Biochemistry, 4th Edition P105 pot

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 o

The pKaof Lys526is 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

E

F

B C C

N

N

C

C

1 2 3 4

D

Leu525 Lys526 Ser527

Glu528

Lys74 Thr81

Asp127 Cys93

Glu88

Ser89

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 Leu525, which is

sur-rounded by hydrophobic residues from Ubc9; Lys526, which is coordinated by Asp127and Cys93of Ubc9; and

Glu528, which is coordinated by Ubc9 Thr81

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)

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 dodo-main 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 protein

ATP

ATP

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 invaprogres-sion.

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 huhu-man 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 (E1, E2, and E3) mediate transfer of ubiquitin to free ONH2groups 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 777720S 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-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

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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 chapchap-erones

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 Gly76of

one ubiquitin molecule and Lys48of another ubiquitin molecule

13. Assign the 20 amino acids to either of two groups based on their

sus-ceptibility to ubiquitin ligation by E3ubiquitin 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 activqual-ity 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, acetymethy-lation, 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 phosphodegron, 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.

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.

Varshvsky, 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

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 KDvalues in the range of 1012to 106M The hormones are produced at concentrations equivalent to or slightly above these KD 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?

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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 prepro-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 horhor-mones 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

Corticotropin

Anterior pituitary

Nervous tissue

-Lipotropin

-MSH

21

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.

• 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 transcripmodula-tion 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.

1The 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

Active Raf

Raf

P

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

Jun Myc

Gene transcription

Protein synthesis

Transcription factor phosphorylation

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

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