Modulation of cyclic nucleotide-mediated cellular signaling and gene expression using photoactivated adenylyl cyclase as an optogenetic tool

Abstract

Cyclic nucleotide signaling pathway plays a significant role in various biological processes such as cell growth, transcription, inflammation, in microbial pathogenesis, etc. Modulation of cyclic nucleotide levels by optogenetic tools has overcome certain limitations of studying transduction cascade by pharmacological agents and has allowed several ways to modulate biological processes in a spatiotemporal manner. Here, we have shown the optogenetic modulation of the cyclooxygenase 2 (Cox-2) gene expression and their downstream effector molecule (PGE2) in HEK-293T cells and the development process of Dictyostelium discoideum via modulating the cyclic nucleotide (cAMP) signaling pathway utilizing photoactivated adenylyl cyclases (PACs) as an optogenetic tool. Light-induced activation of PACs in HEK-293T cells increases the cAMP level that leads to activation of cAMP response element-binding protein (CREB) transcription factor and further upregulates downstream Cox-2 gene expression and their downstream effector molecule prostaglandin E2. In D. discoideum, the light-regulated increase in cAMP level affects the starvation-induced developmental process. These PACs could modulate the cAMP levels in a light-dependent manner and have a potential to control gene expression and their downstream effector molecules with varying magnitude. It would enable one to utilize PAC as a tool to decipher cyclic nucleotide mediated signaling pathway regulations and their mechanism.

The C-terminal region affects the activity of photoactivated adenylyl cyclase from Oscillatoria acuminata

“…(bPAC) 5–10 , which have expanded the use of PAC as an optogenetic tool and in the control of intracellular cAMP levels by blue light in hippocampal neurons, Caenorhabditis elegans , pancreatic β-cells, etc. 7,10–14 .…”

Optogenetic modulation of real-time nanoscale dynamics of HCN channels using photoactivated adenylyl cyclases

“…PACs have also been successfully deployed in diverse biological systems to modulate the intracellular cAMP levels to control signal transduction of selected molecular pathways, to program cellular responses, to modulate cellular morphology and to control animal behavior. [19][20][21][22][23][24][25] Despite these advances, there has been limited understanding on how the local or global activation of PACs can control the lateral organization and real-time trafficking of transmembrane molecules such as HCN channels whose activity is directly controlled by cAMP. This is in part due to the lack of information on the characterization of biophysical properties of PACs in living cells and due to the lack of development of novel paradigms to investigate real-time changes in nanoscale localization and dynamics of molecules in combination with optogenetic approaches.…”

Establishment of Optogenetic Modulation of cAMP for Analyzing Growth, Biofilm Formation, and Virulence Pathways of Bacteria Using a Light-Gated Cyclase

“…The photoactivated adenylate cyclases (PACs) were genetically engineered into the heterologous systems and blue light illumination was used to alter the cAMP-mediated cellular and behavioral responses of the systems [13][14][15]. PACs possess a BLUF (blue light using flavin) domain-coupled adenylate cyclase and is utilized for blue light-dependent modulation of cAMP [16,17].…”

“…However, these large-sized (110 kDa) PACs showed significant cyclase activity in the dark. Tanwar and coworkers characterized smaller-sized (38 kDa) PAC, which showed fast kinetics for light-regulated cyclase activity [13]. PACs have been used for altering cAMP levels in Drosophila [15,17] and for finding the mechanism underlying cAMP-dependent axonal morphogenesis [14].…”