Numerical method for vesicle movement analysis in a complex cytoskeleton network

Abstract

The detection of the precise movement of a vesicle during transport in a live cell provides key information for the intracellular delivery process. Here we report a novel numerical method for analyzing three-dimensional vesicle movement. Since the vesicle moves along a linear cytoskeleton during the active transport, our method first detects the orientation and position of the cytoskeleton as a linear section based on angle correlation and linear regression, after noise reduction. Then, the precise vesicle movement is calculated using vector analysis, in terms of rotation angle and translational displacement. Using this method, various vesicle trajectories obtained via high spatiotemporal resolution microscopy can be understood..

3D rotational motion of an endocytic vesicle on a complex microtubule network in a living cell

“…For the vesicle trajectory data, the linear sections were detected where the local curvature of the trajectory was greater than 3π 4 , after removing the noise by applying the Gaussian noise filter (window size = 16 data points). The orientation and position of the microtubule for each detected linear section were estimated by using principal component analysis (PCA) (21).…”

“…However, since microtubules form a three-dimensional network structure, measurement of three dimensional angles considering the geometry of the microtubules in a living cell is required. To measure accurate angles between microtubules in the network structure, the locations of microtubules were estimated from the 3D trajectory of vesicles that apparently interacted with a multiple number of microtubules, based on our numerical method (21). To do this, first, the microtubules labeled by GFP in a living cell were imaged at 1 frame per second (fps) for 60 s (Fig.…”

“…For each trajectory of vesicle that we detected on the microtubule, active transport sections were distinguished as partial linear domains based on the local curvature, and the locations of microtubules were estimated for these linear sections, by using our numerical analysis method (21). A representative vesicle movement on a microtubule was reconstructed in three dimensions, and the data points in the active transport section were colored in pink, while the intermittent pauses or direction changes between active transports were colored in mint green ( Fig.…”

“…(C) φ 2D represents the angle that can be roughly estimated from 2D tracking data. (D) φ 3D indicates the angle between the 3D segments acquired by numerical analysis of vesicle tracking data based on principal component analysis for detected linear sections (21).…”

“…S1), achieved by our microscope stage position feedback system (22) that guarantees approximately 3 nm of x, y and 5 nm of axial spatial precision at 100 frames per second. First, the 3D angles between adjacent microtubules where the vesicle sequentially interacted are calculated based on our novel numerical analysis method reported previously (21). Additionally, the relationship between the transfer angle and the time scale is investigated.…”