Rux Caleb J, Chong Megan K, Myers Sean, Cho Nathan H, Dumont Sophie
Department of Bioengineering & Therapeutic Sciences, San Francisco, CA 94158, USA.
UC Berkeley-UCSF Graduate Program in Bioengineering, San Francisco, CA 94158, USA.
bioRxiv. 2025 Jun 6:2025.06.05.657915. doi: 10.1101/2025.06.05.657915.
To segregate chromosomes at cell division, the spindle must maintain its structure under force. How it does so remains poorly understood. To address this question, we use microneedle manipulation to apply local force to spindle microtubule bundles, kinetochore-fibers (k-fibers), inside mammalian cells. We show that local load directly fractures k-fibers, and that newly created plus-ends often have arrested dynamics, resisting depolymerization. Force alone, without fracture, is sufficient for spindle microtubule stabilization, as revealed by laser ablating k-fibers under local needle force. Doublecortin, which binds a compacted microtubule lattice, is lost around the force application site, suggesting local force-induced structural remodeling. In turn, EB1, which recognizes GTP-tubulin, is locally enriched at stabilization sites, both before and after force-induced fracture. Together, our findings support a model where force-induced damage leads to local spindle microtubule lattice remodeling and stabilization, which we propose reinforces the spindle where it experiences critical loads.
为了在细胞分裂时分离染色体,纺锤体必须在受力情况下维持其结构。其具体机制仍知之甚少。为了解决这个问题,我们使用微针操作对哺乳动物细胞内的纺锤体微管束,即动粒微管(k纤维)施加局部力。我们发现局部负载会直接使k纤维断裂,并且新产生的正端通常具有停滞的动力学,抵抗解聚。如在局部针力作用下激光消融k纤维所显示的,仅有力而无断裂就足以使纺锤体微管稳定。与紧密微管晶格结合的双皮质素在力施加部位周围消失,表明局部力诱导了结构重塑。反过来,识别GTP微管蛋白的EB1在力诱导断裂前后均在稳定部位局部富集。总之,我们的研究结果支持一种模型,即力诱导的损伤导致局部纺锤体微管晶格重塑和稳定,我们认为这加强了纺锤体承受关键负载的部位。