Actin polymerization driven mitochondrial transport in mating S. cerevisiae

Abstract

The dynamic microenvironment of cells depends on macromolecular architecture, equilibrium fluctuations, and nonequilibrium forces generated by cytoskeletal proteins. We studied the influence of these factors on the motions of mitochondria in mating S. cerevisiae using Fourier imaging correlation spectroscopy (FICS). Our measurements provide detailed length-scale dependent information about the dynamic behavior of mitochondria. We investigate the influence of the actin cytoskeleton on mitochondrial motion and make comparisons between conditions in which actin network assembly and disassembly is varied either by using disruptive pharmacological agents or mutations that alter the rates of actin polymerization. Under physiological conditions, nonequilibrium dynamics of the actin cytoskeleton leads to 1.5-fold enhancement of the long-time mitochondrial diffusion coefficient and a transient subdiffusive temporal scaling of the mean-square displacement (MSD proportional, variant tau (alpha), with alpha = 2/3). We find that nonequilibrium forces associated with actin polymerization are a predominant factor in driving mitochondrial transport. Moreover, our results lend support to an existing model in which these forces are directly coupled to mitochondrial membrane surfaces.

Fig. 1.

(A) Composite fluorescence micrograph of mitochondria in mating yeast cells. (B) and (C) Digital image analysis of sequenced images show that the majority of mitochondria are immobilized on the time scale of successive image frames (20 s/frame).

Fig. 2.

Comparison between TCFs for mitochondrial fluctuations in healthy cells (MYY290, black) and cells depleted of MTs (blue), and cells depleted of MFs (green). Each panel represents the indicated length scale probed by the FICS measurement. In panel (C) we include error bars in our decays. The insets show difference TCFs between drug-treated and healthy cells to emphasize the effects of the cytoskeletal inhibitors.

Fig. 3.

MSD of mitochondrial fluctuations calculated from the data presented in Fig. 2. Color assignments are the same as in that figure. A control measurement of a 0.5 μm colloid sample in viscous, concentrated sorbitol solution is presented at the top of the figure. Various lines with slopes representing the temporal scaling parameter α are also provided to guide the eye.

Fig. 4.

(A) Comparison between mitochondrial MSDs for the temperature-sensitive actin mutant strain act1-V159N (DDY1493, gray curves) and its wild-type parent ACT1 (DDY1495, black curves), measured at the nonpermissive temperature 36° C. (B) Similar comparison between MSDs for the deletion mutant strain Arc18pΔ (gray curves) and its wild-type parent BY4741 (ARC18, black curves).