• Introduction
• Dual Signalling Pathway: Receptor to PLC
• The Dual Signalling Pathway: IP3, DAG & Calcium
• Phospholipase C: The Production of IP3 & DAG
• PLC Isoforms/Isozymes
• Protein Kinase C
• PKC & Gene Activation

In this lecture we continue with another pathway involving G protein linked receptors: the dual signaling pathway (Berridge, 1987. Ann. Rev. Biochem. 56: 159). In this pathway, after receptor activation initiates the G protein cycle, the first effector enzyme leads to the production of two signaling molecules. One of these enters the cytoplasm to cause the release of calcium ions from the endoplasmic reticulum while the other remains in the membrane to activate a membrane-associated protein kinase. Originally called the dual signaling pathway, it is now often referred to as the inositol-lipid signaling pathway. Let's look at the details of this signaling pathway. But first, we'll need to know some common acronyms. It is important to note that there are variations on these acronyms that are used by different journals and textbooks. For example, IP3 is also written Ins 1,4,5 P3. In both cases the acronym often has the "3" as a subscript. Also you may notice that subscripts or superscripts are presented in some figure but not in the corresponding text. Not all web-browsers show subscripts or superscripts properly so we use them sparingly in the text. The key is to know the basic acronyms and define them the first time you use them in any work.

We're going to look at the following figure in two parts. First we'll summarize the initial sequence events that are fundamentally the same as those that occur in all types of GPLR signaling. If these points aren't clear, then review the detailed events of the G protein cycle covered in the previous lecture.

• Receptor binds ligand
• Ligand/Receptor Complex undergoes conformational change
• G Protein binds receptor
• G protein dissociates; G binds to PLC
• PLC is activated

Next we'll look at the unique aspects of the signaling pathway in detail.

Listed are some widely used acronyms that are used in the figure below.

• PLC = Phospholipase C; note the beta isoform/isoenzyme is shown in the figure
• PIP2 = Phosphatidylinositol 1,4 bisphosphate
• IP3 = Inositol 1,4,5 Trisphosphate
• DAG = Diacylglycerol
• PKC = Protein Kinase C (C = Kinase); note the gamma isoform/isoenzyme is shown
• CaBP = Calcium Binding Protein
• CaM = Calmodulin

• PIP2 is cleaved by PLC
• Generates IP3 + DAG
• IP3 diffuses into cytosol
• IP3 will bind to IP3 receptor on the endoplasmic reticulum to release Ca2+ through receptor channel
• Ca2+ binds CaM & other CaBPs
• DAG diffuses in membrane
• DAG activate certain types of PKC which phosphorylate certain proteins
• Calcium ions also activate certain types of PKC

The following animation shows the full Dual Signalling Pathway

The following diagram shows the biochemical reaction that generates IP3 and DAG from PIP2. If you think back to the structure of the cell membrane, this information will be more meaningful.

• Isozymes = Different forms of the same enzyme
• Isoforms = Molecular variants of same protein

When you studied the dual signaling pathway, you should have noticed that phospholipase C was labelled PLC . This is because this was the beta isoform of the PLC protein. Since the protein is also an enzyme PLC is one isozyme in the PLC family. In the lectures on cancer, another isoform, PLC , will be introduced. We'll be discussing isoforms and isozymes for many other proteins as we progress through this course.

• Large family of related Ser/Thr Kinases (Nishizuka, 1992. Science 258: 607-614).
• At least 11 mammalian PKC isozymes with varied distributions
• Divided into three major groups (Dekker & Parker, 1994. Trends Biochem. Sci. 19: 73-77):
• Conventional PKCs (α, βI, βII, γ) activated by Ca2+, phospholipids and diacylglycerol (DAG).
• Novel PKCs (δ, &epsilon, υ and η)and activated by phospholipids and DAG; insensitive to Ca2+
• Atypical PKCs (ζ, ι, λ and μ) do not require Ca2+ or DAG for activation
• typically localized to the cytosol but several translocate to cell membrane when activated
• Phosphorylate a large number of substrates (e.g., MARCKS)
• Implicated in many critical processes (e.g., learning & memory, cell division & cancer)
• PKC phosphorylates proteins leading to changes in physiology (e.g., glucose mobilization) or gene activity (e.g., phosphorylation of transcription factors)

PKC lies upstream of signaling pathways that directly lead to gene activation.

• PKC Initiates chain of phosphorylations in MAPK/ ERK Pathway
• MAPK = Mitogen Activated Protein Kinase
• ERK = Extracellular Signal Related Kinase
• All are Ser/Thr Kinases
• Leads to Activation of Genes
• Other pathways can do same events: cross-talk (e.g., Ras)