Structural domains and conformational changes in nuclear chromatin: A quantitative thermodynamic approach by differential scanning calorimetry

Cecilia Balbi, Maria L. Abelmoschi, Luca Gogioso, Silvio Parodi, Paola Barboro, Barbara Cavazza, Eligio Patrone

Research output: Contribution to journalArticle

Abstract

A good deal of information on the thermodynamic properties of chromatin was derived in the last few years from optical melting experiments. The structural domains of the polynucleosomal chain, the linker, and the core particle denature as independent units. The differential scanning calorimetry profile of isolated chromatin is made up of three endotherms, at ∼74, 90, and 107°C, having an almost Gaussian shape. Previous work on this matter, however, was mainly concerned with the dependence of the transition enthalpy on external parameters, such as the ionic strength, or with the melting of nuclei from different sources. In this paper we report the structural assignment of the transitions of rat liver nuclei, observed at 58, 66, 75, 92, and 107°C. They are representative of the quiescent state of the cell. The strategy adopted in this work builds on the method developed for the investigation of complex biological macromolecules. The heat absorption profile of the nucleus was related to the denaturation of isolated nuclear components; electron microscopy and electrophoretic techniques were used for their morphological and molecular characterization. The digestion of chromatin by endogenous nuclease mimics perfectly the decondensation of the higher order structure and represented the source of several misinterpretations. This point was carefully examined in order to define unambiguously the thermal profile of native nuclei. The low-temperature transitions, centered around 58 and 66°C, arise from the melting of scaffolding structures and of the proteins associated with heterogeneous nuclear RNA. The conformational change of the linker domain occurs at 75°C. Finally, the endotherms at 92 and 107°C reflect the denaturation of the core particle placed within an expanded loop and the 30-nm fiber, respectively. The stability of chromatin higher order structure appears to be enthalpy dependent, because a large decrease in the denaturation heat is associated with unfolding. Starting from these assignments, we gained an insight into the structural changes underlying the progress of the cell along the cycle. The 107°C endotherm is dominant in both G0 and S phase, but actively dividing hepatocytes are characterized by a reproducible broadening of the thermal profile at 100°C, related to the regional unwinding of the 30-nm fiber occurring during transcription and DNA replication.

Original languageEnglish
Pages (from-to)3220-3227
Number of pages8
JournalBiochemistry
Volume28
Issue number8
Publication statusPublished - 1989

ASJC Scopus subject areas

  • Biochemistry

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    Balbi, C., Abelmoschi, M. L., Gogioso, L., Parodi, S., Barboro, P., Cavazza, B., & Patrone, E. (1989). Structural domains and conformational changes in nuclear chromatin: A quantitative thermodynamic approach by differential scanning calorimetry. Biochemistry, 28(8), 3220-3227.