A role for the p34cdc2 kinase and phosphatases in the regulation of phosphorylation and disassembly of lamin B2 during the cell cycle.

Abstract

While the p34cdc2 kinase is considered to be a critical regulator of mitosis, its function has not yet been directly linked to one of the key events during the onset of mitosis: nuclear envelope breakdown. Here we show that a major structural protein of the nuclear envelope, lamin B2, is phosphorylated by p34cdc2. Results from two‐dimensional phosphopeptide mapping experiments demonstrate that the p34cdc2‐specific phosphopeptides represent both mitotic and interphase specific phosphorylations of lamin B2 and include the major interphase phosphorylation site. In mitotic cells we detected two distinct forms of lamin B2 which differ in electrophoretic mobility and in degree of phosphorylation. The phosphorylation pattern of lamin B2 generated in vitro by p34cdc2 was more closely related to the less phosphorylated mitotic lamin B2, suggesting that another kinase(s) in addition to p34cdc2 is involved in generating the mitotic phosphorylation pattern. In addition, we show that treatment of interphase cells with okadaic acid, a potent phosphatase inhibitor, leads to the acquisition of mitosis‐specific phosphopeptides and can reversibly increase the detergent‐solubility of lamin B2. However, the M‐phase‐like phosphorylation of lamin B2 in itself is not sufficient to induce its disassembly from the nuclear lamina suggesting that an additional event(s) besides phosphorylation is required.

M-phase-specific phosphorylation and structural rearrangement of the cytoplasmic cross-linking protein plectin involve p34cdc2 kinase.

“…Best understood among p34cdc2-dependent events involving the reorganization of structural components is the breakdown of the nuclear envelope at the onset of mitosis. Several lines of evidence, including in vivo and in vitro studies, indicate that direct phosphorylation of lamins by p34cdc2/cyclin B causes the disassembly of the nuclear lamina (Heald and McKeon, 1990;Peter et al, 1990Peter et al, , 1991Ward and Kirschner, 1990;Dessev et al, 1991;Enoch et al, 1991;Luscher et al, 1991). Also, cytoplasmic IFs have been described as substrates for p34cdc2kinase.…”

Observation of nuclei reassembled from demembranated Xenopus sperm nuclei and analysis of their lamina components

“…Extracts derived from activated Xenopus laevis eggs able to asemble nuclei around exogeneous naked DNA, isolated chromosomes and demembranated sperm chromatin have been widely used to study the pathways and biochemistry of nuclear envelope and pore complexes assembly, lamina formation, DNA replication and nuclear cytoplasmic transportation [3][4][5][6]. Meanwhile, extracts prepared from mataphase eggs were adopted to study spindle organization and nuclear disassembly [7][8][9][10]. Plenty of information on cell cycle regulation concerning nuclear reassembly and mitosis has also been obtained from the studies using cell-free system [9,10].…”

“…Meanwhile, extracts prepared from mataphase eggs were adopted to study spindle organization and nuclear disassembly [7][8][9][10]. Plenty of information on cell cycle regulation concerning nuclear reassembly and mitosis has also been obtained from the studies using cell-free system [9,10]. Since 1989, colleagues Cell Research(1994), 4,[163][164][165][166][167][168][169][170][171][172] in our laboratory have devoted to this field and achieved a number of results [11][12][13][14].…”

Dynamic properties of nuclear lamins: lamin B is associated with sites of DNA replication.

“…Furthermore, phosphorylation of lamin B by p34 ~2 prevents its self-association in vitro (Peter et al, 1991). There is also evidence for other mitotic lamin kinases which have yet to be identified (Luscher et al, 1991). Other well-characterized kinases such as protein kinase C and protein kinase A have been shown to influence lamin assembly during interphase in different ways (Fields et al, 1987;Lamb et ai., 1991;Lourim and Lim, 1992;Hennekes et ai., 1993).…”