CHAPTER 15

CHAPTER 15Cell Signaling and Signal

Transduction: Communication Between Cells

Signal transduction• Extracellular ligand: 1st messenger

– Autocrine: ligand binds to producing cell– Paracrine: ligand binds to a neighbor cell– Endocrine: ligand binds to a distant cell after traveling through the circulatory

system

A Survey of Extracellular Messengers

• Extracellular messengers (the signal) include:– Small molecules such as amino acids, steroids, lipids and

their derivatives

– Gases such as NO and CO

– Various peptides and proteins

Edn1

serotonin

Signal transduction: theme 1 (GPCR)• Cells respond to a ligand only if

expressing the cognate receptor1) 1st messenger2) binds to cell surface receptor3) triggers conformation change in the

intracellular domain(s)

4) Activates intracellular ‘Effector’ enzymes

5) Effector generates 2nd messengers molecules inside the cell

6) 2nd messengers alter the function of further downstream factors

Signal transduction: theme 2 (RTK)• Cells respond to a ligand only if

expressing the cognate receptor1) 1st messenger2) binds to cell surface receptor3) triggers conformation change in

the intracellular domain(s)

4) Intracellular enzyme domain becomes activated (kinase)

5) Triggers a cascade of kinases activating additional downstream kinases• Target proteins ultimately alter

cell activity– E.g. TXN, TLN, Enzymes, etc

G protein-coupled receptors (GPCRs)BINDING

– GPCRs involved in vision, smell, emotion (1000s of genes)– Respond to a wide variety of ligands

• Proteins, small chemical compounds, metabolites, photons

G protein-coupled receptors (GPCRs)BINDING

– GPCRs involved in vision, smell, emotion (1000s of genes)– Respond to a wide variety of ligands

• Proteins, small chemical compounds, metabolites, photons– Ligands bind to extracellular side of receptor

• Induces a conformational change in intracellular domains– 7 transmembrane (7TM) receptors

• serpentine structure passed through membrane 7 times

1 2 3 4 5 6 7

N

C

GPCRsACTIVATION / SIGNAL TRANSMISSION

– Receptor is ‘coupled’ to a large heterotrimeric G protein “switch”• Alpha, beta and gamma subunits• Alpha subunit = GTP hydrolyzing enzyme (GTPase)• GTP bound form is switched ‘on’• GDP bound form is switched ‘off’• The ligand-bound receptor promotes exchange of GDP for GTP

1 2 3 4 5 6 7

N

C

G-GTPG-GDP

GTP GDP

Pi

Activereceptor

RGS

“off ” “on”

GPCRsACTIVATION / SIGNAL

TRANSMISSION– Four classes of G-alpha subunit

– G-alpha-s increases activity of Adenylate Cyclase (AC)• An “Effector”• ATP --> cAMP + PPi• cAMP is a 2nd messenger

generated within the cytoplasm

– G-alpha-i decreases activity of AC

ACTIVATION / SIGNAL TRANSMISSION– G-alpha-q increases activity of the effector Phospholipase C (PLC)

• PI(4,5)P2 --> DAG + IP3• DAG and IP3 are both 2nd messengers

– G-alpha-12/13 is not well understood, but is linked to cancer

GPCRsTERMINATION• Regulators of G protein

Signaling (RGS)– Enhance GTPase activity

• GPCR-kinases (GRKs)– Phosphorylate internal

portions of active receptors• Arrestins

– Compete with G-alpha for binding to phosphorylated GPCR

• Desensitization: loss of response to a stimulus in spite of the continued presence of the stimulus

GPCR: Gs• Epinephrine: glucose mobilization

– G-alpha-s– Increased AC activity– Increased [cAMP]– cAMP allosterically activates Protein

Kinase A (PKA)• PKA inhibits Glycogen Synthase• PKA activates Glycogen

Phosphorylase– Leads to glycogen breakdown

and release of glucose• PKA stimulates TXN factor activity

– Phosphorylates cAMP Response Element Binding (CREB) protein

– Drives TXN of various target genes

GPCR: Gq• Increased Phospholipase C (PLC)-beta activity

– Hydrolyzes specific phospholipids

– Increased [diacylglycerol (DAG)] and [inositol triphosphate (IP3)]

GPCR: Gq• DAG allosterically activates Protein Kinase C (PKC)• IP3 allosterically opens a Calcium channel on the smooth Endoplasmic

Retriculum– Increased [Ca2+] in cytoplasm

• Increased [Ca2+] in cytoplasm– Ca2+ binds many cytoplasmic

proteins– Calmodulin, regulator of many

proteins

GPCR: Gq

• Increased [Ca2+] in cytoplasm– Ca2+ can pass through GAP junctions into neighboring cells– Ca2+ gated Ca2+ channels in neighboring cells open further increasing [Ca2+]– Propagation of Ca2+ effects through GAP junctions integrates tissue response

(Ca2+ waves)

Video

GPCR: Gq

Receptor Tyrosine Kinases (RTKs)• Over 90 different genes• Extracellular ligand binding triggers receptor dimerization

Receptor Tyrosine Kinases (RTKs)• Intracellular tyrosine (Y) kinase domain• Dimerization allows for trans-autophosphorylation of the receptors

• Intracellular tyrosine (Y) kinase domain• Dimerization allows for trans-autophosphorylation of the receptors• Phospho-Y (PY) sequences are binding sites• SH2 domains in proteins such as Src and Grb2 bind to PY-receptor

Receptor Tyrosine Kinases (RTKs)

• Intracellular tyrosine (Y) kinase domain• Dimerization allows for trans-autophosphorylation of the receptors• Phospho-Y (PY) sequences are binding sites• SH2 domains in proteins such as Src and Grb2 bind to PY-receptor

• Drag partner proteins along with them from the cytoplasm to the membrane

Receptor Tyrosine Kinases (RTKs)

The Ras-MAPK pathway• Ras is a small G protein “switch” (no beta or gamma)

– Over 100 Ras family genes– Lipid anchor to plasma

membrane– GEF = Guanine Nucleotide

Exchange Factor (e.g. Sos)– GAP = GTPase Activating

Protein

• Mitogen Activated Protein Kinases (MAPKs)– A large family of kinases subject to regulation by phosphorylation

Ras-GTPRas-GDP

GTP GDP

Pi

GEF

GAP

“off ” “on”

Disease and the Ras-MAPK pathway• Ras G12V mutations

– Insensitive to GAP activity– Ras stuck in “on” state

• Neurofibromatosis type I– NF1 gene encodes a GAP

gene for Ras– NF1 mutations leave Ras

stuck in the “on” state

Ras-GTPRas-GDPGTP GDP

GEF

GAP

“off ” “on”

X

RTK

RTK: Insulin receptor

Convergence in signal transduction• Examples:

• Integrins:• ECM --> Integrin --> Ras

• RTKs:• EGF --> EGFR --> Ras

Crosstalk in signal transduction• Examples:

• GPCR RTK– PKA-cAMP can inhibit

Raf but activate CREB

Apoptosis (Programmed Cell Death)• ~10^10 cell deaths per day in the average human

Extrinsic: Receptor-mediated Apoptosis• Ligand: Tumor Necrosis Factor (TNF)• TNF --> TNF-Receptor -->

Procaspase recruitment• Procaspase proteolytically activated to

form “initiator” caspase• Initiator proteolytically activates

“executioner” caspases• Caspase Activated DNase (CAD)

• Irreparable DNA damage, sustained high [Ca2+]

• Activates proapoptotic factor: BAD/Bax– Punches holes in mitochondria– Release of cytochrome c from

mitochondria• Activates initiator caspase complex• Activates executioner caspases• Activates CAD

Intrinsic pathway