Medical Physiology, 3rd Edition

Mechanisms of Cellular Communication

Cells can communicate with one another via chemical signals

Cells secrete chemical signals that can induce a physiological response in any (or all) of three ways (Fig. 3-1): by entering the circulation and acting on distant tissues (endocrine), by acting on a neighboring cell in the same tissue (paracrine), or by stimulating the same cell that released the chemical (autocrine). Secreted factors that are produced by cells of one organ and enter the circulation to induce a response in a separate organ are called hormones, and the organs that secrete them—such as the pituitary, adrenal, and thyroid glands—are parts of the endocrine system. However, many other cells and tissues not classically thought of as endocrine in nature also produce hormones. For example, the kidney produces 1,25-dihydroxyvitamin D3, and the salivary gland synthesizes nerve growth factor. Finally, physical contact between one cell and another, or between a cell and the matrix secreted by another cell, can transmit a signal (juxtacrine).

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FIGURE 3-1 Modes of intercellular communication.

For paracrine and autocrine signals to be delivered to their proper targets, their diffusion must be limited. This restriction can be accomplished by rapid endocytosis of the chemical signal by neighboring cells, its destruction by extracellular enzymes, or its immobilization by the extracellular matrix. The events that take place at the neuromuscular junction are excellent examples of paracrine signaling. When an electrical impulse travels down an axon and reaches the nerve terminal (Fig. 3-2), it stimulates release of the neurotransmitter acetylcholine (ACh). In turn, ACh transiently activates a ligand-gated cation channel on the muscle cell membrane. The resultant transient influx of Na+ causes the membrane potential (Vm) to shift locally in a positive direction (i.e., depolarization), initiating events that result in propagation of an action potential along the muscle cell. The ACh signal is rapidly terminated by the action of acetylcholinesterase, which is present in the synaptic cleft. This enzyme degrades the ACh that is released by the neuron.

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FIGURE 3-2 Example of paracrine signaling. The release of ACh at the neuromuscular junction is a form of paracrine signaling because the nerve terminal releases a chemical (i.e., ACh) that acts on a neighboring cell (i.e., the muscle).

Soluble chemical signals interact with target cells via binding to surface or intracellular receptors

Four types of chemicals can serve as extracellular signaling molecules:

1. Amines, such as epinephrine

2. Peptides and proteins, such as angiotensin II and insulin

3. Steroids, including aldosterone, estrogens, and retinoic acid

4. Other small molecules, such as amino acids, nucleotides, ions (e.g., Ca2+), and gases (e.g., nitric oxide)

For a molecule to act as a signal, it must bind to a receptor. Most receptors are proteins on the cell surface or within the cell that specifically bind a signaling molecule (the ligand) and induce a cellular response by interacting with an effector. In some cases, the receptor is itself the effector, as in ligand-gated ion channels that alter transmembrane ion conductance in response to an extracellular signal. In most cases, however, interaction of the ligand with its receptor results in association of the receptor with one or more intracellular effector molecules that in turn initiate the cellular response. Effectors include enzymes, channels, transport proteins, contractile elements, and transcription factors. The ability of a cell or tissue to respond to a specific signal is dictated by the complement of receptors it possesses and by the chain of intracellular reactions that are initiated by the binding of any one ligand to its receptor. Receptors can be divided into five categories on the basis of their mechanisms of signal transduction (Table 3-1).

1. Ligand-gated ion channels. Integral membrane proteins, these hybrid receptors/channels are involved in signaling between electrically excitable cells. The binding of a neurotransmitter such as ACh to its receptor—which in fact is merely part of the channel—results in transient opening of the channel and thus alters the ion permeability of the cell.

2. G protein–coupled receptors. These integral plasma-membrane proteins work indirectly—through multiple intermediaries—to activate or inactivate downstream effectors, such as membrane-associated enzymes or channels. This group of receptors is named for the initial intermediary, which is a heterotrimeric GTP-binding complex called a G protein.

3. Catalytic receptors. When activated by a ligand, these integral plasma-membrane proteins are either enzymes themselves or part of an enzymatic complex. In many cases, these receptors are either kinases that add phosphate groups to their substrates or phosphatases that remove substrate phosphate groups.

4. Nuclear receptors. These proteins, located in the cytosol or nucleus, are ligand-activated transcription factors. These receptors link extracellular signals to gene transcription.

5. Receptors that undergo cleavage. In response to ligand binding, some transmembrane proteins undergo regulated intramembranous proteolysis (RIP; see pp. 87–88), image N3-1which liberates one or more fragments of their cytosolic domain that signal a cellular response by entering the nucleus to modulate gene expression.

TABLE 3-1

Classification of Receptors and Associated Signal-Transduction Pathways

CLASS OF RECEPTOR

SUBUNIT COMPOSITION OF RECEPTOR

LIGAND

SIGNAL-TRANSDUCTION PATHWAY DOWNSTREAM FROM RECEPTOR

Ligand-gated ion channels (ionotropic receptors)

Heteromeric or homomeric oligomers

Extracellular

Ion current

GABA

Cl > image

Glycine

Cl > image

ACh: muscle

Na+, K+, Ca2+

ACh: nerve

Na+, K+, Ca2+

5-HT

Na+, K+

Glutamate: non-NMDA

Na+, K+, Ca2+

Glutamate: NMDA

Na+, K+, Ca2+

ATP (opening)

Ca2+, Na+, Mg2+

Intracellular

 

cGMP (vision)

Na+, K+

cAMP (olfaction)

Na+, K+

ATP (closes channel)

K+

IP3

Ca2+

Ca2+ or ryanodine

Ca2+

Receptors coupled to heterotrimeric (αβγ) G proteins

Single polypeptide that crosses the membrane seven times

Small transmitter molecules

ACh

Norepinephrine

Peptides

Oxytocin

PTH

NPY

Gastrin

CCK

Odorants

Certain cytokines, lipids, and related molecules

βγ directly activates downstream effector

Muscarinic AChR activates atrial K+ channel

α activates an enzyme

Cyclases that make cyclic nucleotides (cAMP, cGMP)

Phospholipases that generate IP3 and diacylglycerols

Phospholipases that generate AA and its metabolites

Catalytic receptors

Single polypeptide that crosses the membrane once

ANP
TGF-β

Receptor guanylyl cyclase
Receptor serine/threonine kinases

May be dimeric or may dimerize after activation

NGF, EGF, PDGF, FGF, insulin, IGF-1

Receptor tyrosine kinase

IL-3, IL-5, IL-6, EPO, LIF, CNTF, GH, IFN-α, IFN-β, IFN-γ, GM-CSF

Tyrosine kinase–associated receptor

CD45

Receptor tyrosine phosphatase

Intracellular (or nuclear) receptors

Homodimers of polypeptides, each with multiple functional domains

Steroid hormones

Bind to regulatory DNA sequences and directly or indirectly increase or decrease the transcription of specific genes

Mineralocorticoids
Glucocorticoids
Androgens
Estrogens
Progestins

Heterodimers of polypeptides, each with multiple functional domains

Others

Thyroid hormones
Retinoic acid
Vitamin D
Prostaglandin

Cleavage-activated receptors

Single polypeptide that crosses the membrane once

Jagged
Delta

After receptor cleavage, cytosolic domain of receptor translocates to nucleus and regulates gene transcription

CCK, cholecystokinin; NMDA, N-methyl-D-aspartate; NPY, neuropeptide Y; PTH, parathyroid hormone.

N3-1

Examples of RIP

Contributed by Peter Igarashi

In addition to sterol regulatory element–binding protein (SREBP) noted in the text on pp. 87–88, other proteins that undergo RIP are Notch and APP—all span the membrane at least once.

Notch is a plasma-membrane receptor whose cytoplasmic domain is released in response to Delta, a membrane-bound ligand that regulates cell fate during development.

Amyloid precursor protein (APP) is a protein of unknown function that is cleaved in the membrane to produce the extracellular amyloid β peptide implicated in Alzheimer disease. For Notch and APP, the intramembrane cleavage does not take place until a primary cleavage event removes the bulk of the protein on the extracytoplasmic face. Although the cleaved sites differ in these proteins, the net effect of the first step is to shorten the extracytoplasmic domain to <30 amino acids, which allows the second cleavage to release a portion of the cytoplasmic domain.

The epithelial Na+ channel ENaC—a heterotrimer that consists of α, β, and γ subunits—also undergoes intramembrane proteolysis. While the ENaC is still in the vesicular subcompartment of the secretory pathway, the protease furin cleaves the extracellular domain of the α subunit twice (after consensus RXXR motifs), releasing a peptide of 26 amino acids. By itself, this modification increases the open probability of ENaC to ~0.30. Furin also cleaves the γ subunit, but only once. A variety of extracellular proteases—including prostasin, elastase, and plasmin—can make a second cut after the protein has reached the plasma membrane. The result of the γ-subunit cleavages is to increase the open probability to nearly 1.0. Thus, both examples of proteolysis greatly increase the activity of the channel.

Reference

Hughey RP, Carattino MD, Kleyman TR. Role of proteolysis in the activation of epithelial sodium channels. Curr Opin Nephrol Hypertens. 2007;16:444–450.

Signaling events initiated by membrane-associated receptors can generally be divided into six steps:

Step 1: Recognition of the signal by its receptor. The same signaling molecule can sometimes bind to more than one kind of receptor. For example, ACh can bind to both ligand-gated channels and G protein–coupled receptors. Binding of a ligand to its receptor involves the same three types of weak, noncovalent interactions that characterize substrate-enzyme interactions. Ionic bonds are formed between groups of opposite charge. In van der Waals interactions, a transient dipole in one atom generates the opposite dipole in an adjacent atom and thereby creates an electrostatic interaction. Hydrophobic interactions occur between nonpolar groups.

Step 2: Transduction of the extracellular message into an intracellular signal or second messenger. Ligand binding causes a conformational change in the receptor that triggers the catalytic activities intrinsic to the receptor or causes the receptor to interact with membrane or cytoplasmic enzymes. The final consequence is the generation of a second messenger or the activation of a catalytic cascade.

Step 3: Transmission of the second messenger's signal to the appropriate effector. The second messenger can be amplified and transmitted to distant regions within the cell by myriad events, such as activation of intracellular kinases and phosphatases that alter the activity of other enzymes and proteins, release or sequestration of intracellular ions, or regulation of metabolic pathways that generate ATP.

Step 4: Modulation of the effector. These effectors represent a diverse array of molecules, such as enzymes, ion channels, cytoskeletal components, and transcription factors. The second message can modulate their expression or activity as well as alter their location or substrate availability.

Step 5: Response of the cell to the initial stimulus. This collection of actions represents the summation and integration of input from multiple signaling pathways.

Step 6: Termination of the response by feedback mechanisms at any or all levels of the signaling pathway.

Cells can also communicate by direct interactions—juxtacrine signaling

Gap Junctions

Neighboring cells can be electrically and metabolically coupled by means of gap junctions formed by the interaction of connexins in two closely apposed cell membranes (see pp. 158–159). These water-filled channels facilitate the passage of inorganic ions and small molecules, such as Ca2+ and cyclic 3′,5′-adenosine monophosphate (cAMP), from the cytoplasm of one cell into the cytoplasm of an adjacent cell. Mammalian gap junctions permit the passage of molecules that are less than ~1200 Da but restrict the movement of molecules that are greater than ~2000 Da. Gap junctions are also excellent pathways for the flow of electrical current between adjacent cells, playing a critical role in cardiac and smooth muscle.

The permeability of gap junctions can be rapidly regulated by changes in cytosolic concentrations of Ca2+, cAMP, and H+ as well as by the voltage across the cell membrane or Vm. This type of modulation is physiologically important for cell-to-cell communication. For example, if a cell's plasma membrane is damaged, Ca2+ passively moves into the cell and raises [Ca2+]i to toxic levels. Elevated intracellular [Ca2+] in the damaged cell triggers closure of the gap junctions, which prevents the flow of excessive amounts of Ca2+ into the adjacent cell.

Adhering and Tight Junctions

Adhering junctions form as the result of the Ca2+-dependent interactions of the extracellular domains of transmembrane proteins called cadherins (see p. 17). The clustering of cadherins at the site of interaction with an adjacent cell causes secondary clustering of intracellular proteins known as catenins, which in turn serve as sites of attachment for the intracellular actin cytoskeleton. Thus, adhering junctions provide important clues for the maintenance of normal cell architecture as well as the organization of groups of cells into tissues.

In addition to playing a homeostatic role, adhering junctions can serve a signaling role during organ development and remodeling. In a cell that is stably associated with its neighbors, a catenin known as β-catenin is mainly sequestered at the adhering junctions, so that the concentration of free β-catenin is minimized. However, disruption of adhering junctions following stimulation by certain growth factors, for example, causes β-catenin to disassociate from cadherin. The resulting rise in free β-catenin levels promotes the translocation of β-catenin to the nucleus. There, β-catenin regulates the transcription of multiple genes, including ones that promote cell proliferation and migration. image N3-2

N3-2

β-Catenins

Contributed by Lloyd Cantley

The concentration of free β-catenin in the cytoplasm is regulated by a cluster of cytosolic proteins (including the adenomatous polyposis coli [APC] protein that is mutated in some colon cancers) that bind and phosphorylate β-catenin, targeting it for degradation. Thus, inhibition of this regulatory pathway, such as occurs following activation of Wnt signaling during organ development, will increase β-catenin levels and enhance nuclear localization and transcriptional regulation.

Similar to adhering junctions, tight junctions (see pp. 43–44) comprise transmembrane proteins that link with their counterparts on adjacent cells as well as intracellular proteins that both stabilize the complex and have a signaling role. The transmembrane proteins—including claudins, occludin, and junctional adhesion molecule (JAM)—and their extracellular domains create the diffusion barrier of the tight junction. One of the integral cytoplasmic proteins in tight junctions, zonula occludin 1 (ZO-1), colocalizes with a serine/threonine kinase known as WNK1, image N3-3 which is found in certain renal tubule epithelial cells that reabsorb Na+ and Cl from the tubule lumen. By phosphorylating specific claudins in the tight-junction complex, WNK1 determines the paracellular Cl permeability and thereby helps regulate NaCl uptake from the glomerular filtrate. Mutations in WNK1 that increase the movement of Cl through the tight junctions (see pp. 754–755) cause increased salt reabsorption by the kidney and lead to hypertension.

N3-3

WNK Kinases

Contributed by Lloyd Cantley

WNK kinases are serine/threonine kinases that are unique in that they lack an otherwise highly conserved lysine residue in the catalytic domain (specifically, subdomain II) of the enzyme. Hence, they are kinases “with nK”—that is, no lysine residue (the single-letter code for lysine is K). These kinases have emerged as highly important regulators of epithelial ion transport in the kidney, predominantly via phosphorylation-dependent changes in ion channel surface localization and activity.

Membrane-Associated Ligands

Another mechanism by which cells can directly communicate is the interaction of a receptor in the plasma membrane with a ligand that is itself a membrane protein on an adjacent cell. Such membrane-associated ligands can provide spatial clues in migrating cells. For example, an ephrin ligand expressed on the surface of one cell can interact with an Eph receptor on a nearby cell. The resulting activation of the Eph receptor can in turn provide signals for regulating such developmental events as axonal guidance in the nervous system and endothelial-cell guidance in the vasculature.

Ligands in the Extracellular Matrix

Cells also receive input from their extracellular environment via cell-surface receptors that interact with the extracellular matrix. These receptors, called integrins (see p. 17), have an extracellular domain that interacts with amino-acid sequences that are specific to certain matrix components, such as collagen, fibronectin, and laminin. This ligand-receptor interaction results in a conformational change in the integrin that promotes the accumulation of a cluster of signaling molecules—called a focal adhesion—on the cytosolic side of the membrane. The focal adhesion includes small GTP-binding proteins (see p. 56) such as Rac and Rho that regulate actin cytoskeletal anchoring and turnover, as well as intracellular kinases such as focal-adhesion kinase (FAK) and Src that control such diverse processes as cell proliferation, cell migration, and cell differentiation. Thus, by sensing a change in the matrix composition, cells can activate migratory responses needed for organ repair or can undergo terminal differentiation during organ development.

Second-messenger systems amplify signals and integrate responses among cell types

Once a signal has been received at the cell surface, it is typically amplified and transmitted to specific sites within the cells via second messengers. For a molecule to function as a second messenger, its concentration, activation, and location must be finely regulated. The cell achieves this control by rapidly producing or activating the second messenger and then inactivating or degrading it. To ensure that the system returns to a resting state when the stimulus is removed, counterbalancing activities function at each step of the cascade.

The involvement of second messengers in catalytic cascades provides numerous opportunities to amplify a signal. For example, the binding of a ligand to its receptor can generate hundreds of second-messenger molecules, which can in turn alter the activity of thousands of downstream effectors. This modulation usually involves the conversion of an inactive species into an active molecule or vice versa. An example of creating an active species would be G proteins that elicit the formation of the second messenger cAMP (see pp. 56–57), whereas an example of destroying an active species would be G proteins that elicit the breakdown of a related second messenger, cGMP; (see p. 58).

Second-messenger systems also allow both specificity and diversity. Distinct ligand-receptor combinations that activate the same intracellular signaling pathways often produce the same cellular response. For example, epinephrine, adrenocorticotropic hormone (ACTH), glucagon, and thyroid-stimulating hormone (TSH) all signal via cAMP to induce triacylglycerol breakdown. In contrast, a single signaling molecule can produce distinct responses in different cells, depending on the complement of receptors and signal-transduction pathways that are available in the cell as well as the specialized function that the cell carries out in the organism. For example, ACh stimulates contraction of skeletal muscle cells, inhibits contraction of heart muscle cells, and facilitates the exocytosis of secretory granules by pancreatic acinar cells. This signaling molecule achieves these different end points by interacting with distinct receptors on each cell. Finally, even within a single cell, a second messenger such as cAMP can act within microdomains to target a specific set of effector molecules, thereby conferring spatially based specificity. image N3-4

N3-4

Compartmentalization of Second-Messenger Effects

Contributed by Laurie Roman

In the textbook, we referred only to whole-cell levels of intracellular second messengers (e.g., cAMP), as if these messengers were uniformly distributed throughout the cell. However, some cell physiologists and cell biologists believe that local effects of intracellular second messengers may be extremely important in governing how signal-transduction processes work. One piece of evidence for such local effects is that the receptors for hormones and other extracellular agonists often are a part of macromolecular clusters of proteins that share a common physiological role. For example, a hormone receptor, its downstream heterotrimeric G protein, an amplifying enzyme (e.g., adenylyl cyclase) that generates the intracellular second messenger (e.g. cAMP), other proteins (e.g., the A kinase anchoring protein [or AKAP]), and the effector molecule (e.g., protein kinase A) may all reside in a microdomain at the cell membrane. Thus, it is possible that a particular hormone could act by locally raising [cAMP]i to levels much higher than in neighboring areas, so that—of all the cellular proteins potentially sensitive to cAMP—only a local subset of these targets may be activated by the newly formed cAMP.

A second piece of evidence for the local effects of cAMP is the wide distribution of phosphodiesterases, which would be expected to break down cAMP and limit its ability to spread throughout the cell.

The diversity and specialization of second-messenger systems are important to a multicellular organism, as can be seen in the coordinated response of an organism to a stressful situation. Under these conditions, the adrenal gland releases epinephrine. Different organ systems respond to epinephrine in a distinct manner, such as activation of glycogen breakdown in the liver, constriction of the blood vessels of the skin, dilation of the blood vessels in skeletal muscle, dilation of airways in the lung, and increased rate and force of heart contraction. The overall effect is an integrated response that readies the organism for attack, defense, or escape. In contrast, complex cell behaviors, such as proliferation and differentiation, are generally stimulated by combinations of signals rather than by a single signal. Integration of these stimuli requires crosstalk among the various signaling cascades.

As discussed below, most signal-transduction pathways use elaborate cascades of signaling proteins to relay information from the cell surface to effectors in the cell membrane, the cytoplasm, or the nucleus. In Chapter 4, we discuss how signal-transduction pathways that lead to the nucleus can affect the cell by modulating gene transcription. These are genomic effects. Signal-transduction systems that project to the cell membrane or to the cytoplasm produce nongenomic effects, the focus of this chapter.