Krisenko Mariya O, Cartagena Alexander, Raman Arvind, Geahlen Robert L
Department of Medicinal Chemistry and Molecular Pharmacology, ‡School of Mechanical Engineering, §Purdue Center for Cancer Research, and ∥Birck Nanotechnology Center, Purdue University , West Lafayette, Indiana 47907, United States.
Biochemistry. 2015 Jan 13;54(1):60-8. doi: 10.1021/bi500325n. Epub 2014 Jun 18.
The Syk protein-tyrosine kinase, a well-characterized modulator of immune recognition receptor signaling, also plays important, but poorly characterized, roles in tumor progression, acting as an inhibitor of cellular motility and metastasis in highly invasive cancer cells. Multiharmonic atomic force microscopy (AFM) was used to map nanomechanical properties of live MDA-MB-231 breast cancer cells either lacking or expressing Syk. The expression of Syk dramatically altered the cellular topography, reduced cell height, increased elasticity, increased viscosity, and allowed visualization of a more substantial microtubule network. The microtubules of Syk-expressing cells were more stable to nocodazole-induced depolymerization and were more highly acetylated than those of Syk-deficient cells. Silencing of MAP1B, a major substrate for Syk in MDA-MB-231 cells, attenuated Syk-dependent microtubule stability and reversed much of the effect of Syk on cellular topography, stiffness, and viscosity. This study illustrates the use of multiharmonic AFM both to quantitatively map the local nanomechanical properties of living cells and to identify the underlying mechanisms by which these properties are modulated by signal transduction machinery.
脾酪氨酸激酶（Syk）是一种已被充分表征的免疫识别受体信号调节因子，在肿瘤进展中也发挥着重要但尚未完全明确的作用，它在高侵袭性癌细胞中作为细胞运动和转移的抑制剂。利用多谐波原子力显微镜（AFM）绘制了缺乏或表达Syk的活MDA-MB-231乳腺癌细胞的纳米力学特性。Syk的表达显著改变了细胞形貌，降低了细胞高度，增加了弹性和粘度，并使更丰富的微管网络得以可视化。表达Syk的细胞中的微管对诺考达唑诱导的解聚更稳定，并且比缺乏Syk的细胞中的微管乙酰化程度更高。在MDA-MB-231细胞中，Syk的主要底物MAP1B的沉默减弱了Syk依赖的微管稳定性，并逆转了Syk对细胞形貌、硬度和粘度的大部分影响。本研究说明了多谐波AFM在定量绘制活细胞局部纳米力学特性以及识别这些特性受信号转导机制调节的潜在机制方面的应用。
