Abstract
Vapor nanobubble (VNB) photoporation is a physical method for intracellular delivery that has gained significant interest in the past decade. It has successfully been used to introduce molecular cargo of diverse nature into different cell types with high throughput and minimal cytotoxicity. For translational purposes, it is important to understand whether and how photoporation affects cell homeostasis. To obtain a comprehensive view on the transcriptional rewiring that takes place after VNB photoporation, we performed a longitudinal shotgun RNA-sequencing experiment. Six hours after photoporation, we found a marked upregulation of LMNA transcripts as well as their protein products, the A-type lamins. At the same time point, we observed a significant increase in several heterochromatin marks, suggesting a global stiffening of the nucleus. These molecular features vanished 24 h after photoporation. Since VNB-induced chromatin condensation was prolonged in LMNA knockout cells, A-type lamins may be required for restoring the nucleus to its original state. Selective depletion of A-type lamins reduced cell viability after VNB photoporation, while pharmacological stimulation of LMNA transcription increased the percentage of successfully transfected cells that survived after photoporation. Therefore, our results suggest that cells respond to VNB photoporation by temporary upregulation of A-type lamins to facilitate their recovery.
Highlights
Efficient and safe introduction of exogenous cargo into the cytoplasm of the cell is an unrelenting quest
The transcriptional response to vapor nanobubble (VNB) photoporation and photothermal heating, we only found a limited number of shared differentially expressed genes (DEGs)
We confirmed upregulation of A-type lamins at the protein level 6 h after VNB treatment, but we found this upregulation to be rather transient, as protein levels returned to baseline after 24 h
Summary
Efficient and safe introduction of exogenous cargo into the cytoplasm of the cell is an unrelenting quest. At a sufficiently high laser fluence, the AuNP temperature exceeds the critical temperature of the surrounding liquid, causing it to vaporize. This results in the formation of VNBs around the AuNP of which the subsequent collapse leads to mechanical shock waves that generate transient pores in the plasma membrane. While both photothermal heating and VNB formation have been successfully used to permeabilize cell membranes, the latter is more often used as it leads to more efficient intracellular delivery than photothermal heating in combination with the typically used AuNPs [11]. Delivery can be achieved with high throughput and spatiotemporal control [13]
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