Guizzunti Gianni, Seemann Joachim
Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390.
Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390
Proc Natl Acad Sci U S A. 2016 Oct 25;113(43):E6590-E6599. doi: 10.1073/pnas.1610844113. Epub 2016 Oct 10.
During mitosis, the mammalian Golgi vesiculates and, upon partitioning, reassembles in each daughter cell; however, it is not clear whether the disassembly process per se is important for partitioning or is merely an outcome of mitotic entry. Here, we show that Golgi vesiculation is required for progression to metaphase. To prevent Golgi disassembly, we expressed HRP linked to a Golgi-resident protein and acutely triggered the polymerization of 3,3'-diaminobenzidine (DAB) in the Golgi lumen. The DAB polymer does not affect interphase cell viability, but inhibits Golgi fragmentation by nocodazole and brefeldin A and also halts cells in early mitosis. The arrest is Golgi specific and does not occur when DAB is polymerized in the endosomes. Cells with a DAB polymer in the Golgi enter mitosis normally but arrest with an intact Golgi clustered at a monopolar spindle and an active spindle assembly checkpoint (SAC). Mitotic progression is restored upon centrosome depletion by the Polo-like kinase 4 inhibitor, centrinone, indicating that the link between the Golgi and the centrosomes must be dissolved to reach metaphase. These results demonstrate that Golgi disassembly is required for mitotic progression because failure to vesiculate the Golgi activates the canonical SAC. This requirement suggests that cells actively monitor Golgi integrity in mitosis.
在有丝分裂过程中,哺乳动物的高尔基体形成囊泡,并在分裂时在每个子细胞中重新组装;然而,目前尚不清楚高尔基体的解体过程本身对分裂是否重要,还是仅仅是进入有丝分裂的一个结果。在这里,我们表明高尔基体囊泡化是进入中期所必需的。为了防止高尔基体解体,我们表达了与高尔基体驻留蛋白相连的辣根过氧化物酶(HRP),并在高尔基体腔中急性触发3,3'-二氨基联苯胺(DAB)的聚合。DAB聚合物不影响间期细胞的活力,但能抑制诺考达唑和布雷菲德菌素A诱导的高尔基体碎片化,还能使细胞停滞在有丝分裂早期。这种停滞是高尔基体特异性的,当DAB在内体中聚合时不会发生。高尔基体中有DAB聚合物的细胞正常进入有丝分裂,但会因完整高尔基体聚集在单极纺锤体上且纺锤体组装检查点(SAC)激活而停滞。通过Polo样激酶4抑制剂centrinone耗尽中心体后,有丝分裂进程得以恢复,这表明高尔基体与中心体之间的联系必须解除才能进入中期。这些结果表明,高尔基体解体是有丝分裂进程所必需的,因为高尔基体未能形成囊泡会激活经典的SAC。这一需求表明细胞在有丝分裂过程中会积极监测高尔基体的完整性。