good vaccination coverage. However, SARS-CoV-2 testing by RT-qPCR requires specialized personnel, protection equipment, commercial kits, and dedicated facilities, which represent significant challenges for massive testing in resource-limited settings. It is therefore important to develop testing protocols that are inexpensive, fast, and sufficiently sensitive. Here, we optimized the composition of a buffer (PKTP), containing a protease, a detergent, and an RNase inhibitor, which is compatible with the RT-qPCR chemistry, allowing for direct SARS-CoV-2 detection from saliva without extracting RNA. PKTP is compatible with heat inactivation, reducing the biohazard risk of handling samples. We assessed the PKTP buffer performance in comparison to the RNA-extraction-based protocol of the US Centers for Disease Control and Prevention in saliva samples from 70 COVID-19 patients finding a good sensitivity (85.7% for the N1 and 87.1% for the N2 target) and correlations (R = 0.77, p < 0.001 for N1, and R = 0.78, p < 0.001 for N2). We also propose an auto-collection protocol for saliva samples and a multiplex reaction to minimize the PCR reaction number per patient and further reduce costs and processing time of several samples, while maintaining diagnostic standards in favor of massive testing