Abstract
Conditions have been found for the isolation of rat liver nuclei which maintain chromatin in its native state and suppresses endogenous nuclease activity. Chromatin prepared in this way can be dispersed into buffers containing various concentrations of monovalent or divalent cations so that the 30-nm fiber is either totally or partially decondensed. When probed with micrococcal nuclease, the digestion profiles show that although the 30-nm fiber can be cleaved periodically to generate superbead-like particles this only occurs under certain ionic conditions when the fiber is partially decondensed. It is likely that this cleavage pattern reflects the transient exposure of specific nuclease sensitive sites as the 30-nm fiber condenses, rather than the existence of a specific subunit of a beaded 30-nm fiber. The periodicity of these nuclease-sensitive sites appear to be related to the asymmetric distribution of histone H1 molecules along the length of the fiber.
Highlights
From the Cell Physiology Group, Divisionof Biological Sciences, National Research Counciol f Canada, Ottawa KIA OR6, Canada
Electronmicrographs that are considered to support these models have been generated, they invariably show that the 30-nm fiber is irregular or “knobby” in appearance [3, 10, 11].This observation is difficult to reconcile with the above models which would be expected to generate a smooth fiber of constant diameter
In thisreport we have critically examined the use of micrococcal nuclease as a probe of the structure of chromatin in interphase rat liver nuclei
Summary
NUCLEASE DIGESTION PROFILESREFLECTINTERMEDIATE STAGES INTHE FOLDING OF THE 30-NM FIBERRATHERTHAN THE EXISTENCE OF SUBUNIT BEADS*. Principally micrococcal nuclease (MNasel) which played a critical role in the elucidation of the subunit structure of the 10-nm fiber, are currentlybeing used as probes of the structure of the 30-nm fiber [12,13,14,15,16,17,18,19] These studies have shown that the fiber can, under certain conditions, be digested into a discrete class of particles variously described as nucleomers or superbeads [14, 18] that may represent subunits of a beaded 30-nm fiber. Thedata show thatthe ionic environment in which the nuclei are isolated has a great influence over the pattern of digestion by altering the structures of the 30-nm fiber during preparation of the nuclei This is noticeable when nuclei are isolated in the presence of the low concentrations of monovalent cations (25 mM) that are used in virtually all conventional nuclear isolation procedures.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
