The Condensation of Chromatin in Apoptotic Thymocytes Shows a Specific Structural Change

Abstract

Chromatin condensation and DNA cleavage at internucleosomal sites have been recognized early as hallmarks of apoptosis, and it has been suggested that extensive DNA chain scission could directly result in the formation of dense chromatin bodies. Here we have shown that no causal relationship exists between DNA degradation and chromatin condensation in glucocorticoid-induced thymocyte apoptosis. The chromatin rearrangement occurred independent of as well as prior to DNA cleavage and involved a specific conformational change at the nucleosome level. In the early stages of the process, the core particles appeared to be tightly packed face-to-face in smooth 11-nm filaments that progressively folded to generate a closely woven network. The network finally collapsed, producing dense apoptotic bodies. Since trypsin digestion relaxed condensed chromatin and histone H4 underwent appreciable deacetylation in the apoptotic cell, we suggest that changes in the DNA-histone interactions represented a major modulating factor of condensation.Although the term "apoptosis" was originally derived from the Greek to emphasize cytoplasmic and nuclear alterations peculiar to the process of programmed cell death (1), no attempt has been made thus far to search for the molecular events underlying these changes, particularly the collapse of the bulk of chromatin into dense domains. The reasons for this delay in the development of a fundamental approach are manifold. In the first place, the unique condensed appearance of the apoptotic nucleus is closely associated with the cleavage of chromatin at internucleosomal sites (2), a circumstance that supports the hypothesis of a causal relationship between extensive chromatin digestion and condensation (3). This early view has recently been challenged on the basis of more refined determinations of the chain length of the DNA isolated from apoptotic cells (4 -6) but has long distracted from the search for the molecular mechanisms involved in the process of condensation. Moreover, since apoptosis plays a key regulatory role in several physiological and pathological processes, major efforts are currently being directed to the elucidation of the biochemical aspects and to the identification of the genes involved in the activation of the cell death program. The onset of chromatin condensation might direct the orderly turning off of genes required for the execution of metabolic suicide, therefore warranting a detailed structural characterization of apoptotic chromatin and a search for terminal modulating factors.Bearing in mind the spatial distribution of interphase chromatin, the appearance of the apoptotic nucleus immediately suggests the occurrence of a structural change involving extremely large domains, and the question arises whether a specific conformational transition at the nucleosome level might account for such a catastrophic phenomenon. In the first place, is chromatin in apoptosis characterized by a three-dimensional array of nucleosomes different from that prevailing in the interphase 3...

Ultrasound imaging of apoptosis: high-resolution non-invasive monitoring of programmed cell death in vitro, in situ and in vivo

“…Due to the observed correspondence of apoptotic nuclear condensation (Allera et al, 1997) and subsequent nuclear fragmentation with changes in ultrasound images, we hypothesized that the nuclear material in the cells was responsible for the increased ultrasound backscatter. This hypothesis was tested using the same cell culture system, but with the pharmaco-active agent colchicine (Dustin, 1980), which resulted in condensed nuclear material in the form of metaphasic chromosomes, but not fragmentation.…”

“…Some 25 million of the protein units in combination with 2 m of DNA are packed into chromosomes which are approximately 6-8 µm in length when condensed (van Holde, 1988). The process of nuclear condensation, which takes place in the early stages of apoptosis (Allera et al, 1997), similar to mitosis, compacts chromosomes from forms which are distributed throughout most of the cell's nucleus to more canonical condensed forms. Other previous studies also support the postulate that the cellular nuclear material is the major scatterer of high frequency ultrasound (Bérubé et al, 1992;Hunt et al, 1995;Czarnota et al, 1997a).…”

Histone H2B Phosphorylation in Mammalian Apoptotic Cells

“…Using inhibitors or activators of protein kinases, apoptosis can be induced by staurosporine without considerable histone phosphorylation (48) or with H3 phosphorylation by gliotoxin, a protein kinase A activator (51). It was reported that histone H3 and H4 were deacetylated in rat apoptotic thymocytes (52). It would be of interest to ascertain the change in apoptotic histone acetylation under the influence of the caspase inhibitor, Z-Asp-CH 2 -DCB.…”

Relationship between in vitro drug sensitivity and clinical response of patients to treatment in chronic lymphocytic leukemia

“…Differential scanning calorimetry (DSc) is a relatively fast thermal technique that provides data on physical and energetic properties of cellular structures/compounds (19)(20)(21). earlier, it was documented that this method could be useful for the monitoring of native chromatin in cell nuclei (19).…”