Using Light to Increase cAMP

Science Signaling  18 Jan 2011:
Vol. 4, Issue 156, pp. ec17
DOI: 10.1126/scisignal.4156ec17

Noting the success of the light-sensitive protein channelrhodopsin in providing a nontoxic, inheritable mechanism for the selective manipulation of cell membrane potential, Stierl et al. were interested in exploring the possibilities of using such light-sensitive proteins to manipulate other cellular parameters. They identified a sequence in the genome of the bacterium Beggiatoa as likely encoding a protein with a light-sensing BLUF (blue light receptor using FAD) domain linked directly to a type III adenylyl cyclase. When cloned into a cyclic adenosine monophosphate (cAMP)–deficient strain of Escherichia coli, the putative Beggiatoa photoactivated adenylyl cyclase (bPAC) rescued maltose metabolism, indicating that it was a functional adenylyl cyclase. Expression of bPAC in Xenopus oocytes enabled the light-induced activation of currents through coexpressed cyclic nucleotide-gated cation (CNG) channels and, more slowly, through coexpressed cystic fibrosis transmembrane conductance regulator (CFTR) anion channels, which are responsive to phosphorylation by cAMP-dependent protein kinase. ELISA assays revealed that oocytes expressing bPAC had increased cAMP relative to control oocytes and that a 1-min light pulse elicited a marked increase in cAMP. Purified bPAC was highly sensitive to light; moreover, its activity could be stimulated 300-fold by light (from about 33 pmol of cAMP/min/mg of protein in the dark to a maximal activity of about 10 nm of cAMP/min/mg of protein). When expressed in pyramidal neurons in vitro, bPAC enabled the activation of currents through coexpressed CNG channels by 100-ms pulses of dim light. Light-evoked currents appeared more rapidly than did currents in response to treatment with forskolin and IBMX (respectively, an activator of adenylyl cyclase and an inhibitor of phosphodiesterase). In transgenic Drosophila, bPAC under the control of a neuron-specific promoter conferred light-dependent cessation of grooming behavior. The authors thus conclude that bPAC represents a promising tool for optogenetic manipulation of cAMP, with various advantages compared with the previously identified Euglena gracilis PACα, including smaller size, less activity in the dark, and better stimulation by light.

M. Stierl, P. Stumpf, D. Udwari, R. Gueta, R. Hagedorn, A. Losi, W. Gärtner, L. Petereit, M. Efetova, M. Schwarzel, T. G. Oertner, G. Nagel, P. Hegemann, Light modulation of cellular cAMP by a small bacterial photoactivated adenylyl cyclase, bPAC, of the soil bacterium Beggiatoa. J. Biol. Chem. 286, 1181–1188 (2011). [Abstract] [Full Text]