IP3 and DAG Signaling Pathway

The IP3/DAG signaling pathway is a crucial mechanism within cells that helps regulate various biological processes. It involves the activation of inositol trisphosphate (IP3) and diacylglycerol (DAG), which act as secondary messengers to transmit signals within the cell. This pathway plays a fundamental role in processes such as cell growth, metabolism, and intracellular communication. In this article, we will study about IP3/DAG pathway, its mechanism, functioning, and steps involved in the IP3 DAG pathway.

What is IP3/DAG Pathway?

The IP3 DAG pathway is a vital process in cell function. It starts when extracellular ligands attach to receptors on the cell’s surface, triggering a series of events inside the cell. This leads to the creation of two important messengers: inositol trisphosphate (IP3) and diacylglycerol (DAG).

IP3 diffuses through the cytoplasm to the endoplasmic reticulum, releasing calcium ions, which control many cell activities. Meanwhile, DAG stays near the cell’s surface, activating protein kinase C (PKC), which starts other signals affecting cell growth, division, and metabolism.

Mechanism of IP3 Signaling

These processes are followed from ligands to downstream targets. Thyroid-stimulating hormone and acetylcholine bind to and activate either heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors (GCPRs) or tyrosine kinase receptors (rTKs). When a receptor is activated, phospholipase C (PLC) is activated, which converts phosphatidylinositol 4,5-bisphosphate (PIP2) to IP3 and diacylglycerol (DAG).

The release of calcium from the endoplasmic reticulum is then stimulated by IP3, and calcium regulates the activity of multiple downstream targets. Protein kinase C is one of the downstream targets (PKC). Calcium helps PKC to bind to DAG and hence be activated by it. Following that, PKC phosphorylates downstream substrates such as glycogen synthase and the calmodulin-binding protein neurogranin.

IP3 (Inositol Triphosphate)

An inositol phosphate signaling molecule is also known as inositol 1,4,5-triphosphate and abbreviated InsP3, Ins3P, or IP3. It is created by phospholipase carbon breakdown of the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) (PLC).

IP3 is a second messenger molecule utilized in signal transduction in biological cells together with diacylglycerol (DAG). IP3 is soluble and diffuses into the cell, where it interacts with its receptor, a calcium channel found in the endoplasmic reticulum, whereas DAG remains inside the membrane. Calcium is released into the cytosol when IP3 binds to its receptor, activating a variety of intracellular calcium-regulated processes.

Acetylcholine and thyroid-stimulating hormone are ligands that bind to and activate either heterotrimeric G protein-coupled receptors (GCPRs) or tyrosine kinase receptors (rTKs). Phospholipase C (PLC) is activated as a result of receptor activation, cleaving phosphatidylinositol 4,5-bisphosphate (PIP2) into IP3 and diacylglycerol (DAG).

The release of calcium from the endoplasmic reticulum is then stimulated by IP3, and calcium regulates the activity of many downstream targets. Protein kinase C is one of the downstream targets (PKC). PKC can bind to DAG and then be activated by it thanks to calcium. Following that, downstream substrates like glycogen synthase and the calmodulin-binding protein neurogranin are phosphorylated by PKC.

Extracellular main messengers such as adrenaline, acetylcholine, and hormones AGT, GnRH, GHRH, oxytocin, and TRH attach to their specific receptors to initiate the circuit.

DAG (Diacylglycerol)

Using ester bonds, two fatty acid chains are covalently joined to a glycerol molecule to form a diglyceride, also known as diacylglycerol (DAG). There are two varieties that could exist: 1,2 and 1,3 diacylglycerols. DAGs are frequently utilized as emulsifiers in processed foods and can function as surfactants. Due to its potential to prevent the buildup of body fat, DAG-enriched oil, in particular 1,3-DAG, has received substantial research as a fat alternative, with total yearly sales in Japan reaching over USD 200 million from its launch in the late 1990s to 2009.

In order to transmit signals downstream of the many receptors expressed by hematopoietic cells, diacylglycerol (DAG) is a crucial secondary lipid messenger. Adaptive and innate immune cells’ activation, proliferation, migration, and effector capabilities have been demonstrated to be significantly influenced by DAG.

Bioactive lipids like diacylglycerol and phosphatidic acid are created when the T cell receptor interacts with a cognate peptide-MHC complex. Ras guanyl-releasing protein 1, PKC, and other effectors are recruited by DAG to initiate signaling, whereas PA binds to effector molecules such as the mechanistic target of rapamycin, Src homology region 2 domain-containing phosphatase 1, and Raf1.

While it has been demonstrated that DAG-mediated pathways are crucial for T cell growth and operation, the significance of PA-mediated signals is still unclear. The family of enzymes known as diacylglycerol kinases (DGK) phosphorylates DAG to create PA, acting as a molecular switch to control the relative levels of these vital second messengers.

IP3/DAG Signaling Pathway

1. In many cases, IP3 activation leads to increases in intracellular Ca²⁺ concentrations. The Gq heterotrimeric G protein’s -subunit can bind to and activate the PLC isozyme PLC-, leading to the cleavage of PIP2 into IP3 and DAG when a ligand binds to a G protein-coupled receptor (GPCR) that is connected to Gq. 
2. If a receptor tyrosine kinase (RTK) is required for pathway activation, the isozyme PLC- possesses tyrosine residues that can be phosphorylated upon activation of an RTK. This will activate PLC- and permit it to cleave PIP2 into DAG and IP3 if an RTK is involved in pathway activation. Due to the fact that growth factors are the ligands that activate the RTK, this happens in cells that can respond to growth factors like insulin.
3. After being created by PLC, IP3 (also known as Ins(1,4,5)P3) is a soluble molecule that can diffuse through the cytoplasm to the ER or the sarcoplasmic reticulum (SR) in the case of muscle cells. 
4. Once inside the ER, IP3 can attach to the ligand-gated Ca²⁺ channel on the ER’s surface via the IIns(1,4,5)P3 receptor, or Ins(1,4,5)P3R. When IP3 (the ligand in this instance) binds to Ins(1,4,5) P3R, the Ca²⁺ channel opens, releasing Ca²⁺ into the cytoplasm. 
5. This rise in Ca²⁺ causes calcium-induced calcium release, which causes additional Ca²⁺ increases in cardiac muscle cells by activating the ryanodine receptor-operated channel on the SR. IP3 may indirectly open Ca²⁺ channels on cell membranes by raising intracellular Ca2+ concentration.

Pathway for IP3

The other secondary messenger produced by PIP2 cleavage, IP3, is a tiny polar molecule that is released into the cytosol and functions to signal the release of Ca²⁺ from intracellular storage, whereas diacylglycerol stays connected to the plasma membrane. Ca²⁺ pumps that actively export Ca²⁺ from the cell keep the cytosolic concentration of Ca²⁺ at a very low level (approximately 0.1 M).

Ca²⁺ is pushed into the endoplasmic reticulum as well as across the plasma membrane, acting as an intracellular Ca²⁺ store as a result. IP3 binds to ligand-gated Ca²⁺ channel receptors, which release Ca²⁺ from the endoplasmic reticulum. The result is an increase in cytosolic Ca²⁺ concentrations to about 1 M, which has an impact on the actions of numerous target proteins, including protein kinases and phosphatases. For instance, some members of the protein kinase C family need both Ca²⁺ and diacylglycerol to activate, so both arms of the PIP2 signaling pathway work together to control these protein kinases.

Pathway for DAG

Protein-serine/threonine kinases from the protein kinase C family, many of which play crucial roles in the regulation of cell development and differentiation, are activated by the diacylglycerol created by the hydrolysis of PIP2. The activity of phorbol esters, which have been the subject of in-depth research because they aid in the development of tumors in animals, serves as an excellent example of this function of protein kinase C.

The phorbol esters’ capacity to stimulate protein kinase C by functioning as diacylglycerol analogs underlies their tumor-promoting effect. Then, protein kinase C activates additional intracellular targets, such as the MAP kinase pathway, a chain of protein kinases, which results in the activation of transcription factors, modifications to gene expression, and promotion of cell proliferation. In addition to triggering PKC, diacylglycerol performs a variety of other tasks in cells, including:

• Prostaglandins source
• The endocannabinoid 2-arachidonoylglycerol’s precursor
• A TRPC3/6/7 cation channel activator, a member of the TRPC (Transient Receptor Potential Canonical) cation channel family.

Functions of IP3 Signaling

1. Human: In humans, IP3 primarily controls free calcium-dependent cellular processes such as cell proliferation and the release of Ca²⁺ from storage organelles. For instance, a rise in the cytoplasmic Ca²⁺ concentration causes the contraction of the muscle cell in smooth muscle cells. The cerebellum has the highest concentration of IP3 receptors in the neurological system, where IP3 functions as a second messenger. Evidence suggests that IP3 receptors are crucial for promoting plasticity in cerebellar Purkinje cells.
2. Sea urchin eggs: The PIP2 secondary messenger system mediates the gradual impediment to polyspermy in sea urchins. The binding receptors activate PLC, which then breaks down PIP2 in the egg plasma membrane and releases IP3 into the cytoplasm of the egg cell. In the ER, IP3 diffuses and activates Ca²⁺ channels.
3. Activating protein kinase C: Phospholipase C (PLC), a membrane-bound enzyme, hydrolyzes the phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) to create inositol trisphosphate, which serves as a second messenger in biochemical signaling. Phospholipase C is a membrane-bound enzyme (IP3). Diacylglycerol stays inside the plasma membrane due to its hydrophobic characteristics, whereas inositol trisphosphate diffuses into the cytosol. While DAG is a physiological activator of protein kinase C, IP3 increases the release of calcium ions from the smooth endoplasmic reticulum (PKC). The membrane’s synthesis of DAG makes it easier for PKC to go from the cytosol to the plasma membrane.
4. Activation of Munc13: Diacylglycerol has been found to interact with the presynaptic priming protein family Munc13 to exert part of its excitatory effects on vesicle release. DAG binding to Munc13’s C1 domain improves synaptic vesicle fusion capability, leading to potentiated release. The tumor-promoting substances phorbol esters can resemble diacylglycerol.

Translocation of PK-C

RACK proteins help protein kinase C enzymes go to the plasma membrane after activation (membrane-bound receptor for activated protein kinase C proteins). The Ca²⁺ wave or the initial activation signal is no longer present, yet the protein kinase C enzymes continue to be active. This is most likely accomplished by a phospholipase converting phosphatidylcholine into diacylglycerol; fatty acids may also contribute to long-term activation.

They could attach to various RACK proteins and perform varied RACK functions depending on their isoenzyme forms. The activity of PK-C may be inhibited by inhibiting the RACK binding domain of the protein. Bronchoconstriction, latelet aggregation, CSF secretion, H⁺ secretion, Na⁺ reabsorption, and are all aided by Protein Kinase-C activity.

The CaM kinases

The Ca2+/calmodulin complex is principally responsible for controlling serine/threonine-specific protein kinases, also known as CaM kinases. The activation of these kinases exhibits a memory effect. CaM kinase comes in two varieties:

1. Particular CaM kinases: The myosin light chain kinase (MLCK), which phosphorylates myosin to cause muscles to contract, is one example
2. Multipurpose CaM kinases: They are sometimes referred to as CaM kinase II collectively, and they have a variety of functions, including the regulation of transcription factors, glycogen metabolism, and the release of neurotransmitters. CaM kinase II makes up 1% to 2% of the proteins in the brain.

Regulation of IP3 Signaling

• P38-MAPK signaling pathway: The p38-MAPK signaling system can be activated in adipocytes to increase intracellular calcium transport, control adipocyte metabolism, and lessen obesity.
• Calmodulin: Calmodulin influences the energy metabolism of adipocytes to decrease obesity. In hypothalamic neurons, CaMKK2 activation can control food behavior to lower obesity. Following Ca²⁺ binding, calmodulin is only partially activated; phosphorylation results in full activation. It binds to a short peptide after being activated, which causes modifications in its own structure and boosts its activity. Additionally, it modifies the protein’s structure to activate it.
• IP3-Ca²⁺ signaling pathway: In order to control lipolysis and the buildup of adipose, the IP3 pathway can be activated, increasing the intracellular calcium ions concentration in adipocytes.
• Other calcium signaling pathways: Calcium ions can be released by Rya receptor channels in the ER/SR to control neuronal excitability, which in turn controls energy metabolism. Voltage-dependent calcium channels that are open allow extracellular calcium ions to enter cells and control PKA’s ability to affect obesity. Through the activation of PKC and CaMK2, the Wnt-Ca2+ signaling pathway can decrease obesity by raising intracellular calcium concentration.

Conclusion – IP3/DAG Pathway

The IP3/DAG signaling pathway emerges as a critical mechanism controlling diverse cellular functions. Through the activation of inositol trisphosphate (IP3) and diacylglycerol (DAG), this pathway regulates processes such as cell growth, metabolism, and intercellular communication. From the initial binding of extracellular ligands to the subsequent activation of downstream targets, each step in the IP3 DAG pathway contributes to the dynamic coordination of cellular responses. 

FAQs on IP3 Signaling

Why is IP3 Involved in Cell Signaling?

The primary jobs of IP3 are to release Ca²⁺ from storage organelles and to control cell division and other cellular processes that need free calcium.

What are the Roles of DAG and IP3?

DAG and IP3 both serve as significant second messengers. The protein kinase C is attracted to and activated by DAG while it is still in the membrane. The endoplasmic reticulum and other intracellular organelles that store Ca²⁺ are among those where IP3 stimulates the opening of IP3-mediated Ca²⁺ channels.

How is IP3 Regulated?

We propose that cytosolic Ca(2+), which binds to two different locations, biphasically regulates the activity of all IP(3) receptors. By deciding whether Ca(2+) binds to the stimulatory or inhibitory sites, IP(3) promotes channel opening.

How is DAG Regulated?

The diacylglycerol kinases and phosphatidic acid phosphatases are responsible for closely regulating the intracellular levels of DAG and PA due to their significance. As a result, these enzymes play a variety of crucial physiological tasks.

What Kind of Receptors does IP3 Attach to?

IP3 is produced by phospholipase C (PLC) when receptor tyrosine kinases (RTKs) are stimulated. IP3 is then produced and binds to IP3Rs in the endoplasmic reticulum.

Which Hormone Acts Through IP3 DAG?

Acetylcholine, Epinephrine, Vasopressin , and Oxytocin are the hormones that acts through IP3 DAG Pathway.

What Function does Diacylglycerol DAG Serve?

Diacylglycerol (DAG) serves as a crucial activator of protein kinase C (PKC), regulating diverse cellular processes including growth, proliferation, and metabolism. Additionally, DAG plays a role in membrane trafficking and lipid signaling.