Spatio-temporal Dynamics and Mechanisms of Stress Granule Assembly

Stress granules (SGs) are non-membranous cytoplasmic aggregates of mRNAs and related proteins, assembled in response to environmental stresses such as heat shock, hypoxia, endoplasmic reticulum (ER) stress, chemicals (e.g. arsenite), and viral infections. SGs are hypothesized as a loci of mRNA triage and/or maintenance of proper translation capacity ratio to the pool of mRNAs. In brain ischemia, hippocampal CA3 neurons, which are resilient to ischemia, assemble SGs. In contrast, CA1 neurons, which are vulnerable to ischemia, do not assemble SGs. These results suggest a critical role SG plays in regards to cell fate decisions. Thus SG assembly along with its dynamics should determine the cell fate. However, the process that exactly determines the SG assembly dynamics is largely unknown. In this paper, analyses of experimental data and computer simulations were used to approach this problem. SGs were assembled as a result of applying arsenite to HeLa cells. The number of SGs increased after a short latent period, reached a maximum, then decreased during the application of arsenite. At the same time, the size of SGs grew larger and became localized at the perinuclear region. A minimal mathematical model was constructed, and stochastic simulations were run to test the modeling. Since SGs are discrete entities as there are only several tens of them in a cell, commonly used deterministic simulations could not be employed. The stochastic simulations replicated observed dynamics of SG assembly. In addition, these stochastic simulations predicted a gamma distribution relative to the size of SGs. This same distribution was also found in our experimental data suggesting the existence of multiple fusion steps in the SG assembly. Furthermore, we found that the initial steps in the SG assembly process and microtubules were critical to the dynamics. Thus our experiments and stochastic simulations presented a possible mechanism regulating SG assembly.Author Summary: Cells suffer from various environmental stresses such as heat shock and viral infection. In response to a stress, small non-membranous cytoplasmic aggregates, stress granules (SGs), are assembled. SGs contain mRNAs and related proteins. Hippocampal CA1 neurons located in the brain, which are vulnerable to ischemia, do not assemble SGs, while CA3 neurons, which are resilient to ischemia, assemble SGs. The dysfunction of SGs has been reported in human diseases including pathogenic viral infection. These observations led to a hypothesis that SGs play an important role in cell fate decisions, and the dynamics of SG assembly would regulate cell fate. However, the conditions that determine the number and distribution of SGs in a cell in response to a stress are largely unknown. We approached this problem by experiments and simulations. Our stochastic simulations replicated the observations. Furthermore, we found that initial steps in the SG assembly process were important to the dynamics of SG assembly, and that SG size resembled the gamma distribution both in simulations and experiments, suggesting the existence of multiple steps in the SG assembly process. To the best of our knowledge, this work was the first to show SG assembly in a whole cell by stochastic simulations.