Spatial distribution of lamin A and B1 in the K562 cell nuclear matrix stabilized with metal ions

When the nucleus is stripped of most DNA, RNA, and soluble proteins, a stru cture remains that has been referred to as the nuclear matrix, which acts a s a framework to determine the higher order of chromatin organization. Howe ver, there is always uncertainty as to whether or not the nuclear matrix, i solated in vitro, could really represent a skeleton of the nucleus in vivo. In fact, the only nuclear framework of which the existence is universally accepted is the nuclear lamina, a continuous thin layer that underlies the inner nuclear membrane and is mainly composed of three related proteins: la mins A, B, and C. Nevertheless, a number of recent investigations performed on different cell types have suggested that nuclear lamins are also presen t within the nucleoplasm and could be important constituents of the nuclear matrix. In most cell types investigated, the nuclear matrix does not spont aneously resist the extraction steps, but must rather be stabilized before the application of extracting agents. In this investigation, by immunochemi cal and morphological analysis, we studied the effect of stabilization with different divalent cations (Zn2+, Cu2+, Cd2+) On the distribution of lamin A and B1 in the nuclear matrix obtained from K562 human erythroleukcmia ce lls. In intact cells, antibodies to both lamin A and B1 mainly stained the nuclear periphery, although some immunoreactivity was detected in the nucle ar interior. The fluorescent lamin A pattern detected in Cu2+- and Cd2+-sta bilized nuclei was markedly modified, whereas Zn2+-incubated nuclei showed an unaltered pattern of lamin A distribution. By contrast, the distribution of lamin B1 in isolated nuclei was not modified by the stabilizing cations . When chromatin was removed by nuclease digestion and extraction with solu tions of high ionic strength, a previously masked immunoreactivity for lami n A, but not for lamin B1, became evident in the internal part of the resid ual structures representing the nuclear matrix. Our results indicate that w hen metal ions are used as stabilizing agents for the recovery of the nucle ar matrix, the distribution of both lamin A and lamin B1 in the final struc tures, corresponds to the pattern we have very recently reported using diff erent extraction procedures. This observation strengthen the concept that i ntranuclear lamins may act as structural components of the nuclear matrix. (C) 1999 Wiley-Liss, Inc.