Tracking the motion of single nanoparticles inside living cells: New insights into intracellular crowdedness

生物分子如何在活细胞内在空间和时间中移动，不仅是一个基本的研究问题，而且是理解许多生物过程的关键。随着先进的单粒子跟踪技术的发展，很明显，细胞内的生物分子表现出复杂的运动类型，这不能简单地描述为随机的热扩散或定向运动通过分子马达。这是因为细胞内的细胞质中充满了大的生物分子和细胞器，以及细胞骨架蛋白质细丝的异质网络。深入了解活细胞内生物分子运动的关键是开发能够以高定位精度和速度在3D中跟踪单个粒子的新技术，以及能够根据物理定律描述实验的数学模型，在这个项目中，你将应用一种先进的激光显微光谱技术，称为共振四波混频（resonant Four-Wave Mixing），由监督团队首创，以纳米级的精度在3D中成像和跟踪细胞内的单个小金纳米颗粒。纳米颗粒将被显微注射到卵子（卵母细胞）中。这些是由监督小组积极研究的大细胞（来自哺乳动物和无脊椎动物）。它们的ATP水平和代谢率与它们成熟、受精和进一步发育成优质胚胎的能力密切相关。值得注意的是，粒子的运动可以受到ATP驱动的分子马达的影响。因此，对这些细胞内运动的研究将成为监测胚胎代谢和预测存活能力的新技术的平台。在不同的代谢条件下，将在卵子中测量实验性单粒子轨迹，并将其与扩散的数学模型进行比较，反映出监督团队在数学和计算方法方面的专业知识。测量的时间轨迹将揭示与拥挤环境相关的随机、定向、瞬时停滞和约束运动的组合。目的是在环境的纳米结构和粒子运动之间建立因果关系。这反过来又会刺激假设被实验验证，并将提供对细胞内部前所未有的洞察力。

How biomolecules move inside living cells, in space and time, is not only a fundamental research question but is key to the understanding of many biological processes. With the development of advanced single-particle tracking techniques, it has become apparent that biomolecules inside cells exhibit complicated types of motion, which cannot be simply described as random from thermal diffusion or directed motion via molecular motors. This is because the cell contains a cytoplasm crowded with large biomolecules and organelles, and a heterogeneous network of cytoskeletal protein filaments.Key to the in depth understanding of biomolecular motion inside living cells is the development of new techniques capable to track single particles in 3D with high localisation precision and speed, alongside mathematical models that can describe the experiments according to physics' law, and eventually reveal new insights into the highly complex living cell's interior.In this project, you will apply an advanced laser micro-spectroscopy technique called resonant Four-Wave Mixing, pioneered by the supervisory team, to image and track single small gold nanoparticles background free inside cells with precision at the nanoscale in 3D. Nanoparticles will be micro-injected into eggs (oocytes). These are large cells actively studied by the supervisory team (from mammals and invertebrates). They have ATP levels and metabolic rates closely linked to their ability to mature, be fertilised, and further develop into good quality embryos. Notably, movement of particles can be influenced by molecular motors that are ATP-driven. Hence, studies of intracellular motion in these cells will be a platform for new technologies to monitor embryo metabolism and predict viability.Experimental single particle trajectories will be measured in eggs under different metabolic conditions, and will be compared with mathematical models of diffusion, reflecting expertise in mathematical and computational methods by the supervisory team. Measured time trajectories will reveal combinations of random, directed, transiently stalled and constrained motions related to crowded environments. The aim will be to develop a causal link between the nanostructure of the environment and particle motion. This in turn will stimulate hypothesis to be verified experimentally, and will provide an unprecedented insight on the cell's interior.