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HMGN5, an RNA and Nucleosome binding protein required for chromatin de-condensation and gene activation

ABSTRACT: The packaging of DNA into chromatin and its compaction within cells renders the underlying DNA template un-accessible for processes like transcription, replication and repair. Active mechanisms as chromatin modifying activities or the association with non-coding RNAs can de-condense chromatin, rendering it accessible for the DNA dependent processes. High mobility group proteins (HMG) are small architectural chromatin proteins that were shown to contribute to the regulation of chromatin accessibility and condensation. Here we show that HMGN5, a member of the HMGN family is capable to de-compact chromatin and to activate gene expression. We identified and characterized a novel RNA binding domain within HMGN5 that overlaps with its nucleosome binding domain (NBD). HMGN5 binds exclusively to nucleosomes or RNA, suggesting a switching mechanism and different functionalities depending on its binding partner. For this we show that HMGN5 is bound to regulatory regions of the genes that it tends to activate and also is found on the primary transcript of these genes. The results suggest that HMGN5 is switching from the chromatin binding to the RNA binding state after gene activation, potentially improving RNA synthesis. Furthermore, HMGN5 co-localizes and directly interacts with CTCF, suggesting a cooperative role of both proteins in organizing higher order structures of chromatin and active chromatin domains.

ORGANISM(S): Homo sapiens

PROVIDER: GSE166103 | GEO | 2024/05/08

REPOSITORIES: GEO

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Project description:In most metazoan nuclei, heterochromatin is located at the nuclear periphery in contact with the nuclear lamina, which provides mechanical stability to the nucleus. We show that in cultured cells, chromatin de-compaction by the nucleosome binding protein HMGN5 decreases the sturdiness, elasticity, and rigidity of the nucleus. Mice overexpressing HMGN5, either globally or only in the heart, are normal at birth but develop hypertrophic heart with large cardiomyoctyes, deformed nuclei and disrupted lamina, and die of cardiac malfunction. Chromatin de-compaction is seen in cardiomyocytes of newborn mice but misshaped nuclei with disrupted lamina are seen only in adult cardiomyocytes, suggesting that loss of heterochromatin diminishes the ability of the nucleus to withstand the mechanical forces of the contracting heart. Thus, heterochromatin enhances the ability of the nuclear lamina to maintain the sturdiness and shape of the eukaryotic nucleus; a structural role for chromatin that is distinct from its genetic functions. The full length mouse Hmgn5 cDNA, containing the entire 5’ UTR and FLAG tag on the 3’ end was amplified by PCR and inserted between BglII and XhoI sites of the pCALL vector (Novak et al., 2000). This vector contains a floxed LacZ cDNA driven by the CAG (a hybrid CMV enhancer and chicken beta–actin) promoter which is highly active in early mouse embryos. The vector was linearized by NotI digestion and injected into fertilized oocytes of C57BL/6 mice. Total embryonic excision of LacZ sequence was done by breeding floxed HMGN5-TG mice with SoxCre mice (Hayashi et al., 2002). Heart-specific excision was performed by breeding floxed HMGN5-TG mice with alphaMHC-cre mice (Oka et al., 2006).Total RNA from the left ventricle of newborn and adult Hmgn5-hTG mice which overexpress HMGN5 only in the heart as well as control mice was collected from biological triplicates (duplicates, in the case of newborn wild-type mice) and analyzed by Affymetrix expression arrays.

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HMGN5, an RNA and Nucleosome binding protein required for chromatin de-condensation and gene activation

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