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agonists generally have other, undesirable effects. To minimize such side effects, the goal is to
find an agonist that is specific for the subtype of b-adrenergic receptors found in bronchial
smooth muscle.
6. Termination of Hormonal Signals Signals carried by hormones must eventually be terminated.
Describe several different mechanisms for signal termination.
Answer A hormone can be degraded by extracellular enzymes (such as acetyl-
cholinesterase). The GTP bound to a G protein can be hydrolyzed to GDP. A second messen-
ger can be degraded (cAMP, cGMP), further metabolized (IP3), or resequestered (Ca2�, in the
endoplasmic reticulum). A receptor can be desensitized (acetylcholine receptor/channel),
phosphorylated/inactivated, bound to an arrestin, or removed from the surface (b-adrenergic
receptor, rhodopsin).
7. Using FRET to Explore Protein-Protein Interactions in Vivo Figure 12–8 shows the interaction
between �-arrestin and the �-adrenergic receptor. How would you use FRET (see Box 12–3) to
demonstrate this interaction in living cells? Which proteins would you fuse? Which wavelengths would
you use to illuminate the cells, and which would you monitor? What would you expect to observe if
the interaction occurred? If it did not occur? How might you explain the failure of this approach to
demonstrate this interaction?
Answer Fuse CFP to �-arrestin and YFP to the cytoplasmic domain of the �-adrenergic re-
ceptor, or vice versa. In either case, illuminate at 433 nm and observe at both 476 and 527 nm.
If the interaction occurs, emitted light intensity will decrease at 476 nm and increase at 527 nm
on addition of epinephrine to cells expressing the fusion proteins. If the interaction does not
occur, the wavelength of the emitted light will remain at 476 nm. There are several reasons
why this might fail; for example, the fusion proteins (1) are inactive or otherwise unable to
interact, (2) are not translocated to their normal subcellular location, or (3) are not stable to
proteolytic breakdown.
8. EGTA Injection EGTA (ethylene glycol-bis(b-aminoethyl ether)-N,N,N�,N�-tetraacetic acid) is a
chelating agent with high affinity and specificity for Ca2�. By microinjecting a cell with an appropriate
Ca2�-EGTA solution, an experimenter can prevent cytosolic [Ca2�] from rising above 10�7
M. How
would EGTA microinjection affect a cell’s response to vasopressin (see Table 12–4)? To glucagon?
Answer Vasopressin acts through a PLC-coupled GPCR. The IP3 released by PLC normally el-
evates cytosolic [Ca2�] to 10�6 
M, activating (with diacylglycerol) protein kinase C. Preventing
this elevation of [Ca2�] by using EGTA to “buffer” the internal [Ca2�] would block vasopressin
action, but should not directly affect the response to glucagon, which uses cAMP, not Ca2�, as
its intracellular second messenger.
9. Amplification of Hormonal Signals Describe all the sources of amplification in the insulin receptor 
system.
Answer The amplification results from catalysts activating catalysts—including protein
kinases that act in enzyme cascades. Two molecules of insulin activate an insulin receptor
dimer for a finite period, during which the receptor phosphorylates many molecules of IRS-1.
Through a series of interactions with other proteins (including Grb2, Sos, Ras), IRS-1
activates Raf,which phosphorylates and activates many molecules of MEK, each of which
phosphorylates and activates many molecules of ERK. Each activated ERK phosphorylates
and activates several molecules of a transcription factor, and each of these stimulates the
transcription of multiple copies of mRNA for specific genes. Each mRNA can direct the
synthesis of many copies of the protein it encodes. (See Fig. 12–15.)
Chapter 12 Biosignaling S-133
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