Organization and modulation of nuclear lamina structure

Abstract

The nuclear lamina is a protein meshwork associated with the nucleoplasmic surface of the inner nuclear membrane, that is suggested to be important for organizing nuclear envelope and interphase chromosome architecture. To investigate the structural organization of the lamina, we have analysed rat liver nuclear envelopes by various chemical extraction procedures. From these studies, we have defined conditions that yield a nuclear envelope subfraction that is both highly enriched in the lamina and devoid of pore complexes. This fraction contains mostly lamins A, B and C, the three major lamina polypeptides that are apparently arranged in a polymeric assembly. Our chemical extraction studies also indicate that lamin B has a stronger interaction with nuclear membranes than the other two lamins, and support the possibility that lamin B is important for attaching the lamina to the inner nuclear membrane. We have examined the synthesis and assembly of the lamins during interphase in tissue-culture cells to investigate lamina structure by a second approach. We found that all three lamins are synthesized at similar rates throughout the cell cycle in synchronized Chinese hamster ovary cells, and that their biosynthesis is not temporally coupled to DNA replication. Our studies indicate that newly synthesized lamins are rapidly assembled into an insoluble lamina structure but that the apparent half-time for lamina insertion differs for individual lamins. We have also observed that lamin A is synthesized as an apparent precursor molecule that is converted to mature lamin A only after integration into the lamina structure. The lamina is reversibly depolymerized during cell division, a process that may be mediated by enzymic phosphorylation of the lamins. To investigate this possibility further, we have analysed charge-altering modifications of the lamins on two-dimensional gels, and have found that phosphorylation is the only detectable modification of these proteins that occurs specifically during mitosis. Furthermore, we have determined that when the lamins are disassembled during metaphase, each lamin has approximately 2 moles of associated phosphate/mole lamin, a value that is four to sevenfold higher than the average interphase level. Considering this information, we discuss a model by which depolymerization and reassembly of the lamina can regulate the reversible disassembly of the nuclear envelope during mitosis.