title: Linker histone partial phosphorylation: effects on secondary structure and chromatin condensation.
abstract: Linker histones are involved in chromatin higher-order structure and gene regulation. We have successfully achieved partial phosphorylation of linker histones in chicken erythrocyte soluble chromatin with CDK2, as indicated by HPCE, MALDI-TOF and Tandem MS. We have studied the effects of linker histone partial phosphorylation on secondary structure and chromatin condensation. Infrared spectroscopy analysis showed a gradual increase of β-structure in the phosphorylated samples, concomitant to a decrease in α-helix/turns, with increasing linker histone phosphorylation. This conformational change could act as the first step in the phosphorylation-induced effects on chromatin condensation. A decrease of the sedimentation rate through sucrose gradients of the phosphorylated samples was observed, indicating a global relaxation of the 30-nm fiber following linker histone phosphorylation. Analysis of specific genes, combining nuclease digestion and qPCR, showed that phosphorylated samples were more accessible than unphosphorylated samples, suggesting local chromatin relaxation. Chromatin aggregation was induced by MgCl2 and analyzed by dynamic light scattering (DLS). Phosphorylated chromatin had lower percentages in volume of aggregated molecules and the aggregates had smaller hydrodynamic diameter than unphosphorylated chromatin, indicating that linker histone phosphorylation impaired chromatin aggregation. These findings provide new insights into the effects of linker histone phosphorylation in chromatin condensation.
authors: Departamento de Bioquímica y Biología Molecular, Facultad de Biociencias, Universidad Autónoma de Barcelona, 08193, Bellaterra, Barcelona, Spain.
authors: Rita
authors: R
authors: Lopez
authors: Division of Clinical Biochemistry, Biocenter, Innsbruck Medical University, A-6020, Innsbruck, Austria.
authors: Bettina
authors: B
authors: Sarg
authors: Division of Clinical Biochemistry, Biocenter, Innsbruck Medical University, A-6020, Innsbruck, Austria.
authors: Herbert
authors: H
authors: Lindner
authors: Laboratorio de Luminiscencia y Espectroscopia de Biomoléculas, Universidad Autónoma de Barcelona, 08193, Bellaterra, Barcelona, Spain.
authors: Salvador
authors: S
authors: Bartolomé
authors: Departamento de Bioquímica y Biología Molecular, Facultad de Biociencias, Universidad Autónoma de Barcelona, 08193, Bellaterra, Barcelona, Spain.
authors: Inma
authors: I
authors: Ponte
authors: Departamento de Bioquímica y Biología Molecular, Facultad de Biociencias, Universidad Autónoma de Barcelona, 08193, Bellaterra, Barcelona, Spain.
authors: Pedro
authors: P
authors: Suau
authors: Departamento de Bioquímica y Biología Molecular, Facultad de Biociencias, Universidad Autónoma de Barcelona, 08193, Bellaterra, Barcelona, Spain alicia.roque@uab.es.
authors: Alicia
authors: A
authors: Roque
mesh_terms: D000818:Animals; D002645:Chickens; D002843:Chromatin / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D051357:Cyclin-Dependent Kinase 2 / Q000378:metabolism; D006657:Histones / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D015636:Magnesium Chloride / Q000494:pharmacology; D008836:Micrococcal Nuclease; D010766:Phosphorylation; D017433:Protein Structure, Secondary
chemical_list: D002843:Chromatin; D006657:Histones; D015636:Magnesium Chloride; D051357:Cyclin-Dependent Kinase 2; D008836:Micrococcal Nuclease