European Molecular Biology Laboratory, 69117 Heidelberg, Germany.
J Cell Biol. 2012 Sep 17;198(6):981-90. doi: 10.1083/jcb.201205103.
Disassembly of the nuclear lamina is a key step during open mitosis in higher eukaryotes. The activity of several kinases, including CDK1 (cyclin-dependent kinase 1) and protein kinase C (PKC), has been shown to trigger mitotic lamin disassembly, yet their precise contributions are unclear. In this study, we develop a quantitative imaging assay to study mitotic lamin B1 disassembly in living cells. We find that CDK1 and PKC act in concert to mediate phosphorylation-dependent lamin B1 disassembly during mitosis. Using ribonucleic acid interference (RNAi), we showed that diacylglycerol (DAG)-dependent PKCs triggered rate-limiting steps of lamin disassembly. RNAi-mediated depletion or chemical inhibition of lipins, enzymes that produce DAG, delayed lamin disassembly to a similar extent as does PKC inhibition/depletion. Furthermore, the delay of lamin B1 disassembly after lipin depletion could be rescued by the addition of DAG. These findings suggest that lipins activate a PKC-dependent pathway during mitotic lamin disassembly and provide evidence for a lipid-mediated mitotic signaling event.
核层板的解体是高等真核生物有丝分裂中一个关键步骤。已有研究表明，包括细胞周期蛋白依赖性激酶 1（CDK1）和蛋白激酶 C（PKC）在内的几种激酶的活性可触发有丝分裂核层板的解体，但它们的确切贡献尚不清楚。在这项研究中，我们开发了一种定量成像测定法来研究活细胞中的有丝分裂核层板 B1 解体。我们发现 CDK1 和 PKC 协同作用，介导有丝分裂过程中磷酸化依赖性核层板 B1 的解体。通过核糖核酸干扰（RNAi），我们发现二酰基甘油（DAG）依赖性 PKC 触发了核层板解体的限速步骤。RNAi 介导的 Lipin 耗竭或化学抑制，Lipin 是产生 DAG 的酶，可使核层板解体延迟到与 PKC 抑制/耗竭相似的程度。此外，在 Lipin 耗竭后，添加 DAG 可挽救核层板 B1 解体的延迟。这些发现表明，Lipin 在有丝分裂核层板解体过程中激活了一个依赖于 PKC 的途径，并为脂质介导的有丝分裂信号事件提供了证据。
