Biochemistry, 4th Edition P105 pot

31.4 How Does Protein Degradation Regulate Cellular Levels of Specific Proteins? 1003 The pK a of Lys 526 is lowered in this complex, activating the lysine amino group for nucleophilic attack to form the SUMO–target protein conjugate. HtrA Proteases Also Function in Protein Quality Control The discussion thus far has stressed the importance of protein quality control to cel- lular health. HtrA proteases are a class of proteins involved in quality control that combine the dual functions of chaperones and proteasomes. (The acronym Htr comes from “high temperature requirement” because E. coli strains bearing muta- tions in HtrA genes do not grow at elevated temperatures.) In addition to their novel ability to be either chaperones or proteases, HtrA proteases are the only known pro- tein quality control factor that is not ATP-dependent. Prokaryotic HtrA proteases act as chaperones at low temperatures (20°C) where they have negligible protease ac- tivity. However, as the temperature increases, these proteins switch from a chaperone function to a protease function to remove misfolded or unfolded proteins from the cell. With this functional duality, HtrA proteases have the potential to mediate qual- ity control through protein triage (see A Deeper Look box on page 1004). The E. coli HtrA protein DegP is the best characterized HtrA protease. DegP is lo- calized in the E. coli periplasmic space, where it oversees quality control of proteins in- (b)(a) A A B D E F B C C N N C C 1 2 3 4 D Leu 525 Lys 526 Ser 527 Glu 528 Lys 74 Thr 81 Asp 127 Cys 93 Glu 88 Ser 89 FIGURE 31.14 (a) The complex formed by the E2 enzyme, Ubc9 (yellow), and a target protein, RanGAP1 (blue). (b) In the Ubc9–RanGAP1 complex, the exposed loop of RanGAP1 lies in the binding pocket of Ubc9.The exposed loop contains the consensus sequence for sumoylation (ψKXD/E), including Leu 525 , which is sur- rounded by hydrophobic residues from Ubc9; Lys 526 , which is coordinated by Asp 127 and Cys 93 of Ubc9; and Glu 528 , which is coordinated by Ubc9 Thr 81 . HUMAN BIOCHEMISTRY Proteasome Inhibitors in Cancer Chemotherapy Proteasome inhibition offers a promising approach to treating cancer. The counterintuitive rationale goes like this: The protea- some is responsible for the regulated destruction of proteins in- volved in cell cycle progression and the control of apoptosis (pro- grammed cell death). Inhibition of proteasome function leads to cell cycle arrest and apoptosis. In clinical trials, proteasome in- hibitors have retarded cancer progression by interfering with the programmed degradation of regulatory proteins, causing cancer cells to self-destruct. Bortezomib, a small-molecule proteasome in- hibitor developed by Millenium Pharmaceuticals, Inc., has re- ceived FDA approval for the treatment of multiple myeloma, a cancer of plasma cells that accounts for 10% of all cancers of the blood (see Figure 31.10a). Source: Adams, J., 2003. The proteasome: Structure, function, and role in the cell. Cancer Treatment Reviews Supplement 1:3–9. 1004 Chapter 31 Completing the Protein Life Cycle: Folding, Processing, and Degradation volved in the cell envelope. It is a 448-residue protein containing a central protease domain with a classic Ser protease Asp-His-Ser catalytic triad (see Chapter 14) and two C-terminal PDZ domains. These domains are structural modules involved in protein– protein interactions, and they recognize and bind selectively to the C-terminal three or four residues of target proteins. Like other quality control systems, HtrA proteases have a central cavity where proteolysis occurs (Figure 31.15); the height of this cavity is 1.5 nm, which excludes folded proteins from access to the proteolytic sites. Thus, HtrA proteases can act only on misfolded proteins. As we have seen, limited access to proteolytic sites is an important regulatory feature of quality control proteases; only (a) 90° (b) PDZ PDZ FIGURE 31.15 The HtrA protease structure. (a) A trimer of DegP subunits represents the HtrA functional unit.The different domains are color- coded:The protease domain is green,PDZ do- main 1 (PDZ1) is yellow, and PDZ domain 2 (PDZ2) is orange. Protease active sites are high- lighted in blue.The trimer has somewhat of a funnel shape, with the protease in the center and the PDZ domains on the rim. (b) Two HtrA trimers come together to form a hexameric structure in which the two protease domains form a rigid molecular cage (blue) and the six PDZ domains are like tentacles (red) that both bind protein substrate targets and control lateral access into the protease cavity. (Adapted from Fig- ure 3 in Clausen,T., Southan, C., and Ehrmann, M., 2002. The HtrA family of proteases: Implications for protein composition and cell fate. Molecular Cell 10:443–455.) A DEEPER LOOK Protein Triage—A Model for Quality Control Triage is a medical term for the sorting of patients according to their need for (and their likelihood to benefit from) medical treat- ment. Sue Wickner, Michael Maurizi, and Susan Gottesman have pointed out that cells control the quality of their proteins through a system of triage based on the chaperones and the ubiquitination- proteasome degradation pathway. These systems recognize non- native proteins (proteins that are only partially folded, misfolded, incorrectly modified, damaged, or in an inappropriate compart- ment). Depending on the severity of its damage, a non-native pro- tein is directed to chaperones for refolding or targeted for de- struction by a proteasome (see accompanying figure). Adapted from Figures 2 and 3 in Wickner, S., Maurizi, M., and Gottesman, S., 1999. Posttranslational quality control: Folding, refolding, and degrad- ing proteins. Science 286:1888–1893. ATP Unfolded and misfolded proteins Ubiquitination system Ubiquitin and Ubiquitinated protein Eukaryotic chaperones Bound protein Protein remodeling Native protein Protein bound to proteasome Degraded p rotein ATP ATP Problems 1005 proteins targeted for destruction have access to such sites. The PDZ domains of the HtrA proteases act as gatekeepers, determining access of protein substrates to the pro- teolytic centers. Human HtrA proteases are implicated in stress response pathways. Human HtrA1 is expressed at higher levels in osteoarthritis and aging and lower lev- els in ovarian cancer and melanoma. Secreted human Htr1 may be involved in degra- dation of extracellular matrix proteins involved in arthritis as well as tumor progres- sion and invasion. SUMMARY 31.1 How Do Newly Synthesized Proteins Fold? Most proteins fold spontaneously, as anticipated by Anfinsen, whose experiments suggested that all of the information necessary for a polypeptide chain to assume its active, folded conformation resides in its amino acid sequence. How- ever, some proteins depend on molecular chaperones to achieve their folded conformation within the crowded intracellular environment. Hsp70 chaperones are ATP-dependent proteins that bind to exposed hydrophobic regions of polypeptides, preventing nonproductive associa- tions with other proteins and keeping the protein in an unfolded state until productive folding steps can take place. Hsp60 chaperones such as GroEL–GroES are ATP-dependent cylindrical chaperonins that provide a protected central cavity or “Anfinsen cage,” where partially folded pro- teins can fold spontaneously, free from the danger of nonspecific hy- drophobic interactions with other unfolded protein chains. Hsp90 chap- erones assist a subset of “client proteins” involved in signal transduction pathways in assuming their active conformations. 31.2 How Are Proteins Processed Following Translation? Nascent pro- teins are seldom produced in their final, functional form. Maturation typically involves proteolytic cleavage and may require post-translational modification, such as phosphorylation, glycosylation, or other covalent substitutions. Removal of nascent N-terminal methionine residues is a common form of protein processing by proteolysis. The number of hu- man proteins is believed to exceed the number of human genes (20,000 or so) by an order of magnitude or more. The great number of proteins available from a fixed set of genes is attributed to a variety of processes, including alternative splicing of mRNAs and post-translational modifica- tion of proteins. 31.3 How Do Proteins Find Their Proper Place in the Cell? Most pro- teins destined for compartments other than the cytosol are synthesized with N-terminal signal sequences that target them to their proper desti- nations. These signal sequences are recognized by signal recognition particles as they emerge from translating ribosomes. The signal recog- nition particle interacts with a membrane-bound signal receptor, deliv- ering the translating ribosome to a translocon, a multimeric integral membrane protein structure having at its core the Sec61p complex. The translocon transfers the growing protein chain across the membrane (or in the case of nascent integral membrane proteins, inserts the pro- tein into the membrane). Signal peptidases within the membrane com- partment clip off the signal sequence. Other post-translational modifi- cations may follow. Mitochondrial protein import and membrane insertion are mediated by specific translocon complexes in the outer mitochondrial membrane called TOM and SAM. Proteins destined for the inner mitochondrial membrane or mitochondrial matrix must in- teract with inner mitochondrial translocons (either TIM23 or TIM22) as well as TOM. 31.4 How Does Protein Degradation Regulate Cellular Levels of Specific Proteins? Protein degradation is potentially hazardous to cells, because cell function depends on active proteins. Therefore, protein degradation is compartmentalized in lysosomes or in proteasomes. Proteins targeted for destruction are selected by ubiquitination. A set of three enzymes (E 1 , E 2 , and E 3 ) mediate transfer of ubiquitin to free ONH 2 groups in tar- geted proteins. Proteins with charged or hydrophobic residues at their N-termini are particularly susceptible to ubiquitination and destruction. The ubiquitin moieties are recognized by 19S cap structures found at either end of 26S proteasomes. Protein degradation occurs when, in an ATP-dependent process, the ubiquitinated protein is unfolded and threaded into the central cavity of the cylindrical ␣ 7 ␤ 7 ␤ 7 ␣ 7 20S part of the 26S proteasome. The ␤-subunits possess protease active sites that chop the protein substrate into short (seven- to nine-residue) fragments; the ubiquitin moieties are recycled. Linkage of SUMO (small ubiquitin-like protein modifiers) to target proteins has the ability to alter their protein- protein interactions. HtrA proteases also function in protein quality con- trol. HtrA proteins are novel in two aspects: Unlike other chaperones and proteasomes, they are ATP-independent; and unlike the others, they have dual chaperone and protease activities and switch between these two functions in response to stress conditions. PROBLEMS Preparing for an exam? Create your own study path for this chapter at www.cengage.com/login. 1. (Integrates with Chapter 30.) Human rhodanese (33 kD) consists of 296 amino acid residues. Approximately how many ATP equivalents are consumed in the synthesis of the rhodanese polypeptide chain from its constituent amino acids and the folding of this chain into an active tertiary structure? 2. A single proteolytic break in a polypeptide chain of a native protein is often sufficient to initiate its total degradation. What does this fact suggest to you regarding the structural consequences of proteo- lytic nicks in proteins? 3. Protein molecules, like all molecules, can be characterized in terms of general properties such as size, shape, charge, solubility/hydropho- bicity. Consider the influence of each of these general features on the likelihood of whether folding of a particular protein will require chap- erone assistance or not. Be specific regarding just Hsp70 chaperones or Hsp70 chaperones and Hsp60 chaperonins. 4. Many multidomain proteins apparently do not require chaperones to attain the fully folded conformations. Suggest a rational scenario for chaperone-independent folding of such proteins. 5. The GroEL ring has a 5-nm central cavity. Calculate the maximum molecular weight for a spherical protein that can just fit in this cav- ity, assuming the density of the protein is 1.25 g/mL. 6. (Integrates with Chapter 24.) Acetyl-CoA carboxylase has at least seven possible phosphorylation sites (residues 23, 25, 29, 76, 77, 95, and 1200) in its 2345-residue polypeptide (see Figure 24.4). How many different covalently modified forms of acetyl-CoA car- boxylase protein are possible if there are seven phosphorylation sites? 1006 Chapter 31 Completing the Protein Life Cycle: Folding, Processing, and Degradation 7. (Integrates with Chapter 30.) In what ways are the mechanisms of action of EF-Tu/EF-Ts and DnaK/GrpE similar? What mechanistic functions do the ribosome A-site and DnaJ have in common? 8. The amino acid sequence deduced from the nucleotide sequence of a newly discovered human gene begins: MRSLLILVLCFLPLAALGK… . Is this a signal sequence? If so, where does the signal peptidase act on it? What can you surmise about the intended destination of this protein? 9. Not only is the Sec61p translocon complex essential for transloca- tion of proteins into the ER lumen, it also mediates the incorpora- tion of integral membrane proteins into the ER membrane. The mechanism for integration is triggered by stop-transfer signals that cause a pause in translocation. Figure 31.5 shows the translocon as a closed cylinder spanning the membrane. Suggest a mechanism for lateral transfer of an integral membrane protein from the protein-conducting channel of the translocon into the hydrophobic phase of the ER membrane. 10. The Sec61p core complex of the translocon has a highly dynamic pore whose internal diameter varies from 0.6 to 6 nm. In post- translational translocation, folded proteins can move across the ER membrane through this pore. What is the molecular weight of a spherical protein that would just fit through a 6-nm pore? (Adopt the same assumptions used in problem 5.) 11. (Integrates with Chapters 6, 9, and 30.) During co-translational translocation, the peptide tunnel running from the peptidyl trans- ferase center of the large ribosomal subunit and the protein- conducting channel are aligned. If the tunnel through the ribosomal subunit is 10 nm and the translocon channel has the same length as the thickness of a phospholipid bilayer, what is the minimum number of amino acid residues sequestered in this common conduit? 12. Draw the structure of the isopeptide bond formed between Gly 76 of one ubiquitin molecule and Lys 48 of another ubiquitin molecule. 13. Assign the 20 amino acids to either of two groups based on their sus- ceptibility to ubiquitin ligation by E 3 ubiquitin protein ligase. Can you discern any common attributes among the amino acids in the less susceptible versus the more susceptible group? 14. Lactacystin is a Streptomyces natural product that acts as an irre- versible inhibitor of 26S proteasome ␤-subunit catalytic activity by covalent attachment to N-terminal threonine OOH groups. Predict the effects of lactacystin on cell cycle progression. 15. HtrA proteases are dual-function chaperone-protease protein qual- ity control systems. The protease activity of HtrA proteases depends on a proper spatial relationship between the Asp-His-Ser catalytic triad. Propose a mechanism for the temperature-induced switch of HtrA proteases from chaperone function to protease function. 16. As described in this chapter, the most common post-translational modifications of proteins are proteolysis, phosphorylation, methy- lation, acetylation, and linkage with ubiquitin and SUMO pro- teins. Carry out a Web search to identify at least eight other post- translational modifications and the amino acid residues involved in these modifications. 17. Fluorescence resonance energy transfer (FRET) is a spectroscopic technique that can be used to provide certain details of the confor- mation of biomolecules. Look up FRET on the Web or in an intro- ductory text on FRET uses in biochemistry, and explain how FRET could be used to observe conformational changes in proteins bound to chaperonins such as GroEL. A good article on FRET in protein folding and dynamics can be found here: Haas, E., 2005. The study of protein folding and dynamics by determination of in- tramolecular distance distributions and their fluctuations using en- semble and single-molecule FRET measurements. ChemPhysChem 6:858–870. Studies of GroEL using FRET analysis include the fol- lowing: Sharma, S., et al., 2008. Monitoring protein conformation along the pathway of chaperonin-assisted folding. Cell 133:142–153; and Lin, Z., et al., 2008. GroEL stimulates protein folding through forced unfolding. Nature Structural and Molecular Biology 15:303–311. 18. The cross-talk between phosphorylation and ubiquitination in pro- tein degradation processes is encapsulated in the concept of the “phosphodegron.” What is a phosphode gron, and how does phos- phorylation serve as a recognition signal for protein degradation? (A good reference on the phosphodegron and crosstalk between phosphorylation and ubiquitination is Hunter, T., 2007. The age of crosstalk: Phosphorylation, ubiquitination, and beyond. Molecular Cell 28:730–738.) Preparing for the MCAT Exam 19. A common post-translational modification is removal of the univer- sal N-terminal methionine in many proteins by Met-aminopeptidase. How might Met removal affect the half-life of the protein? 20. Figure 31.6 shows the generalized amphipathic ␣-helix structure found as an N-terminal presequence on a nuclear-encoded mito- chondrial protein. Write out a 20-residue-long amino acid sequence that would give rise to such an amphipathic ␣-helical secondary structure. FURTHER READING Protein-Folding Diseases Bates, G., 2003. Huntingdin aggregation and toxicity in Huntington’s disease. Lancet 361:1642–1644. Bucciantini, M., Giannoni, E., et al., 2002. Inherent toxicity of aggre- gates implies a common mechanism for protein misfolding. Nature 416:507–511. Gamblin, T. C., Chen, F., et al., 2003. Caspase cleavage of tau: Linking amyloid and neurofibrillary tangles in Alzheimer’s disease. Proceed- ings of the National Academy of Sciences U.S.A. 100:10032–10037. Herczenik, E., and Gebbink, M. F., 2008. Molecular and cellular aspects of protein misfolding and disease. FASEB Journal 22:2115–2133. Soto, C., Estrada, L., et al., 2006. Amyloids, prions, and the inherent in- fectious nature of misfolded protein aggregates. Trends in Biochemi- cal Sciences 31:150–156. Winklhofer, K. F., Tatzelt, J., et al., 2008. The two faces of protein mis- folding: Gain- and loss-of-function in neurodegenerative diseases. EMBO Journal 27:336–349. Chaperone-Assisted Protein Folding Bigotti, M. G., and Clarke, A. R., 2008. Chaperonins: The hunt for the group II mechanism. Archives of Biochemistry and Biophysics 474: 331–339. Bukau, B., et al., 2000. Getting newly synthesized proteins into shape. Cell 101:119–122. Bukau, B., Weissman, J., et al., 2006. Molecular chaperones and protein quality control. Cell 125:443–451. Ellis, J. R., 2001. Molecular chaperones: Inside and outside the Anfinsen cage. Current Biology 11:R1038–R1040. Frydman, J., 2001. Folding of newly translated proteins in vivo: The role of molecular chaperones. Annual Review of Biochemistry 70:603–647. Hartl, F. U., and Hayer-Hartl, H., 2002. Molecular chaperones in the cytosol: From nascent chain to folded chain. Science 295:1852–1858. Jahn, T., and Radford, S. E., 2007. Folding versus aggregation: Polypep- tide conformations on competing pathways. Archives of Biochemistry and Biophysics 469:100–117. Further Reading 1007 Kramer, G., et al., 2002. L23 protein functions as a chaperone docking site on the ribosome. Nature 419:171–174. Lin, Z., Madan, D., et al., 2008. GroEL stimulates protein folding through forced unfolding. Nature Structural and Molecular Biology 15:303–311. Luheshi, L. M., Crowther, D. C., et al., 2008. Protein misfolding and dis- ease: From the test tube to the organism. Current Opinion in Chemi- cal Biology 12:25–31. Sharma, S., Chakraborty, K., et al., 2008. Monitoring protein conformation along the pathway of chaperonin-assisted folding. Cell 133:142–153. Vogel, M., Bukau, B., et al., 2006. Allosteric regulation of Hsp70 chap- erones by a proline switch. Molecular Cell 21:359–367. Protein Translocation Bolender, N., Sickmann, A., et al., 2008. Multiple pathways for sorting mitochondrial precursor proteins. EMBO Reports 9:42–49. Bornemann, T., Jockel, J., et al., 2008. Signal sequence-independent membrane targeting of ribosomes containing short nascent pep- tides within the exit tunnel. Nature Structural and Molecular Biology 15:494–499. Brodersen, D. E., and Nissen, P., 2005. The social life of ribosomal pro- teins. FEBS Journal 272:2098–2108. Kutik, S., Guiard, B., et al., 2007. Cooperation of translocase complexes in mitochondrial protein import. Journal of Cell Biology 179:585–591. Mihara, K., 2003. Moving inside membranes. Nature 424:505–506. Rapoport, T. A., 2007. Protein translocation across the eukaryotic en- doplasmic reticulum and bacterial plasma membranes. Nature 450:663–669. Schnell, D. J., and Hebert, D. N., 2003. Protein translocons: Multi- functional mediators of protein translocation across membranes. Cell 112:491–505. Schwartz, S., and Blobel, G., 2003. Structural basis for the function of the ␤-subunit of the eukaryotic signal recognition particle. Cell 112: 793–803. Skach, W., 2007. The expanding role of the ER translocon in membrane protein folding. Journal of Cell Biology 179:1333–1335. Wirth, A., et al., 2003. The Sec61p complex is a dynamic precursor acti- vated channel. Molecular Cell 12:261–268. Ubiquitin Selection of Proteins for Degradation Bellare, P., Small, E. C., et al., 2008. A role for ubiquitin in the spliceosome assembly pathway. Nature Structural and Molecular Biology 15:444–451. Hershko, A., 1996. Lessons from the discovery of ubiquitin system. Trends in Biochemical Sciences 21:445–449. Hochstrasser, M., 1996. Ubiquitin-dependent protein degradation. An- nual Review of Genetics 30:405–439. Hunter, T., 2007. The age of crosstalk: Phosphorylation, ubiquitination, and beyond. Molecular Cell 28:730–738. Madsen, L., Schulze, A., et al., 2007. Ubiquitin domain proteins in dis- ease. BMC Biochemistry 8: 1–8. V arshvsky, A., 1997. The ubiquitin system. Trends in Biochemical Sciences 22:383–387. Proteasome-Mediated Protein Degradation Borissenko, L., and Groll, M., 2007. 20S proteasome and its inhibitors: Crystallographic knowledge for drug development. Chemical Reviews 107:687–717. Breusing, N., and Grune, T., 2008. Regulation of proteasome-mediated protein degradation during oxidative stress and aging. Biological Chemistry 389:203–209. Dahlmann, B., 2007. Role of proteasomes in disease. BMC Biochemistry 8:1–12. Ferrel, K., et al., 2000. Regulatory subunit interactions of the 26S pro- teasome, a complex problem. Trends in Biochemical Sciences 25:83–88. Groll, M., Berkers, C. R., et al., 2006. Crystal structure of the boronic acid-based proteasome inhibitor bortezomib in complex with the yeast 20S proteasome. Structure 14:451–456. Zhang, F., Paterson, A. J., et al., 2007. Metabolic control of proteasome function. Physiology 22:373–379. HtrA Proteases Clausen, T., Southan, C., et al., 2002. The HtrA family of proteases: Im- plications for protein composition and cell fate. Molecular Cell 10:443–455. Kim, D. Y., and Kim, K. K., 2005. Structure and function of HtrA family proteins, the key players in protein quality control. Journal of Bio- chemistry and Molecular Biology 38:266–274. Sohn, J., Grant, R. A., et al., 2007. Allosteric activation of DegS, a stress sensor PDZ protease. Cell 131:572–583. Walle, L. V., Lamkanfi, M., et al., 2008. The mitochondrial serine pro- tease HtrA2/Omi: An overview. Cell Death and Differentiation 15: 453–460. Post-translational Modification by Sumoylation Anckar, J., and Sistonen, L., 2007. SUMO: Getting it on. Biochemical So- ciety Transactions 35:1409–1413. Bernier-Villamor, V., Sampson, D. A., et al., 2002. Structural basis for E2-mediated SUMO conjugation revealed by a complex between ubiquitin-conjugating enzyme Ubc9 and RanGAP1. Cell 108:345–356. Geiss-Friedlander, R., and Melchior, F., 2007. Concepts in sumoylation: A decade on. Nature Reviews Molecular Cell Biology 8:947–956. Hay, R. T., 2005. SUMO: A history of modification. Molecular Cell 18:1–12. Johnson, E. S., 2004. Protein modification by SUMO. Annual Review of Biochemistry 73:355–382. Ulrich, H. D., 2005. Mutual interactions between the SUMO and ubiq- uitin systems: A plea of no contest. Trends in Cell Biology 15:525–532. Vertegaal, A. C. O., 2007. Small ubiquitin-related modifiers in chains. Biochemical Society Transactions 35:1422–1423. Yunus, A. A., and Lima, D. D., 2006. Lysine activation and functional analysis of E2-mediated conjugation in the SUMO pathway. Nature Structural and Molecular Biology 13:491–499. Florentine Royal Collection, Windsor, England A.K.G., Berlin/Superstock, International 32 The Reception and Transmission of Extracellular Information Hormones are secreted by certain cells, usually located in glands, and travel, either by simple diffusion or circulation in the bloodstream, to specific target cells (Fig- ure 32.1). As we shall see, some hormones bind to specialized receptors on the plasma membrane and induce responses within the cell without themselves enter- ing the target cell (Figure 32.2). Other hormones actually enter the target cell and interact with specific receptors there. By these mechanisms, hormones: •Regulate the metabolic processes of various organs and tissues • Facilitate and control growth, differentiation, reproductive activities, learning, and memory • Help the organism cope with changing conditions and stresses in its environment All these effects of hormonal signals are either metabolic responses or gene expression responses (Figure 32.2). 32.1 What Are Hormones? Many different chemical species act as hormones. Steroid hormones, all derived from cholesterol, regulate metabolism, salt and water balances, inflammatory processes, and sexual function. Several hormones are amino acid derivatives. Among these are epinephrine and norepinephrine (which regulate smooth muscle contraction and relax- ation, blood pressure, cardiac rate, and the processes of lipolysis and glycogenolysis) and the thyroid hormones (which stimulate metabolism). Peptide hormones are a large group of hormones that regulate processes in all body tissues, including the release of yet other hormones. Hormones and other signal molecules in biological systems bind with very high affinities to their receptors, displaying K D values in the range of 10 Ϫ12 to 10 Ϫ6 M. The hormones are produced at concentrations equivalent to or slightly above these K D values. Once hormonal effects have been induced, the hormone is usually rapidly metabolized. Steroid Hormones Act in Two Ways The steroid hormones include the glucocorticoids (cortisol and corticosterone), the mineralocorticoids (aldosterone), and the sex hormones (progesterone and testos- terone, for example) (Figure 32.1; see Chapter 24 for the details of their synthesis). The steroid hormones exert their effects in two ways: First, by entering cells and mi- Drawing of a human fetus in utero, by Leonardo da Vinci. Human sexuality and embryonic development represent two hormonally regulated processes of uni- versal interest. “How little we know, how much to discover, What chemical forces flow from lover to lover. How little we understand what touches off that tingle, That sudden explosion when two tingles intermingle. Who cares to define what chemistry this is? Who cares with your lips on mine How ignorant bliss is, So long as you kiss me and the world around us shatters? How little it matters how little we know” “How Little We Know” by P. Springer and C. Leigh (Excerpt from “How Little We Know” by P. Springer and C. Leigh as recorded by Frank Sinatra, April 5, 1956. Capitol Records, Inc.) KEY QUESTIONS 32.1 What Are Hormones? 32.2 What Is Signal Transduction? 32.3 How Do Signal-Transducing Receptors Respond to the Hormonal Message? 32.4 How Are Receptor Signals Transduced? 32.5 How Do Effectors Convert the Signals to Actions in the Cell? 32.6 How Are Signaling Pathways Organized and Integrated? 32.7 How Do Neurotransmission Pathways Control the Function of Sensory Systems? ESSENTIAL QUESTION Higher life forms must have molecular mechanisms for detecting environmental infor- mation as well as mechanisms that allow for communication at the cell and tissue lev- els. Sensory systems detect and integrate physical and chemical information from the environment and pass this information along by the process of neurotransmission. Control and coordination of processes at the cell and tissue levels are achieved not only by neurotransmission but also by chemical signals in the form of hormones that are secreted by one set of cells to direct the activity of other cells. What are these mechanisms of information transfer that mediate the molecular basis of hormone action and that use excitable membranes to transduce the signals of neurotransmission and sensory systems? Create your own study path for this chapter with tutorials, simulations, animations, and Active Figures at www.cengage.com/login. 32.1 What Are Hormones? 1009 grating to the nucleus, steroid hormones act as transcription regulators, modulating gene expression. These effects of the steroid hormones occur on time scales of hours and involve synthesis of new proteins. Steroids can also act at the cell membrane, di- rectly regulating ligand-gated ion channels and perhaps other processes. These latter processes take place very rapidly, on time scales of seconds and minutes. Somatostatin (anterior pituitary) Testosterone Estradiol (many: vascular system, reproductive organs, central nervous system, gastrointestinal tract, immune system, skin, kidney, and lung) Bradykinin (blood vessels) Prolactin (breast) Glucagon (primarily liver) Gastrin (GI tract, gall bladder, pancreas) Luteinizing hormone (LH) (gonads, ovarian follicle cells) Follicle-stimulating hormone (FSH) (gonads) Adrenocorticotropic hormone (ACTH) (adrenal cortex) Insulin (primarily liver, muscle, fat) Growth hormone (GH) (many: bone, fat, liver) Chorionic gonadotropin (various reproductive tissues) Calcitonin (bone) FIGURE 32.1 The sites of synthesis and action of a few of the polypeptide and steroid hormones. Hormones typically circulate at low concentrations (1 nM or less) and bind with high affinity to their receptor proteins. Nonsteroid hormone Steroid hormone Gene expression Transcription responses Signal transduction pathways Nucleus Metabolic responses FIGURE 32.2 Nonsteroid hormones bind exclusively to plasma membrane receptors, which mediate the cellular responses to the hormone. Steroid hormones exert their effects either by binding to plasma membrane recep- tors or by diffusing to the nucleus, where they modulate transcriptional events. Intracellular responses to hor- mone binding include metabolic changes and alter- ations of gene expression. 1010 Chapter 32 The Reception and Transmission of Extracellular Information Polypeptide Hormones Share Similarities of Synthesis and Processing The largest class of hormones in vertebrate organisms is that of the polypeptide hormones (Figure 32.1). One of the first polypeptide hormones to be discovered, insulin, was described by Banting and Best in 1921. Insulin, a secretion of the pan- creas, controls glucose utilization and promotes the synthesis of proteins, fatty acids, and glycogen. Insulin, which is typical of the secreted polypeptide hormones, is dis- cussed in detail in Chapters 5, 15, and 22. Many other polypeptide hormones are produced and processed in a manner sim- ilar to that of insulin. Three unifying features of their synthesis and cellular process- ing should be noted. First, all secreted polypeptide hormones are originally synthe- sized with a signal sequence, which facilitates their eventual direction to secretory granules, and thence to the extracellular milieu. Second, peptide hormones are usu- ally synthesized from mRNA as inactive precursors, termed preprohormones, which become activated by proteolysis. Third, a single polypeptide precursor or prepro- hormone may produce several different peptide hormones by suitable proteolytic processing. An impressive example of the production of many hormone products from a single precursor is the case of prepro-opiomelanocortin, a 250-residue precursor peptide synthesized in the pituitary gland. A cascade of proteolytic steps produces, as the name implies, a natural opiate substance (endorphin) and several other hor- mones (Figure 32.3). Endorphins and other opiatelike hormones are produced by the body in response to systemic stress. These substances probably contribute to the “runner’s high” that marathon runners describe. 32.2 What Is Signal Transduction? Hormonal regulation depends on the transduction of the hormonal signal across the plasma membrane to specific intracellular sites, particularly the nucleus. Signal transduction consists of a stepwise progression of signaling stages: receptor⎯→ transducers⎯→effectors. The receptor perceives the signal, transducers relay the signal, and the effectors convert the signal into an intracellular response. What are the characteristics of signal transduction systems? Regardless of the organism or the tissue, certain features appear to be universal, or nearly so: • Transmembrane communication of hormonal signals by receptor proteins • Protein interaction domains that selectively recognize specific structural motifs and bind them with high affinity and specificity Signal peptide Signal peptidase 26 48 12 14 Corticotropin Anterior pituitary Nervous tissue ␤-Lipotropin ␤-MSH␣-MSH Endorphin 18 312140 40 Pro-opiomelanocortin Prepro-opiomelanocortin ␥-Lipotropin FIGURE 32.3 The conversion of prepro-opiomelanocortin to a family of peptide hormones, including corticotropin, ␤- and ␥-lipotropin, ␣- and ␤-MSH, and endorphin. 32.2 What Is Signal Transduction? 1011 • Clustering of membrane receptors and their ligands in large aggregates called signalsomes • Reversible covalent modifications that change the function of certain proteins and lipids (including phosphorylation, methylation, acetylation, ubiquitylation, hydroxylation, and cleavage) • Second messengers that bind to specific targets, changing their activity and be- havior • Intracellular signaling pathways, often involving a series of enzymes (such as pro- tein kinases), that link receptors to their downstream functional targets • Cooperativity • Spatial and temporal control of signals and messengers • Integration of signals • Converging and diverging networks •Signal amplification • Desensitization and adaptation Many Signaling Pathways Involve Enzyme Cascades Signaling pathways must operate with speed and precision, facilitating the accurate relay of intracellular signals to specific targets. But how does this happen? Enzyme cascades are one answer to this question. Enzymes can produce (or modify) a large number of molecules rapidly and specifically. Many enzymes of signaling cascades are protein kinases and protein phosphatases, and many steps in signaling pathways involve phosphorylation of serine, threonine, and tyrosine residues on target pro- teins. The complexity of signal transduction is thus manifested in the estimates that the human genome contains about a thousand protein kinase and protein phos- phatase genes. Enzyme cascades act like a series of amplifiers, dramatically increas- ing the magnitude of the intracellular response available from a very small amount of hormone. Signaling Pathways Connect Membrane Interactions with Events in the Nucleus The complete pathway from hormone binding at the plasma membrane to modula- tion of transcription in the nucleus is understood for a few signaling pathways. Figure 32.4 shows a complete signal transduction pathway that connects receptor tyrosine kinases, the Ras GTPase, cytoplasmic Raf, and two other protein kinases with tran- scription factors that alter gene expression in the nucleus. This pathway represents just one component of a complex signaling network that involves many other proteins and signaling factors. The existence of nearly 4000 human genes devoted to signal transduction portends a complex and interwoven network of signaling interactions in nearly all human cells. 1 Signaling Pathways Depend on Multiple Molecular Interactions Each step in a signaling pathway depends on one or more molecular interac- tions. For example, a protein may bind to a small molecule, a peptide, or even another protein (Figure 32.5). In fact, many signaling proteins are constructed in a cassettelike fashion with several distinct modules, termed protein interaction domains (PIDs), each of which mediates a particular interaction (Figure 32.6a). A protein with multiple modules can interact with several binding targets at once to create a complex, called a signalsome, with multiple signaling capabilities (Figure 32.6b). Several signaling systems described in this chapter involve events at a signalsome. 1 The American Association for the Advancement of Science oversees a consortium of researchers who have established a Web site—the Signal Transduction Knowledge Environment (STKE). This site is an up-to-date and ongoing compilation of information about cell signaling and signal transduction. The URL is http://www.stke.org. 1012 Chapter 32 The Reception and Transmission of Extracellular Information GTP GDP Active Raf Raf P MAPKK Active MAPKK 2 2 P Polypeptide hormone MAPK Thr Active MAPK P MAPK kinase Mitogen-activated protein kinase Tyrosine kinase Tyr Tyr Ras Ras GRB2 SOS Receptor tyrosine kinase(RTK) Nucleus Fos ATF-2 Jun Myc Gene transcription Protein synthesis Transcription factor phosphorylation Tyr P ATP ADP ATP ADP ACTIVE FIGURE 32.4 A complete signal transduction pathway that connects a hormone receptor with transcription events in the nucleus. A number of similar pathways have been characterized. Test yourself on the concepts in this figure at www.cengage.com/login. . 14 Corticotropin Anterior pituitary Nervous tissue ␤-Lipotropin ␤-MSH␣-MSH Endorphin 18 312140 40 Pro-opiomelanocortin Prepro-opiomelanocortin ␥-Lipotropin FIGURE 32.3 The conversion of prepro-opiomelanocortin. How Does Protein Degradation Regulate Cellular Levels of Specific Proteins? Protein degradation is potentially hazardous to cells, because cell function depends on active proteins. Therefore, protein. confor- mation of biomolecules. Look up FRET on the Web or in an intro- ductory text on FRET uses in biochemistry, and explain how FRET could be used to observe conformational changes in proteins bound