Ianoul Anatoli, Grant Donna D, Rouleau Yanouchka, Bani-Yaghoub Mahmud, Johnston Linda J, Pezacki John Paul
The Steacie Institute for Molecular Sciences, National Research Council Canada, 100 Sussex Drive, Ottawa, Canada K1A 0R6.
Nat Chem Biol. 2005 Sep;1(4):196-202. doi: 10.1038/nchembio726. Epub 2005 Aug 7.
The contraction of cardiac myocytes is initiated by ligand binding to adrenergic receptors contained in nanoscale multiprotein complexes called signalosomes. The composition and number of functional signalosomes within cardiac myocytes defines the molecular basis of the response to adrenergic stimuli. For the first time, we demonstrated the ability of near-field scanning optical microscopy to visualize beta-adrenergic receptors at the nanoscale in situ. On H9C2 cells, mouse neonatal and mouse embryonic cardiac myocytes, we showed that functional receptors are organized into multiprotein domains of approximately 140 nm average diameter. Colocalization experiments in primary cells at the nanometer scale showed that 15-20% of receptors were preassociated in caveolae. These nanoscale complexes were sufficient to effect changes in ligand-induced contraction rate without the requirement for substantial changes in receptor distribution in the cellular membrane. Using fluorescence intensities associated with these nanodomains, we estimated the receptor density within the observed nanometer features and established a lower limit for the number of receptors in the signalosome.
心肌细胞的收缩是由配体与称为信号小体的纳米级多蛋白复合物中所含的肾上腺素能受体结合引发的。心肌细胞内功能性信号小体的组成和数量决定了对肾上腺素能刺激反应的分子基础。我们首次证明了近场扫描光学显微镜在原位纳米尺度上可视化β-肾上腺素能受体的能力。在H9C2细胞、小鼠新生心肌细胞和小鼠胚胎心肌细胞上，我们表明功能性受体被组织成平均直径约为140 nm的多蛋白结构域。在原代细胞中进行的纳米尺度共定位实验表明，15%-20%的受体预先聚集在小窝中。这些纳米级复合物足以影响配体诱导的收缩速率，而无需细胞膜中受体分布发生实质性变化。利用与这些纳米结构域相关的荧光强度，我们估计了观察到的纳米特征内的受体密度，并确定了信号小体中受体数量的下限。
