Biochemistry: Chapter 20

In the late 1980s, Robert Weinberg isolated DNA from bladder cancer cells and used this to transfect normal mouse fibroblasts. Which of the following statements is NOT true of this experiment?

Sequence analysis of the transforming gene H-ras was found to contain multiple mutations.

Which of the following statements about hormones are correct? 1) Hormones can be peptides, steroids, or amino acid derivatives. 2) Hormones can stimulate the synthesis of target proteins through the activation of specific genes. 3) Hormones can directly activate or inhibit enzymes through the action of signal molecules. 4) Hormones can increase the cellular uptake of metabolites.

1, 2, and 4 are correct.

Lithium ion inhibits the synthesis of inositol trisphosphate by inhibiting a reaction in the breakdown of inositol trisphosphate. Choose the correct explanation of this apparent paradox.

Because inositol trisphosphate is derived from phosphatidylinositol, which in turn is derived from diacylglycerol and inositol, inositol phosphates must be completely dephosphorylated in order to be reincorporated into phosphatidylinositol for another round of synthesis.

Hormone action is ultimately controlled by the cells that secrete the hormone.

False

Most synaptic transmission events are mediated electrically through gap junctions between cells.

False

Receptor tyrosine kinases are membrane-spanning proteins with an intrinsic tyrosine kinase domain on the extracellular side of the membrane.

False

Which of the following is NOT a secondary messenger? cAMP nitric oxide sn-1,2-diacylglycerol inositol 1,4,5-trisphosphate

nitric oxide

The membrane-bound receptor in G protein-coupled signal transduction is quite separate to the effector enzyme which catalyses the formation of an intracellular ________.

second messenger

Nuclear receptors:

are transcriptional regulators.

Bisphenol A (BPA) is widely used as a building block in polymer synthesis and is found in the polycarbonate hard plastics of reusable drink containers, DVDs, cell phones, and other consumer goods. Bisphenol A is reported to have estrogenic activity, and its widespread occurrence in our environment is a potential concern. Choose all the suggestions that correctly describe biochemical experiments that could be done to compare the activity of bisphenol A with that of its estradiol, its structural relative. Select all that apply.

- Look directly for binding of radiolabeled BPA to receptors for estrogens and related hormones. - Check for competitive or synergistic effects of coadministration of BPA and estradiol. - Compare the effects on transcription of estradiol and bisphenol A in estrogen-responsive cells.

List factors that make it advantageous for peptide hormones to be synthesized as inactive prohormones that are activated by proteolytic cleavage. Check all that apply.

- Signal sequence on the prohormone molecule could direct the protein to the correct subcellular site, then be cleaved off when no longer needed. - Suppression of the hormonal signal until it is needed. - Additional polypeptide sequence may promote correct folding of the polypeptide chain.

Describe a mechanism by which a steroid hormone might act to increase intracellular levels of cyclic AMP. Check all that apply.

- Steroid hormones could activate transcription of the gene for adenylate cyclase and, hence, increase the steady-state level of this enzyme. - The mechanism could include induced synthesis of a phosphodiesterase inhibitor or repressed synthesis of cAMP phosphodiesterase.

With a better understanding of these three types of hormone signaling, the next question you should be asking yourself is how can a transmembrane protein either allow a molecule or ion to transverse the membrane, or how can they initiate the production of a secondary messenger or a metabolic pathway. Complete the paragraph to describe how hormone interaction with a transmembrane protein can signal the cell. Match the words in the left column to the appropriate blanks in the sentences on the right.

1. Hormone interaction with a transmembrane protein to signal the cell involves ALLOSTERY, which is the idea that the binding of a ligand to a protein at one site on the protein can cause a(n) 3-DIMENSIONAL CHANGE in the protein. 2. In the case of ion channels, the hormone binding causes a 3-DIMENSIONAL CHANGE that results in a(n) pore forming which allows an ion to transverse the membrane. 3. In the two other types, the hormone molecule that binds on the extracellular side of the transmembrane protein causes a conformational change located on the INTRACELLULAR portion of that same protein. The change result in the activation or opening of an active site. This new, active site then catalyzes the synthesis of a second messenger, or initiates the first step of a specific metabolic pathway.

The Foundation Figure introduces classes of plasma membrane-bound receptors that play key roles in signal transduction. Signal transduction refers to the reception of an extracellular stimulus that leads to a metabolic change within a cell. Fill in the blanks in the paragraph below to given an overview regarding how extracellular stimulus can transmit signals. Match the words in the left column to the appropriate blanks in the sentences on the right.

1. There are three main methods of transmitting an extracellular signal into a cell where metabolic change occurs. The simplest might be to create a CHANNEL from one side of the membrane to the other. This pore could allow a specific extracellular molecules (often ions) into the cytosol initiating metabolic changes. 2. Of the three methods, two involve transmitting an extracellular stimulus through membrane bound proteins or receptors. An extracellular stimulus could interact with a membrane-bound protein and directly initiate a metabolic change through a SIGNAL CASCADE within the cell. 3. The second method involving transmitting an extracellular stimulus through membrane bound proteins or receptors would be that the signal induces a SECOND MESSENGER and that molecule initiates the metabolic change.

Match the correct hormone class (on the left) with the correct protein (on the right) whose synthesis is increased by hormonal stimulation. 1) progesterone 2) 1,25-dihydroxy-vitamin D3 3) glucocorticoids 4) androgens A) calcium binding protein B) phosphoenolpyruvate carboxykinase C) uteroglobin D) aldolase

1:C; 2:A; 3:B; 4:D

Arrange the steps of G-protein activation in the proper order. 1) cAMP synthesis results from the activation of adenylate cyclase. 2) GTP exchanges with bound GDP. 3) The signal binds to the extracellular component of the receptor, inducing a conformational change. 4) cAMP activates several kinases. 5) The G protein activates adenylate cyclase.

3, 2, 5, 1, 4

Arrange the events of glycogen synthesis in the proper order. 1) The receptor dimerizes. 2) Proteins, such as IRS-1, are recruited to the phosphorylated tyrosine domain of the receptor. 3) Autophosphorylation of the intracellular domain takes place. 4) Insulin binds to the insulin receptor. 5) IRS-1 is phosphorylated, which then activates phosphoinositide 3-kinase. 6) Protein kinase B is activated. 7) Phosphatidylinositol 4,5-bisphosphate is converted to phosphatidylinositol 3,4,5-trisphosphate. 8) Protein kinase B phosphorylates glycogen synthase kinase 3, inactivating it.

4, 1, 3, 2, 5, 7, 6, 8

Mutations that inactivate p53 have a recessive phenotype, whereas mutations affecting Ras are dominant. Complete the explanation of this difference. Drag the terms on the left to the appropriate blanks on the right to complete the sentences. Terms can be used once, more than once, or not at all.

A cell heterozygous for p53 could still make sufficient p53 from its one functional gene to carry out its SIGNALING FUNCTION, giving the heterozygote a wild-type function. Both genes must be inactive to block a p53 damage response. With respect to RAS, the presence of one functional gene is sufficient to initiate the abnormal GROWTH RESPONSE. Similar phenomena are seen generally in genetic analysis, where a LOSS-OF-FUNCTION mutation is typically recessive, whereas a GAIN-OF-FUNCTION mutation is dominant.

A. Choose correct statements describing features of insulin signaling that affect glucose utilization. Select all that apply. B. A β-adrenergic response can be modulated through the actions of a receptor kinase and arrestin, because phosphorylation by the kinase desensitizes the receptor. How might signaling by a tyrosine receptor kinase, such as the insulin receptor, be modulated?

A. - The action of phosphoinositide 3-kinase stimulates an enzyme phosphorylation cascade that results in enhanced glycogen synthesis. - Over the long term insulin receptor activation stimulates protein synthesis and cell division. - Glucose uptake into cells is stimulated through mobilization of glucose transporters to the membrane. B. Modulation occurs through action of protein tyrosine phosphatases on phosphorylated tyrosine residues.

Signaling molecules interact with cells through specific macromolecular receptors. For each of the four receptors identified below, check all characteristics, which accurately describe that receptor. A. An adrenergic receptor __________ Check all that apply. B. A steroid receptor __________ Check all that apply. C. The LDL receptor __________ Check all that apply. D. The insulin receptor __________ Check all that apply.

A. - is located at the cell surface - is a transmembrane protein - activation can inhibit the synthesis of glycogen - is a receptor whose biological activity involves interaction with guanine nucleotide-binding proteins B. - is a DNA-binding protein - is located in the cell interior - is a receptor whose receptor-ligand complex becomes concentrated in the nucleus - is a receptor whose the hormone-receptor complex activates specific gene transcription - is not known to act through a second messenger C. - is located at the cell surface - is associated with the protein clathrin - is a receptor whose receptor-ligand complex moves to the lysosome - action diminishes the synthesis and activity of β-hydroxy-β-methylglutaryl-CoA reductase (HMG-CoA reductase) - is not known to act through a second messenger D. - is located at the cell surface - is a transmembrane protein - is a receptor whose the hormone-receptor complex activates specific gene transcription - has a protein kinase activity

A. Which of these is a receptor molecule? B. A signal transduction pathway is initiated when a ________ binds to a receptor. C. Which of these is a signal molecule? D. A signal molecule is also known as a(n) ________. E. Which of these is the second of the three stages of cell signaling?

A. B B. signal molecule C. A D. ligand E. transduction

A. Which of these acts as a second messenger? B. Which of these is responsible for initiating a signal transduction pathway? C. What role does a transcription factor play in a signal transduction pathway? D. Which of these is a membrane receptor? E. A signal transduction pathway is initiated when a ________ binds to a receptor. F. Which of these acts as a second messenger? G. Calcium ions that act as second messengers are stored in ________. H. ________ catalyzes the production of ________, which then opens an ion channel that releases ________ into the cell's cytoplasm. I. A protein kinase activating many other protein kinases is an example of ________.

A. D B. A C. By binding to DNA it triggers the transcription of a specific gene. D. B E. signal molecule F. cyclic AMP G. endoplasmic reticula H. Phospholipase C ... IP3 ... Ca2+ I. amplification

Caffeine is an inhibitor of cyclic AMP phosphodiesterase. A. How would drinking several cups of coffee affect muscle function? B. How might it affect lipid metabolism?

A. Muscle function would be enhanced. B. Lypolysis would be promoted.

Studies on cAMP actions in cultured cells usually involve adding to the cell culture not cAMP, but dibutyryl cAMP (see structure below). A. Why is this structural modification necessary? B. How could you test the premise that di-Bu-cAMP has the same biochemical effects as cAMP? Select all that apply.

A. Negative charge on the phosphate of cAMP prevents its passage through cell membranes. The butyrate chains make the molecule more lipophilic, although the negative charge on the phosphate is unaffected. B. - One could assess the effect of dibutyryl-cAMP on purified cAMP target proteins. - One could compare the actions of cAMP and dibutyryl-cAMP in cells whose plasma membrane had been treated with an agent to increase its permeability.

Suppose that you measured binding to the isolated EGF receptor of EGF at various concentrations. A. Would you expect the binding curve to be hyperbolic? B. Choose the correct explanation of your answer.

A. No, the binding curve will not be hyperbolic. B. The reaction of EGF with its receptor should be trimolecular at low EGFR concentrations, because EGF binding should promote dimerization of the monomeric receptor.

A. Which of the following best describes the mechanism by which steroid hormones control gene expression? B. Which of the following best describes the role of chaperone proteins in the regulation of gene expression by steroid hormones? C. The reason some cells respond to the presence of a steroid hormone which others do not is that ________.

A. Steroid hormones that enter the cell activate receptors. These hormone-receptor complexes then bind HREs and influence gene expression. B. Chaperone proteins maintain functionality of the receptor. C. the receptors necessary for regulation differ among cells of various types

GTPγS is a nonhydrolyzable analog of GTP. Suppose this compound were added to a cell-free system containing active components of an adrenergic signaling system. A. What consequences would you expect? B. What would be the effects on cAMP levels?

A. Stimulation of adenylate cyclase is lost much slower than in case of GTP. B. cAMP levels should increase

Suppose that a G protein undergoes a mutation that allows the exchange of bound GTP for GDP to occur in the absence of G protein binding to a receptor. A. How might this mutation affect signaling involving a GPCR? B. Which subunit of the G protein is most likely affected by the mutation?

A. Such a mutation would cause hyperactivation of downstream signaling. B. α-subunit is most likely to be affected by mutation

A. What is the biochemical basis for the action of Viagra? B. What is the biochemical basis for the action of Prozac? C. Oral administration of S-adenosylmethionine has been reported to be effective in treating depression. Suggest a possible explanation. Select all that apply.

A. Viagra inhibits cyclic GMP phosphodiesterase, so cGMP levels remain high, thereby prolonging the effects of nitric oxide on regional blood flow. B. Prozac selectively inhibits reuptake of serotonin by presynaptic neurons, thereby increasing the amount of the neurotransmitter that reaches the postsynaptic cell, stimulating synaptic transmission in areas related to regulation of mood. C. - AdoMet could affect membrane fluidity through effects on phospholipid metabolism. - If AdoMet could cross the blood-brain barrier, it might conceivably increase methylation of norepinephrine to give epinephrine. - AdoMet could act through folate metabolism, increasing the pool of labile methyl groups.

A. The magnification of the signal from a water-soluble hormone is achieved through an increase in ________. B. Water-soluble hormones affect target cells by binding to ________. C. How do endocrine hormones reach their target cells? Select the best answer. D. What is the role of activated protein kinases? Select the best answer. E. Cyclic AMP is degraded by ________.

A. cAMP in the cytoplasm B. plasma membrane receptors C. Hormones are transported through the blood stream to target cells. D. Phosphorylate proteins E. phosphodiesterase

A. Choose the correct balanced equation for the hydrolysis of cGMP, catalyzed by cGMP phosphodiesterase. B. Would you expect an inhibitor of this enzyme to potentiate or antagonize the action of Viagra? Choose the correct explanation.

A. cGMP + H2O -> 5'-GMP B. Viagra acts by inhibiting this reaction, any other compound acting similarly should potentiate the effect of Viagra.

A. Hormones that use the tyrosine kinase receptor mechanism as shown in the animation are important for B. Autophosphorylation is an important part of the mechanism of the receptor tyrosine kinases. Why is this? C. Sos activates Ras. Sos is an example of a D. Receptor tyrosine kinases are activated by hormones that cause cell division (mitogens). Which of the following describes a way that a protein in the pathway could be mutated to result in its constitutive activation causing unregulated cell division? E. The dominant negative mutation of the receptor in the animation is effective in blocking the hormone signal because

A. cellular growth and survival. B. The phosphorylation provides sites for recruitment of other signal transduction molecules. C. guanine-nucleotide exchange factor (GEF). D. a mutation in Ras that makes it unable to hydrolyze GTP E. it is able to dimerize with a functional receptor without the activation of autophosphorylation.

A. What protein complex or complexes provide initial recognition of the vesicle with the target membrane? B. What is the role of Rab GTPase in the process? C. Botulinum toxin blocks neuromuscular transmission, causing paralysis. How does it do this? D. Consider the mechanism of Botulinum toxin. Is this toxin reversible? E. What is another likely role of the SNARE mechanism of vesicle fusion, in addition to secretion of material?

A. multisubunit tethering proteins and coiled-coil tethering proteins B. Rab GTPase locks the t-SNARE and v-SNARE together, facilitating fusion of the vesicle. C. Botulinum toxin cleaves the t-SNARE, blocking fusion of the vesicle. D. Yes, however the cell must regenerate new t-SNAREs. E. insertion of membrane proteins into the plasma membrane

Hormone action is one element of signal transduction mechanisms. The processes by which signals are transmitted from cell to cell include: A. Hormones may passively diffuse and interact with intracellular receptors; these act at the gene level, with the hormone-receptor complex affecting transcription of specific genes in the target tissue. B. Hormones that interact with plasma membrane receptors that are ion channels, with hormone-binding directly affecting membrane permeability to an ion. C. Hormones that interact with plasma membrane receptors that have a hormone-binding site on the exterior and an enzyme activity on the cytosolic side, with ligand binding stimulating that activity. D. Hormones that interact with plasma membrane receptors that act through G proteins to affect the levels of second messengers. Three types of signal transmission involve chemical messengers (hormones) that interact with plasma membrane receptors, while one diffuses passively. Predict which of the four processes of hormone action is represented by the figures below. Drag the appropriate processes to their respective bins.

A: picture of double helix B: Ion pore/Transmembrane subunits C: two other pictures D: Transmembrane domains

Which of the following are characteristics of G protein-coupled receptors? 1) They typically have seven transmembrane domains that are embedded in the cell membrane. 2) They carry hydrophilic portions of protein that extend into the extracellular matrix as well as the cytosol. 3) The receptor region is located on the extracellular portion of the protein. 4) Most work in concert with G proteins.

All of the listed statements are characteristics of G protein-coupled receptors.

Which of the following compounds can act as second messengers in signal transduction pathways?

Cyclic AMP

Which of the following is an extracellular messenger involved in signal transduction?

Epinephrine

Which statement about G proteins is NOT true?

G proteins possess very active GTPase activity.

The diagram shows the mechanism for hormone signaling in which the hormone molecule binds on the outside of the membrane causing an intracellular conformational change of the transmembrane protein that result in the activation, often depicted as an opening, of an active site. This new, active site then catalyzes the synthesis of a secondary messenger molecule or the first substrate molecule of a specific metabolic pathway. where the substrate and product are as shown Complete the description of the type of signal transduction event depicted. Match the words in the left column to the appropriate blanks in the sentences on the right.

If the intracellular active site then catalyzes the synthesis of an EFFECTOR molecule, which for the system shown would be the conversion of ATP to cAMP, then this hormone signaling system might be a G PROTEIN coupled system.

Suppose that you had a monoclonal antibody that recognized phosphotyrosine. How would you expect that antibody to affect insulin signaling? Drag the terms on the left to the appropriate blanks on the right to complete the sentences. Not all terms will be used.

No effect would likely be observed in a WHOLE-CELL ASSAY system because the phosphotyrosine residues are located on the INTRACELLULAR DOMAIN. But in a cell-free system, it could interfere with the interaction between insulin receptor and INSULIN RECEPTOR SUBSTRATE, IRS-1.

Upon activation by a receptor, a G protein exchanges bound GDP for GTP, rather than phosphorylating GDP that is already bound. Similarly, the α subunit-GTP complex has a slow GTPase activity that hydrolyzes bound GTP, rather than exchanging it for GDP. Complete the description of experimental evidence that would be consistent with these conclusions. Drag the terms on the left to the appropriate blanks on the right to complete the sentences. Terms can be used once, more than once, or not at all.

One could treat a G protein-GDP complex with γ-[32P]ATP and ask whether radiolabeled GTP is synthesized. The fact that it is not indicates that the G protein cannot phosphorylate the bound GDP, at least not with ATP as a phosphate donor. One can also show that radiolabeled GTP can displace bound, unlabeled GDP under various conditions. One can also show that activation of the G protein requires the presence of GTP. Finally, one can demonstrate that the isolated α subunit has GTPase activity, by showing its ability to convert GTP to GDP and Pi in the absence of added GDP.

Now that you have developed three signaling methods involving hormones, lets analyze them in more detail using the foundation figure as a guide. Match the words in the left column to the appropriate blanks in the sentences on the right.

One type involves hormones that interact with a plasma membrane receptor that act through a(n) G PROTEIN to affect the levels of a(n) SECOND MESSENGER, a nonprotein signaling molecule, that modulate a variety of intracellular metabolic processes. One type involves hormones that interact with plasma membrane receptors that have an extracellular hormone-BINDING SITE on a membrane bound protein, which directly initiates ENZYME ACTIVITY on the cytosolic side, with ligand binding stimulating that activity. One type involves hormones that interact with a plasma membrane receptor that is a(n) ION CHANNEL with binding directly affecting membrane permeability to an ion.

G proteins __________. 1) are proteins that bind guanine nucleotides 2) can either stimulate or inhibit downstream enzymes such as adenylyl cyclase 3) are activated by hydrolyzing GTP to GDP 4) are in an inactive state when GTP is bound

Only statements 1 and 2 are correct.

The Foundation Figure introduces classes of chemical messengers and their modes of actions as extracellular messengers. There are several mechanisms that have evolved in which a signal can be transmitted across the cell membrane, with the simplest being the signaling molecule being able to freely diffuse across the membrane. This process is known as passive diffusion. Predict if the following chemical messengers can passively diffuse into the cell. Drag the appropriate chemical messengers to their respective bins.

Passive diffusion possible: - active vitamin D3 - progesterone - RU486 Additional transport mechanism required: - All-trans-retinoic acid - acetylcholine - cyclic AMP (cAMP) - primary structure of bovine insulin

Which of the following statements about the adrenergic receptor is NOT true?

Propranolol is an agonist for this receptor.

In early studies of adrenergic signaling, it was thought that the epinephrine receptor and adenylate cyclase were one and the same protein. What kind of evidence would prove otherwise?

Protein fractionation, with a demonstration that epinephrine-binding activity and adenylate cyclase activity could be physically separated from one another can be done.

β-adrenergic receptors are subject to phosphorylation at several serine residues by β-adrenergic receptor kinase, and this is followed by binding to the phosphorylated sites by a protein called arrestin. Why do you think might be the purpose of these modifications?

These processes control receptor activity and modulate the adrenergic response.

How would you expect this toxin to affect blood pressure? Drag the terms on the left to the appropriate blanks on the right to complete the sentences. Not all terms will be used.

This would INACTIVATE Gi, BLOCKING the inhibition of adenylate cyclase and leading to ACCUMULATION of cAMP, with a corresponding INCREASE in blood pressure.

Which of the following does NOT belong to a receptor tyrosine kinase family?

Thyroid receptor

G proteins are membrane-bound proteins that bind GDP when inactive.

True

The end result of a signal transduction pathway is often a change in gene expression.

True

All chemical synapses exhibit the same general sequence of events during the transmission of information across the synaptic cleft. This sequence is always initiated by an action potential that travels down the presynaptic cell (the sending neuron) to its synaptic terminal(s). Drag the labels onto the flowchart to indicate the sequence of events that occurs in the presynaptic cell (orange background) and the postsynaptic cell (blue background) after an action potential reaches a chemical synapse.

a. Ca2+ channels in presynaptic membrane open briefly b. Ca2+ ions enter presynaptic cells c. neurotransmitter-counting vesicles fuse with presynaptic membrane d. neurotransmitter released into synaptic cleft e. neurotransmitter binds to ligand-gated ion channels in postsynaptic membrane; channels open f. neurotransmitter degraded or removed from cleft; ligand-gated ion channels close

Chemical synapses transmit information from the sending (presynaptic) cell to the receiving (postsynaptic) cell in the form of neurotransmitters. The release of neurotransmitter into the synaptic cleft and the resulting changes in the membrane potential of the postsynaptic cell (postsynaptic potentials) all depend on the presence of several different types of gated ion channels and the distribution of these channels in the pre- and postsynaptic cells. The image here illustrates a chemical synapse. Drag the labels onto the table to indicate which type(s) of gated ion channels are found in each membrane associated with a chemical synapse. (The letters in the table refer to the lettered structures in the image above.) Labels can be used once, more than once, or not at all.

a. voltage-gated Na+ voltage-gated K+ b. voltage-gated Na+ voltage-gated K+ voltage-gated Ca2+ c. ligand-gated d. none e. voltage-gated Na+ voltage-gated K+

Name the amino acid residue that is used to complex zinc ions in the zinc finger motif of steroid receptors.

cysteine