Specific regulatory proteins control both types of GTP-binding proteins. GTPase-activating proteins (GAPs) drive the proteins into an “off” state by increasing the rate of hydrolysis of bound GTP. Conversely, guanine nucleotide exchange factors (GEFs) activate GTP-binding proteins by promoting the release of bound GDP, which allows a new GTP to bind. In the case of trimeric G proteins, the activated receptor serves as the GEF. Figure 15–8 illustrates the regulation of monomeric GTPases.
Not all molecular switches in signaling systems depend on phosphorylation or GTP binding. We see later that some signaling proteins are switched on or off by the binding of another signaling protein or a second messenger such as cyclic AMP or Ca2+, or by covalent modifications other than phosphorylation or dephosphorylation, such as ubiquitylation (discussed in Chapter 3).
For simplicity, we often portray a signaling pathway as a series of activation steps (see Figure 15–1). It is important to note, however, that most signaling pathways contain inhibitory steps, and a sequence of two inhibitory steps can have the same effect as one activating step (Figure 15–9). This double-negative activation is very common in signaling systems, as we will see when we describe specific pathways later in this chapter.
Ideally, an activated intracellular signaling molecule should interact only with the appropriate downstream targets, and, likewise, the targets should only be activated by the appropriate upstream signal. In reality, however, intracellular
Figure 15–7 Two types of intracellular signaling proteins that act as molecular switches. (A) A protein kinase covalently adds a phosphate from ATP to the signaling protein, and a protein phosphatase removes the phosphate. Although not shown, many signaling proteins are activated by dephosphorylation rather than by phosphorylation. (B) A GTP-binding protein is induced to exchange its bound GDP for GTP, which activates the protein; the protein then inactivates itself by hydrolyzing its bound GTP to GDP.
signaling molecules share the cytoplasm with a crowd of closely related signaling molecules that control a diverse array of cellular processes. It is inevitable that an occasional signaling molecule will bind or modify the wrong partner, potentially creating unwanted cross-talk and interference between signaling systems. How does a signal remain strong, precise, and specific under these noisy conditions?
The first line of defense comes from the high affinity and specificity of the interactions between signaling molecules and their correct partners compared to the relatively low affinity of the interactions between inappropriate partners. The binding of a signaling molecule to the correct target is determined by precise and complex interactions between complementary surfaces on the two molecules. Protein kinases, for example, contain active sites that recognize a specific amino acid sequence around the phosphorylation site on the correct target protein, and they often contain additional docking sites that promote a specific, high-affinity interaction with the target. These and related mechanisms help provide a strong and persistent interaction between the correct partners, reducing the likelihood of inappropriate interactions with other proteins.
Another important way that cells avoid responses to unwanted background signals depends on the ability of many downstream target proteins to simply ignore such signals. These proteins respond only when the upstream signal reaches a high concentration or activity level. Consider a signaling pathway in which a protein kinase activates some downstream target protein by phosphorylation. If a response is triggered only when more than half of the target proteins are phosphorylated, then there will be little harm done if a small number of them are occasionally phosphorylated by some inappropriate protein kinase. Furthermore, constitutively active protein phosphatases will further reduce the impact of background phosphorylation by rapidly removing much of it. In these and other ways, intracellular signaling systems filter out noise, generating little or no response to low levels of stimuli.