Bipolar disorder tends to run in families, so researchers are looking for genes that may increase a person’s chance of developing the illness. The Bipolar Disorder Phenome Database was established to link visible signs of the illness with the genes that may influence them. Researchers have used this database to discover environmental and genetic factors that are associated with the disorder.
More recently, an international collaboration of researchers have discovered two new gene regions which show a connection with bipolar disorder susceptibility. Our project is most interested in one of these gene regions, namely adenylate cyclase 2 (ADCY2), a key enzyme in cAMP signalling. This is an important biological pathway in cell communication and specifically, adenylate cyclases (EC 184.108.40.206) are central to hormone responses, playing a pivotal role in signal transduction following stimulation of G-protein coupled receptors (Zhang, 1997).
PDB 1AB8 (ADCY2 rat brain)
Adenylate cyclase 2
Mühleisen (2014) discovered two genome-wide significant SNPs (single nucleotide polymorphisms) located in the ADCY2 gene on chromosome 5p15.31. This chromosome is expressed in the brain and encodes a cell membrane-bound enzyme for the synthesis of the second-messenger molecule cAMP. The transcript contains 6575 base pairs and 25 exons that encode 1091 amino acid residues. rs17826816 is located in intron 2 of the largest protein-encoding transcript while rs13166360 is located in exon 3 where it mediates an amino acid change from nonpolar Val to hydrophobic Leu at position 147. rs13166360 impacts on the fourth α-helix of the first transmembrane domain. This missense mutation is predicted to have a damaging effect on the protein.
The ADCY2 isoform belongs to the Class III of adenylate cyclases (AC). This class of ACs typically has 12 transmembrane segments: 6 membrane segments M1, then the C1 cytoplasmic domain, then another 6 transmembrane segments M2, then a second cytoplasmic domain C2. The well-conserved C1a and C2a subdomains are homologous to each other and form an intramolecular dimer that forms the active site.
The numerous AC isoforms catalyze the conversion of ATP to cAMP and pyrophosphate (PPi). Biochemical and crystallographic studies show that this enzymatic process requires Mg2+ and Mn2+ as cofactors. ACs are activated by the stimulatory G-protein α subunit (Gαs) and other regulatory molecules.
AC-catalyzed reaction converting ATP to cAMP
The catalytic site resides at the interface of two cyclase homology domains (CHD). The model for the prototypical catalytic core is derived from crystal structures of the complex formed by the C1 CHD domain from ADCY5 and the C2 CHD domain from ADCY2 (VC1: IIC2) bound to two activators, namely forskolin (FSK) and GTPγS-activated Gαs. Structural studies of the activated VC1: IIC2 complex show that upon binding to potent substrate analogues, ACs undergo a transition from an “open” to “closed” conformation by moving several structural elements in both CHDs toward the catalytic site (Mou, 2009).
PDB 3C16 (VC1:IIC2 complex with ATP and Ca)
In 1971 Earl Sutherland was awarded the Nobel Prize in Physiology or Medicine “for his discoveries concerning the mechanisms of the action of hormones”. His work included insights into the biological activity of cAMP. He also discovered the key role of AC-III in human liver, where adrenaline indirectly stimulates AC to mobilise energy in the “fight or flight” response. The effect of adrenaline is via a G-protein signalling cascade, which transmits chemical signals from outside the cell across the membrane to the cytoplasm. Adrenaline binds to a receptor, which transmits a signal to the G-protein, which transmits a signal to the AC, which transmits a signal by converting ATP to cAMP (Sutherland, 1970).
Nobel Prize winner Dr Earl Sutherland
Optogenetics is a rapidly evolving field that allows optical control of genetically targeted biological systems at high temporal and spatial resolution (Rein, 2012). It was also named by Nature Methods as its Method of the Year 2010. Optogenetic “tools” (the most notable being channelrhodopsin-2) are of microbial origin and can readily target neurons within heterogeneous tissue, selectively inactivating just one type of neurons while leaving the others more or less unaltered (Fenno, 2011). As the optogenetic toolbox evolves and diversifies, the opportunities for neuroscience continue to grow. The YouTube clip below shows how scientists can control the behaviour of cells simply by switching on a blue light.
Along with channelrhodopsin-2, photoactivatable adenylate cyclase (PAC) is another optogenetic tool that shows significant promise in the field of neuroscience. PACs were fist discovered in E. gracilis and can be expressed in other organisms through genetic engineering. Shining a blue light on a cell containing PAC activates it and abruptly increases the rate of conversion of ATP to cAMP. This allows us to study the effect of increasing intracellular cAMP levels in particular neurons and how this increase in neural activity affects the behaviour of an organism (Stierl, 2011).
Bipolar Disorder Phenome Database. National Institute of Mental Health
Structure of the adenylyl cyclase catalytic core. Nature
Genome-wide association study reveals two new risk loci for bipolar disorder. Nature Communications
Structural basis for inhibition of mammalian adenylyl cyclase by calcium. Biochemistry
On the biological role of cyclic AMP. The Journal of the American Medical Association
The optogenetic (r)evolution. Molecular Genetics and Genomics
The development and application of optogenetics. Annual Review of Neuroscience
Light modulation of cellular cAMP by a small bacterial photoactivated adenylyl cyclase, bPAC, of the soil bacterium Beggiatoa. Journal of Biological Chemistry