Chromatin condensation during apoptosis is accompanied by degradation of lamin A+B, without enhanced activation of cdc2 kinase.

Abstract

Abstract. Chromatin condensation paralleled by DNA fragmentation is one of the most important criteria which are used to identify apoptotic cells. However, comparable changes are also observed in interphase nuclei which have been treated with cell extracts from mitotic cells. In this respect it is known that in mitosis, the lamina structure is broken down as a result of lamin solubilization and it is possible that a similar process is happening in apoptotic cells. The experiments described in this study have used confluent cultures of an embryonic fibroblast cell line which can be induced to undergo either apoptosis at low serum conditions or mitosis.Solubilization of lamin A+B was analyzed by immunoblotting and indirect immunofluorescence. These studies showed that in mitotic cells lamina breakdown is accompanied by lamin solubilization. In apoptotic cells, a small amount of lamin is solubilized before the onset of apoptosis, thereafter, chromatin condensation is accompanied by degradation of lamin A+B to a 46-kD fragment. Analysis of cellular lysates by probing blots with anti-PSTAIR followed by anti-phosphotyrosine showed that in contrast to mitosis, dephosphorylation on tyrosine residues did not occur in apoptotic cells. At all timepoints after the onset of apoptosis there was no significant increase in the activation of p34 ~2 as determined in the histone H1 kinase assay. Coinduction of apoptosis and mitosis after release of cells from aphidicolin block showed that apoptosis could be induced in parallel with S-phase.The sudden breakdown of chromatin structure may be the result of detachment of the chromatin loops from their anchorage at the nuclear matrix, as bands of 50 kbp and corresponding multimers were detectable by field inversion gel electrophoresis (FIGE). In apoptotic cells all of the DNA was fragmented, but only 14% of the DNA was smaller than 50 kbp. DNA strand breaks were detected at the periphery of the condensed chromatin by in situ tailing (ISTAIL). Chromatin condensation during apoptosis appears to be due to a rapid proteolysis of nuclear matrix proteins which does not involve the p34 ~c2 kinase.OPTOSIS waS originally defined as a program of morphological changes accompanying cell death during which the cells round up and the chromatin condenses, leading to the formation of crescent-shaped masses aggregating at the membrane. In parallel the nucleolus dissolves (Kerr, 1971). These initial changes are followed by cytoplasmic and nuclear compaction, and ultimately by fragmentation of the cell. The nuclear fragments formed in this process are still surrounded by the nuclear membranes (Kerr, 1971;Oberhammer et al., 1993a). As a common biochemical marker for apoptosis, an activation of a nonlysosomal, Ca2+/MgZ÷-dependent endonuclease has been suggested (Wyllie, 1980;Cohen and Duke, 1984). The response is so readily identifiable that endonuclease activation

A novel Bcl-2/Bcl-XL/Bcl-w inhibitor ABT-737 as therapy in multiple myeloma

“…Chromatin condensation and nuclear disintegration are typical signs of apoptotic cell death. MM.1S cells treated with ABT-737 (2 mM) for 48 h show a marked increase in nuclear condensation, as indicated (Oberhammer et al, 1994).…”

Mapping the initial DNA breaks in apoptotic Jurkat cells using ligation-mediated PCR

“…34 There have been a number of reports in the literature suggesting that initiation of apoptotic DNA cleavage occurs at matrix attachment regions. 6,7,[35][36][37] The first supporting evidence came from studies that establish that high molecular weight DNA fragments of 50 and 300 kb, produced in early stages of apoptosis, represent structural domains of chromatin, loops and rosettes, respectively. 1,7 It was then suggested that topoisomerase II, located at matrix attachment sites, is responsible for this cleavage since inhibition of its activity generated the same pattern of DNA fragments.…”

Protein expression changes in a cell system of beta-cell maturation reflect an acquired sensitivity to IL-1?

“…After maturation of the beta-cell phenotype protein spots containing lamin B1 and C2 were up-regulated. Proteolysis of lamins, the major structural proteins of the nuclear envelope, are observed in cells undergoing apoptosis [52]. The relevance of this in relation to beta-cell sensitivity to IL-1β is not clear.…”