G protein-coupled inwardly-rectifying potassium (GIRK) channels

My current Master’s thesis research is focussed on understanding the structural and functional properties of G protein-coupled inwardly-rectifying potassium (GIRK or Kir 3.x) channels. This class of potassium ion channels are responsible for regulating the heart rate and modulating the neuronal excitability of certain neurons.

GIRK channels are activated by G-protein coupled receptors (GPCRs) including the muscarinic, dopamine, serotonin, GABA, opiod, and acetylcholine receptors, which are involved in many signal transduction pathways in the cell. The activation of a GPCR by its ligand (neurotransmitter or hormone) results in the release of Gα and Gβγ, two intracellular effector molecules. The activated Gβγ binds to the GIRK channel and opens it up to potassium ions resulting in the hyperpolarization of the cell (increased negative charge due to efflux of K+ ions).

Activation of GIRK channel
GIRK channels are activated upon GPCR stimulation by direct interaction with Gβγ.

Molecular cloning techniques have led to the discovery of four channel subunits – GIRK1 (Kir 3.1), GIRK2 (Kir 3.2), GIRK3 (Kir 3.3) and GIRK 4 (Kir 3.4). GIRK1 through 3 can be found in the central nervous system and GIRK4 is primarily found in the heart. Four of these subunits assemble either as homomers or heteromers (in 1:1 subunit ratio) to form a tetrameric functional channel.

Structurally, the channel is divided into cytoplasmic and transmembrane domains. The amino- (NH2) terminus and the carboxyl- (COOH) terminus are present in the cytoplasm and contribute to the formation of the intracellular/cytoplasmic domain. Each subunit is composed of two transmembrane domains separated by a P-loop containing the “ion-selectivity filter”. This type of channel assembly results in significant interactions between the cytoplasmic domains of the four subunits.

Crystal Structure of GIRK channel. Left - Front view of four GIRK2 subunits (color coded) channel assembly. Right - Top view of the cytoplasmic domain forming the selectivity filter.
Crystal Structure of GIRK channel. Left – Front view of the channel comprised of 4 subunits (color coded). Right – Top view of the cytoplasmic domain forming the selectivity filter. Protein Data Base ID – 3SYQ and 2QKS (Whorton, M. R., Mckinnon, R., 2011)

I have been specifically involved in understanding how certain amino acid residues residing in the hydrophobic pockets of the subunits influence channel activation and function. I use multiple experimental methods to investigate the interaction between the N- and C- termini of the GIRK1 and GIRK4 channel subunits to analyse protein expression and domain association. Previous research (Sarac et al, 2005) has revealed that certain mutations in the amino acid residues of these two subunits alters channel function.

Understanding how the interaction between the different GIRK channel subunits influences the channel formation and activity is critical for the elucidation of certain cellular mechanisms involved in cell physiology as well as in various channelopathies. New studies also suggest that ethanol binds to a hydrophobic pocket in the channel and activates it. (Bodhinathan, K., Slesinger, P. A., 2013 and Aryal, P., et al, 2009) Ethanol activation of the channel can be utilised for developing selective therapeutics to treat alcohol-related disorders like alcohol addiction and abuse.

Selected Resources:

Giraffe and Evolution – Not just a long (neck) story

Feeding on acacia leaves
Feeding high up on acacia leaves

In the early 19th century, Lamarck proposed a theory of evolution by studying the behaviour of giraffes. He believed that giraffes evolved to have long necks as they began reaching for higher leaves on trees. He called this “change through use and disuse”. According to this theory, an organ or a character that is used more often becomes stronger and better. Therefore, over the course of history, giraffe’s neck got longer as it began stretching it a lot more than usual. Lamarck also proposed the “law of inheritance of acquired characteristics” according to which the improved characteristics of an organism are passed on from one generation to the next. These improved features persists and the disadvantageous features disappear.

The Lamarckian theory was eventually abandoned* as it could not explain the genetic basis for inheritance of acquired characteristics (traits obtained after birth due to environmental changes, accidents, use and disuse; these traits are not inheritable). Lamarckism also predicts that simpler life forms will disappear from the earth once organisms become more complex. While we see some organisms evolving into more complex systems with intricate functions, the simpler life forms like the single celled prokaryotic cells still exist to this day.

Darwin’s theory of evolution on the other hand, can account for the continued existence of the simpler life forms on earth. Darwin believed that complexity is a result of adaptation to the environment from one generation to the next. In the Origin of Species, Darwin proposed a theory of evolution driven by natural selection. According to this theory, there is variation seen amongst individuals. Certain environmental conditions favours certain variations and the species exhibiting these variations adaptsurvive. The unadapted species which do not exhibit the favoured variations do not survive and become extinct over generations of time.

Applying this to giraffes, the long neck species are considered dominant and have greater chances of survival during harsh drought conditions compared to the short neck species that have to rely on ground habitation for food. This of course is just one theory amongst many other. The long necks are also used to reach deep inside trees that other competing animals cannot reach, and is therefore more advantageous. One of the latest proposal is the theory of sexual selection. Male giraffes fight with other males by “necking” to compete for female partners. As it turns out, females prefer males with longer and stronger necks. Natural selection again, favours long neck males.

“Animal Autopsy”, a show on National Geography channel dug deep into giraffes – quite literally! – to explore the physiological and anatomical features of this intriguing mammal to unravel some of its evolutionary secrets.

During the autopsy, Richard Dawkins talks about one of the evolutionary disadvantages caused due to the long laryngeal nerve that starts off in the brain and ends in the larynx (which is in fact situated very close to the brain). This nerve runs all the way down the neck, loops around one of the arteries in the chest and returns to the larynx on top. Why does the nerve take such a long route when it can simply connect from the brain to the larynx directly without having to pass through the entire neck? Also, consider this – giraffes with a long necks must also bend much lower to drink water from the ground. The contracting of the muscles along with the tension in the elastic tissue in the neck utilises way more energy, and is considered to be another evolutionary flaw. Another interesting fact to chew upon is that due to its long neck, giraffes have to pump blood to the brain that is ~2.5 meters above the heart – against gravity – by using extremely high blood pressure. How does the long neck favour such distant positioning of the heart and the brain?

Biologists consider different perspectives to understand the evolutionary reasoning behind the advantages as well as the flaws caused due to the long necks of giraffes. As more pieces of this puzzles are put together, it is quite remarkable to think about the imperfections that are caused due to evolution. Imperfections that somehow adds up to the making of such a marvellous mammal on our planet.

*New research reveals the epigenetic basis for the inheritance of acquired characteristics. In the article “A Comeback for Lamarckian Evolution?”, Emily Singer of the Tufts University School of Medicine provides evidence for the largely abandoned Lamarckian theory of evolution. Read article on MIT tech review here.