Cells can survive executioner caspase activation following transient apoptotic stimuli, a process called anastasis. Using whole-transcriptome RNA sequencing, Sun et al. show that anastasis is an active, two-stage program and characterize the cell behaviors and molecular signature involved in the process.
SUMMARY The essential organization of microtubules within neurons has been described, however, less is known about how neuronal actin is arranged and the functional implications of its arrangement. Here we describe in live cells an actin-based structure in the proximal axon that selectively prevents some proteins from entering the axon, while allowing the passage of others. Concentrated patches of actin in proximal axons are present shortly after axonal specification in rat and zebrafish neurons imaged live, and mark positions where anterogradely traveling vesicles carrying dendritic proteins halt and reverse. Patches colocalize with the ARP2/3 complex, and when ARP2/3-mediated nucleation is blocked a dendritic protein mislocalizes to the axon. Patches are highly dynamic, with few persisting longer than 30 minutes. In neurons in culture and in vivo, actin appears to form a contiguous, semipermeable barrier, despite its apparently sparse distribution, preventing axonal localization of constitutively active Myosin Va, but not Myosin VI.
The localization of mRNAs within axons and dendrites allows neurons to manipulate protein levels in a time and location dependent manner and is essential for processes such as synaptic plasticity and axon guidance. However, an essential step in the process of mRNA localization, the decision to traffic to dendrites and/or axons, remains poorly understood. Here we show that Myosin Va and actin filaments are necessary for the dendritic localization of the mRNA binding protein Staufen 1 and of mRNA encoding the microtubule binding protein Map2. Blocking the function or expression of Myosin Va or depolymerizing actin filaments leads to localization of Staufen 1 and of Map2 mRNA in both axons and dendrites. Furthermore, interaction with Myosin Va plays an instructive role in the dendritic localization of Hermes 1, an RNA binding protein. Wild-type Hermes 1 localizes to both axons and dendrites, whereas Hermes 1 fused with a Myosin Va binding peptide, localizes specifically to dendrites. Thus, our results suggest that targeting of mRNAs to the dendrites is mediated by a mechanism that is dependent on actin and Myosin Va.
Executioner-caspase activation has been considered a point-of-no-return in apoptosis. However, numerous studies report survival from caspase activation after treatment with drugs or radiation. An open question is whether cells can recover from direct caspase activation without pro-survival stress responses induced by drugs. To address this question, we engineered a HeLa cell line to express caspase-3 inducibly and combined it with a quantitative caspase activity reporter. While high caspase activity levels killed all cells and very low levels allowed all cells to live, doses of caspase activity sufficient to kill 15 to 30% of cells nevertheless allowed 70 to 85% to survive. At these doses, neither the rate, nor the peak level, nor the total amount of caspase activity could accurately predict cell death versus survival. Thus, cells can survive direct executioner-caspase activation, and variations in cellular state modify the outcome of potentially lethal caspase activity. Such heterogeneities may underlie incomplete tumor cell killing in response to apoptosis-inducing cancer treatments.
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