Growth factors, cytokines (growth factors of the immune system), and some hormones are signaling molecules that use catalytic or enzymatic receptors to stimulate their target cells (see also LIR Immunology, Chapter 6). Most catalytic receptors are single-chain transmembrane proteins that associate with other single-chain transmembrane proteins upon ligand binding and signal via phosphorylation of tyrosine (Tyr) residues. The Tyr kinase activity responsible for the production of phosphotyrosines may belong to the receptor itself or to a Tyr kinase that associates with the receptor.
As a consequence of ligand binding to the receptor, intracellular portions of the receptor protein become phosphorylated on Tyr residues, then in a regulated manner, protein Tyr phosphatases rapidly dephosphorylate phosphotyrosines. The short-lived appearance of phosphotyrosines is a potent signal to the cell. Phosphorylated Tyr residues within the receptor induce binding of other proteins that serve as adaptors in the relay of the signal deeper into the cell. The original signal from the ligand may be split along several intracellular pathways as the signal is sent further onward with the goal of evoking a biological response to the ligand.
RECEPTORS WITH INTRINSIC TYROSINE KINASE ACTIVITY
Many growth factors signal via receptors with intrinsic Tyr kinase activity. Examples include transforming growth factor, epidermal growth factor, and platelet-derived growth factor. Insulin, a hormone, also signals via intrinsic catalytic receptors. Receptors with intrinsic Tyr kinase activity contain latent catalytic or enzymatic domains that are activated upon ligand binding. While many distinct catalytic receptors exist, they all share some common structural characteristics.
Most catalytic receptors are formed by associations of two or more single transmembrane protein chains. Each of the single transmembrane chains has three domains: a ligand-binding portion that contains the amino- (NH2) terminus of the protein, an ? helical domain that spans the lipid bilayer, and an effector region that extends into the cytoplasm that contains the catalytic domain with Tyr kinase activity (Figure 18.1).
FIGURE 18.1. Structure of catalytic receptors.
Mechanism of signaling
In response to ligand binding, the individual transmembrane protein chains are brought physically closer together, often forming dimers (Figure 18.2). Tyr kinase domains of each receptor chain activate the other and the Tyr kinase within each receptor tail phosphorylates Tyr residues on intracellular portions of the other receptor chain. Tyr residues within the receptor itself serve as substrates for the receptor’s Tyr kinase. This process is known as autophosphorylation since Tyr residues within the receptor protein are phosphorylated by enzymatic activity within the receptor itself.
As a consequence, phosphotyrosine residues are present on each of the receptor cytoplasmic tails. Tyr phosphorylation triggers assembly of an elaborate intracellular signaling complex on the receptor tails (cytoplasmic domains). Intracellular proteins, known as adaptor proteins, which contain highly conserved domains (regions) known as SH2 and SH3 domains (named for their Src homology) dock with phosphotyrosines of the cytoplasmic tails of the receptor chains (Figure 18.3). Different receptors recruit distinct collections of SH2-containing adaptor proteins.
FIGURE 18.2. Initial steps in signaling by catalytic receptors with intrinsic tyrosine kinase activity.
FIGURE 18.3. Binding of adaptor proteins.
Important adaptor molecules
Adaptor molecules can function in signaling initiated by different growth factors or hormones. Certain adaptor molecules are known to be critically important in multiple signaling processes.
1. Ras: Ras is a small GTP-binding protein and serves as a molecular switch for key signaling in the control of growth and differentiation (see also Chapter 17, Figure 17.9). Ras does not contain an SH2 domain itself but it binds to SH2-containing adaptor proteins that bind to phosphorylated Tyr residues within receptor tails. Ras activates a serine/threonine phosphorylation cascade called the MAP kinase cascade. Serine and threonine phosphorylations are longer lived than Tyr phosphorylations. The final enzyme in this cascade, mitogen-activated protein kinase (MAPK), becomes phosphorylated, translocates to the nucleus, and phosphorylates transcription factors. Phosphorylated transcription factors induce transcription of genes that allow the cell to proliferate or differentiate, depending on the nature of the signaling molecule at the cell’s surface.
2. STATs: STATs are named for their function as signal transducers and activators of transcription. They are SH2-containing latent, cytoplasmic proteins that can bind to phosphorylated Tyr residues within the cytoplasmic domains of receptors with their own catalytic activity and are also involved in signaling by nonreceptor Tyr kinases. After ligand binding has induced receptor dimerization and Tyr phosphorylation of receptor tails, STATs dock with a phosphotyrosine on a receptor tail (Figure 18.4). When the STAT is bound to a phosphorylated Tyr residue, the Tyr kinase sees the STAT as a substrate and phosphorylates Tyr residues within the STAT protein. Tyr-phosphorylated STATs form dimers and translocate to the nucleus to bind to DNA and induce transcription of certain responsive genes.
FIGURE 18.4. STATs in signal transduction.
PI3 kinase pathway
Another major signaling pathway stimulated by catalytic receptors is the phosphatidylinositol 3-kinase or PI3 kinase pathway that is important in promoting cell survival and cell growth. Following ligand binding to a catalytic receptor, receptor dimerization, and phosphorylation of Tyr residues within the cytoplasmic tail of the receptor, PI3 kinase binds to the phosphorylated Tyr residues (Figure 18.5). Activated PI3 kinase phosphorylates membrane inositol phospholipids (on their three position), such as phosphatidylinositol 4,5-bisphosphate (PIP2).
PIP2 is converted to PIP3 in response to PI3 kinase action. Phosphorylated inositol lipids are docking sites for intracellular signaling proteins. Akt, also called protein kinase B, is recruited by PIP3 and activated by phosphorylation. Bad is then phosphorylated by Akt, inactivating Bad and preventing it from inducing programmed cell death (apoptosis). Thus, cell survival is promoted. PIP3 remains in the membrane until it is dephosphorylated by inositol phospholipid phosphatases, including PTEN, whose actions halt the signaling mechanism. (If a mutation occurs in PTEN, signaling through PI3 kinase can continue for prolonged periods and can promote cancer development.)
FIGURE 18.5. PI3 kinase pathway.
SIGNALING VIA NONRECEPTOR TYROSINE KINASES
Receptors for cytokines (interleukins and interferons) and for some hormones (such as prolactin and growth hormone) do not possess their own Tyr kinase activity but activate nonreceptor Tyr kinases to carry out their signaling process (see also LIR Immunology, pp. 66–73). The cytoplasmic domains of these receptors noncovalently associate with cytoplasmic Tyr kinase proteins and phosphorylate Tyr residues within the receptor tail. Several nonreceptor Tyr kinases have been identified but two of the best characterized include the Src and Janus kinase families.
Src family tyrosine kinases
Src was the first nonreceptor Tyr kinase discovered. There are at least eight members of this nonreceptor protein Tyr kinase family including Blk, Fgr, Fyn, Hck, Lck, Lyn, Src, and Yes. All of these contain SH2 and SH3 domains that mediate protein to protein interactions as well as an SH1 catalytic domain (Figure 18.6). Different members of the family are found in different cell types. For example, Fyn, Lck, and Lyn function in lymphocyte signal transduction.
Various members of the Src family can phosphorylate Tyr residues of many of the same target proteins. Src family members are regulated by phosphorylation on Tyr residues and by protein to protein interactions. Src proteins are normally inactive and are switched on only at crucial times. If Src remains active, uncontrolled growth and malignancy can result. Mutations in Src are found in many cancers.
FIGURE 18.6. Src family of tyrosine kinases.
Janus kinases, referred to as JAKs, are latent cytosolic Tyr kinases activated by certain cytokine and hormone receptors. JAKs phosphorylate Tyr residues of the intracellular portion of the receptor chains. STATs bind to these phosphotyrosines and are phosphorylated on Tyr residues by the JAK (Figure 18.7). This signaling process is often referred to as the JAK-STAT pathway. Tyr-phosphorylated STATs form dimers and translocate to the nucleus as previously described.
FIGURE 18.7. Janus kinase signaling.
Insulin signals via catalytic receptors with intrinsic Tyr kinase activity. The insulin receptor is preformed in the membrane with all chains joined together prior to hormone binding. After insulin binds to the extracellular ligand-binding domain of its receptor, the insulin receptor Tyr kinase activity is stimulated, inducing Tyr phosphorylation of various insulin receptor substrates (IRS) (Figure 18.8). At least four different IRS proteins are known. IRS-1 and IRS-2 are widely expressed, IRS-3 is found in adipose tissue, pancreatic ? cells, and possibly liver, while IRS-4 is found in thymus, brain, and kidney. Depending on the tissue type and the IRS proteins expressed, insulin will induce different biological responses to its signaling.
FIGURE 18.8. Insulin receptor autophosphorylation and IRS function.
Recent research has shown that Tyr-phosphorylated IRS proteins activate several different intracellular signaling proteins including Ras, STATs, and PI3 kinase. This signal splitting occurs as the message sent to the cell by insulin is diverted down several pathways in order to evoke the biological response in the cell. Activation of Ras and STAT signaling leads to the regulation of transcription of specific genes. It is thought that the activation of the PI3 kinase and its downstream kinases promotes glucose transport (see also Chapter 15, Figure 15.6), protein synthesis, glycogen synthesis, cell proliferation, and cell survival in various cells and tissues.
- Most catalytic receptors are single-chain transmembrane proteins that form dimers when the ligand binds.
- Stimulation of phosphorylation of substrates on Tyr residues is a key feature of signaling by catalytic receptors.
- Some receptors have intrinsic Tyr kinase activity while others associate with nonreceptor Tyr kinases.
- Receptors for growth factors and for the hormone insulin contain intrinsic Tyr kinase activity that is stimulated by ligand binding.
- Adapter molecules containing SH2 domains bind to the phosphorylated Tyr residues within the receptor cytoplasmic tails when the catalytic receptor is activated by its ligand. STATs are such adapter proteins that are activated to stimulate gene transcription.
- The PI3 kinase pathway promotes cell growth and survival and is stimulated by many catalytic receptors. Phosphorylation of inositol phospholipids stimulates additional signaling reactions.
- Src and Janus kinases are intracellular nonreceptor Tyr kinases.
- Insulin signals its catalytic receptor to undergo autophosphorylation on Tyr residues. The activated Tyr kinase domain of the receptor then phosphorylates various IRSs to send the signal onward in the cell.