remains unknown.
An example of the role of cyclic nucleotide–gated ion channels in sea urchin sperm chemotaxis.
Discovery
The discovery of CNG channels is related to the discovery of
cilia of olfactory sensory neurons, and the pineal gland. After the identification of amino acids from purified proteins, cloning and functional expression of CNG channels were performed. Molecular cloning allowed for the discovery of similar channels in many other tissues.[2][3] In 2000, scientists performed studies using mouse retina and molecular cloning to find a new subunit of the channel, CNG6.[4]
Function
CNG channels have important functions in
olfactory receptor neurons. They are directly activated by cyclic nucleotides, and approximately 4 cyclic nucleotides are needed to activate each channel. CNG channels are nonselective and allow many alkali ions to flow into or out of a cell expressing CNG channels on its membrane. This flow of ions can result in either depolarization or hyperpolarization. CNG channels can be activated by cAMP or cGMP exclusively, or sometimes by a combination of both cNMPs, and some channels are more selective than others. Even though the activity of these channels show little voltage dependence, they are still considered voltage-dependent channels. Calcium, calmodulin, and phosphorylation modulate the opening of CNG channels.[3]
The structure of the pore is similar to other ion channels that contain P-loops. The P-loop enters the
inner membrane line the channel. These also form a 6 helix bundle that signifies the entrance. In order to open the pore, a conformational change must occur in the inner 6 helix bundle.[5]
Cyclic nucleotide binding domain
A
disulfide bonds. This occurs mainly in closed channels, inhibiting movement of the α-helix towards the β-pleated sheet. When a ligand binds to the β-pleated sheet, this bound cyclic nucleotide stabilizes the movement of the α-helix toward the β-pleated sheet in each subunit, pulling the α-helices away from each other.[5]
[6]Illustration of a cyclic nucleotide–gated ion channel with a cAMP binding domain.
C-linker
The C-linker is a region that connects the CNBD to the S6 segment. The C-linker region contributes to the contact between channel subunits as well as promotes tetramerization, the forming of
N-terminal of the C-linker region that can reach two C481 residues, making a favorable disulfide bond compared to a C481-C481 bond.[5]
cations are able to move through an opening, which implies that the gate is beyond the helix bundle and that S6 helices are in conjunction with conformational changes in the selectivity filter.[6]
P region
The P region forms a loop, the pore loop, connecting the S5 and S6 regions, which extend to the central axis of the channel. Ionic properties are determined by the residues in the loop between S5 and S6
transmembrane segments. The P region dictates the ion selectivity of the cyclic-nucleotide gated ion channel, which also determine the pore diameter of CNG channels. The P region functions as a channel gate since it prevents ion permeation in the closed state. The pore may be hindered by small conformational changes in this region. The P region acts as an ion selectivity filter that changes structure in the open conformation. In the open state, four identical subunits contribute a single P-loop region, which forms a selectivity filter.[6]
CNG channel family
In vertebrates, the CNG channel gene family consists of six members and belong to a larger group of voltage-gated ion channels. These genes are divided based on sequence similarity into two subtypes CNGA and CNGB.
autosomal recessive form of retinitis pigmentosa, a degenerative form of blindness. CNGB1, previously called the rod β subunit, is a second subunit of the rod channel. Unlike CNGA1, CNGB1 subunits expressed alone do not produce functional CNG channels, but coexpression of CNGA1 and CNGB1 subunits produces heteromeric channels with modulation, permeation, pharmacology, and cyclic-nucleotide specificity comparable to that of native channels.[8]
CNG channels form
congenital disorder characterized by the complete failure in color distinction.[8]
CNGA4, previously called the olfactory β subunit, and CnGB1b are involved in transduction of odorant signals in olfactory sensory neurons for which the subunit stoichiometry and arrangement are unknown.[8]
In invertebrates, a CNG channel subunit called CNG-P1 has been cloned from D. melanogaster and is expressed in antennae and the visual system, an indication that CNG channels may be linked to the transduction of light in invertebrates. A second putative CNG-like subunit called CNGL, cloned from D. melanogaster, is found to be expressed in the brain. Two CNG channel subunits, Tax-2 and Tax-4, have been cloned in C. elegans and are responsible for chemosensation, thermosensation, and normal axon outgrowth of some sensory neurons in C. elegans.[8]
The binding event
The ligand might be placed at the bottom of the cavity due to interactions with the phosphate binding cassette (PBC). This cavity refers to a region in the CNBD formed by the β roll, a two-looped β helix. Changes induced by ligand binding occur in α helices (αA, αB, and αC and PBC helix). The β roll only undergoes small changes during binding. After the ligand is seated, αB and αC helices arrange themselves so that they form a cap over the cavity. How binding affects the αA helix is still unclear.
[9]
Cooperative and non-cooperative activation
The steep concentration between CNG channels and ligand concentration shows that at least two or three cyclic nucleotides are needed. It is believed that the second ligand is required for the channel to transition from closed to open. When the third and fourth ligands bind, the open state of the channel becomes stabilized.[9]
In bacteria, the opening of CNG channels is the result of non-cooperative binding.[9]
With differing concentrations of ligands, cooperative binding and non-cooperative binding arise to adapt to these differing environments. At low ligand concentrations, it is rare for a ligand to cooperatively bind, because cooperative binding at low concentrations weakens the binding between channel and ligand, reducing channel sensitivity.[9]
Ligand selectivity
By measuring the currents activated in excised inside-out membrane patches upon superfusion with varying
heteromeric channels most likely form a tetrameric
complex, a maximum of four ligand molecules can bind to the channel.
Selectivity can be achieved by differential control of the
affinity for binding of the ligand, efficacy of gating, or a combination of both. Binding affinity means how tightly cyclic nucleotides bind to the channel. Efficacy refers to the ability of ligand to activate and open the channel once it is bound. Although these processes are useful in understanding selectivity, they are inextricably coupled to each other that it is very difficult to experimentally separate one from another.[3]
CNG channels do not discriminate between
divalent ions inhibit the current carried by Na+ and K+. A highly conserved residue of glutamic acid in the selectivity filter of CNG channels has been found to form a high-affinity binding site for Ca2+. Moreover, a bacterial nonselective cation channel called the NaK channel hosts a selectivity filter sequence similar to that of CNG channels. In the crystal structure of the NaK channel, a discrete Ca2+-binding site at the extracellular opening of the pore has been identified.[9]
Inhibition of CNG channels
Studies have shown the differential inhibition of CNG channels by
glutamate.[3][5] Studies have shown that over activation of cGMP-dependent CNG channels in photoreceptors can lead to their degeneration. If the CNG channels on a photoreceptor are continuously activated, Ca2+ and Na+ ion flux into the outer segment of the photoreceptor will increase so that it depolarizes beyond the dark current. Through a positive feedback loop, this would then increase the current of Ca2+ into the cell. High concentration of Ca2+ in the photoreceptor cell would lead to its death programmed cell death or apoptosis.[11]
Fundus of patient with retinitis pigmentosa, mid stage (Bone spicule-shaped pigment deposits are present in the mid periphery along with retinal atrophy, while the macula is preserved although with a peripheral ring of depigmentation. Retinal vessels are attenuated.)
Retinitis Pigmentosa (RP) is a genetic disease in which patients suffer degeneration of rod and cone photoreceptors. The loss starts in the patient's peripheral vision and progresses to the central visual field, leaving the patient blind by middle age.
About 1% of RP patients have mutations in
phototransduction cascades. The mutation of these subunits indirectly impairs rod cGMP-gated channel function, which implies that there is a common mechanism of photoreceptor degradation.[12]
In the nervous system, heart, and some visceral organs, cells contain cyclic nucleotide gated channels which determine the rhythm of the organ. These channels, formally called hyperpolarization-activated cyclic nucleotide–gated channels (
funny current), triggering another depolarization event and subsequent cardiac contraction. This gives the heart its automaticity. The primary cyclic nucleotide operating in conjunction with the HCN channel is cAMP.[13]
Olfactory sensory neurons
Almost all responses to odorants in
cilia membrane, it activates a G protein, which causes a downstream reaction activating the enzyme adenylyl cyclase (AC). This enzyme is responsible for an increase in cAMP concentration within the OSN. cAMP binds to the CNG channels in the OSN membrane, opening them, and making the cell highly permeable to Ca2+. Calcium ions flow into the cell causing a depolarization. As in all other cell types, CNG channels in OSNs also allow Na+ to flow into the cell. Additionally, the increased Ca2+ concentration inside the cell activates Ca2+-dependent chloride (Cl−) channels, which causes intracellular Cl− ions to also flow out of the cell augmenting the depolarization event. This depolarization stimulates an action potential that ultimately signals the reception of the odorant. In addition to cAMP gated ion channels, a small subset of OSNs also has cGMP-selective CNG channels that contain the CNGA3 subunit.[3]
. CNG channels are prime candidates for the calcium-entry pathway, due to their high calcium permeability. CNG channels have yet to be detected by homology screening.
In
CatSper need additional subunits to become functional, they are unrelated to CNG channels because CatSper lacks a cAMP/cGMP-binding site. It is possible that CNG and CatSper subunits assemble to form calcium-permeable and cyclic nucleotide-sensitive ion channels.[3]
Kidney
cGMP-sensitive channels have been analyzed in the
arterioles.[14] Differences between retinal and renal cDNA have been implicated in the functional differences between CNG channels in these two tissues.[3]
Gonadotropin-releasing hormone
There has been identification of CNG ion channel subunits A2, A4, and B1 in a neuronal cell line that secretes
cilia of OSNs. In high extracellular calcium, the unit conductance of CNG channels in rods and OSNs are significantly smaller than those measured in the neuronal line. It seems doubtful that CNG channels would create large unit conductance.[3]
CryoEM Structure of a prokaryotic cyclic nucleotide-gated ion channel.
Plants
CNG ions channels in plants are similar in
immunity and response to pathogens or external infectious agents. They have also been implicated in apoptosis in plants. CNG ion channels are also thought to be involved in pollen development in plants, however its exact role in this mechanism is still not known.[15]
Unlike animal CNG channels, plant CNG channels have not been extensively analyzed biochemically with respect to their structure.[15]
Prokaryotes
CNG ion channels share a high degree of sequence and structural similarity to mammalian CNG channels.
cyclic nucleotides to the CNBDs has been shown to regulate channel activity and alter the channel conformational state.[16][17] Because these channels were only recently identified in spirochaeta and leptospira species,[16] their precise physiological function remains unknown in these organisms. In combination with photoactivatedadenylyl cyclases, they have been used as optogenetic tools to inhibit action potential generation in neurons.[18]
Current and future research
Researchers have answered many important questions regarding CNG ion channels functions in vision and
olfaction. In other physiological areas, the role of CNG channels is less defined. With technological growth, there now exists more possibilities for understanding these mechanisms.[3]
Because nitric oxide (NO) is involved in stimulating the synthesis of cGMP, further research is being conducted to understand the physiological interaction of NO with CNG channels, particularly in the covalent modification of CNG channels in OSNs.[3]
Scientists are adding on to the mechanism involved in the interaction of binding sites and interfaces of subunits. This might be nonexistent in non-cooperative CNG channels. It is also possible that binding site and gate are attached to a single subunit. In order to develop these ideas, double electron-electron resonance (DEER) and rapid fixing techniques can show these mechanistic movements.[9]
A 2007 study suggests that because of the various and complex regulatory properties in addition to the large number of CNG channels in plants, a multidisciplinary study to research plant CNG channels should be conducted.[15] Another study in March 2011 recognizes recent reverse genetics data that has been helpful in further understanding CNG channels in plants, and also suggests that additional research be conducted to identify the upstream and downstream factors in CNGC-mediated signal transduction in plants.[19]
Scientist are speculating whether DAG directly binds with CNG channel during inhibition. It is possible that DAG may insert itself into the transmembrane domains in the channel. It is also possible that DAG inserts itself into the interface between the channel and bilayer. The molecular mechanism of DAG inhibition is still not fully understood.[10]
