FRET detection of cellular α4-integrin conformational activation
FRET detection of cellular α4-integrin conformational activation
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DOI：
10.1016/s0006-3495(03)74809-7
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发表时间：
2003-12-01
影响因子：
3.4
通讯作者：
Sklar, LA
中科院分区：
文献类型：
--
作者：
Chigaev, A;Buranda, T;Sklar, LA
整合素是细胞粘附受体，在每一种细胞类型上都有表达，被认为在激活后会发生构象变化。在这里，通过将α(4)-整合素暴露于二价离子或使用趋化因子受体由内向外激活产生不同的亲和力状态。我们利用荧光共振能量转移（FRET）技术在α(4)-整合素特异性结合的肽供体和整合到质膜上的十八烷基罗丹明B受体之间研究了活细胞上整合素的动态结构转化。我们使用一个模型来分析数据，该模型描述了在无限平面上随机分布的供体和受体之间的FRET。最接近的距离随整合素的亲和力而变化。静息与Mn2+激活受体之间的最接近距离变化接近50埃，趋化因子激活后接近25埃。我们使用共聚焦显微镜来探测整合素激活后供体和受体的横向组织。综上所述，FRET和共聚焦的结果表明，FRET效率的变化主要是由于垂直延伸的整合素。α(4)-整合素的延伸与其亲和力之间的协调为Dembo的catch-bond概念提供了一种机制。
Integrins are cell adhesion receptors, expressed on every cell type, that have been postulated to undergo conformational changes upon activation. Here, different affinity states were generated by exposing alpha(4)-integrins to divalent ions or by inside-out activation using a chemokine receptor. We probed the dynamic structural transformation of the integrin on live cells using fluorescence resonance energy transfer (FRET) between a peptide donor, which specifically binds to the alpha(4)-integrin, and octadecyl rhodamine B acceptors incorporated into the plasma membrane. We analyzed the data using a model that describes FRET between a random distribution of donors and acceptors in an infinite plane. The distance of closest approach was found to vary with the affinity of the integrin. The change in distance of closest approach was similar to50 Angstrom between resting and Mn2+ activated receptors and similar to25 Angstrom after chemokine activation. We used confocal microscopy to probe the lateral organization of donors and acceptors subsequent to integrin activation. Taken together, FRET and confocal results suggest that changes in FRET efficiencies are primarily due to the vertical extension of the integrin. The coordination between the extension of alpha(4)-integrin and its affinity provides a mechanism for Dembo's catch-bond concept.