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Chap. 16 Signaling Pathways That Control
Gene Expression
Topics
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Receptor Tyrosine Kinases (RTKs)
The Ras/MAP Kinase Pathway
Phosphoinositide Signaling
TGFß Receptors and Smad Signaling
Goals
• Learn the properties of RTKs.
• Learn about RTK signaling via the
Ras/MAP kinase signaling pathway.
• Learn the general features of the
PI-3 kinase signaling pathway.
• Learn about the TGFß/Smad
signaling pathway.
EGF receptor bound to EGF
Major Classes of Cell-surface Receptors
About one dozen classes of
cell-surface receptors occur
in human cells. An overview
of signaling by the two
receptor systems that are
covered in this chapter
(receptor tyrosine kinases,
RTKs, and TGF-ß receptors)
is shown in (Fig. 16.1a).
The signal transduction
pathways used by RTKs are
summarized in (Fig. 16.2).
Activation of RTKs via Ligand Binding
Receptor tyrosine kinases (RTKs) regulate cell differentiation and
proliferation. Ligands of RTKs include nerve growth factor (NGF),
fibroblast growth factor (FGF), and insulin. In some cases (e.g.,
the epidermal growth factor (EGF) RTK), ligand binding causes
receptor dimerization (Fig. 16.3). In other cases (e.g., the
insulin RTK), binding occurs to pre-existing dimers. RTKs exhibit
intrinsic tyrosine kinase activity located within their cytosolic
domains. The binding of ligand activates the kinase domains which
cross-phosphorylate the two monomers of the dimeric receptor.
Phosphorylation first occurs at a regulatory site known as the
activation lip.
Phosphorylation of
the lip causes
conformational
changes that allow
the kinase domain to
phosphorylate other
tyrosine residues in
the receptor and in
signal transduction
proteins.
Recruitment of Signal Transduction
Proteins to Activated Receptors
Signal transduction system
proteins interact with
phosphorylated RTKs via
phosphotyrosine binding
domains. Two main binding
domains--PTB and SH2
(Src homology domain-2)-within signal transduction
proteins such as the
multi-docking protein
known as the insulin
receptor substrate-1
(IRS-1) perform this
function (Fig. 16.12). The
binding of signaling
proteins either directly to the receptor or to IRS-1 allows
them to be phosphorylated by the receptor. Some of these
signaling proteins are involved in activation of the Ras
GTPase (next slides). Some such as phosphatidylinositol-3
kinase (PI-3 kinase) participate in lipid-mediated signaling
pathways. RTK signaling typically is down-regulated by
endocytosis of receptors from the cytoplasmic membrane.
RTKs and Ras/MAP Kinase Signaling
Nearly all RTKs signal via Ras/MAP kinase pathways. They also
may signal via other pathways. For example, the insulin receptor
uses the Ras/MAP kinase pathway to regulate gene expression
and the PI-3 kinase pathway to regulate enzyme activity (e.g.,
glycogen synthase). RTK-Ras/MAP kinase signaling controls cell
division, differentiation, and metabolism.
Ras is a monomeric (small) GTPase switch protein that unlike
trimeric G proteins does not directly bind to receptors. Ras
typically relies on guanine nucleotide-exchange factors (GEFs)
for binding GTP, and on GTPase-activating proteins (GAPs) for
stimulation of GTP hydrolysis. Once activated, Ras propagates
signaling further inside the cell via a kinase cascade that
culminates in the activation of members of the MAP kinase
family. MAP kinases phosphorylate TFs that regulate genes
involved in the cell cycle and in differentiation. Mutant RTKs or
Ras/MAP kinase signaling proteins are associated with nearly all
cancers. Dominant Ras mutations that block GAP binding and lock
Ras in the "on" state promote cancer.
RTK Activation of Ras
The mechanism by which EGF activates
Ras is illustrated in Fig. 16.17. In
Step 1, EGF binding causes receptor
dimerization and autophosphorylation on
cytosolic tyrosines. In Step 2, the
adaptor protein GRB2 binds receptor
phosphotyrosine residues via its SH2
domain. GRB2 contains SH3 domains
that allow the GEF protein known as
Sos to bind to the membrane complex.
Sos then recruits Ras to the complex.
In the last step of Ras activation (Step
3), Sos promotes GTP exchange for
GDP on Ras. The activated Ras-GTP
complex then dissociates from Sos, but
remains tethered to the inner leaflet
of the cytoplasmic membrane via a lipid
anchor sequence. The active form of
Ras then activates the MAP kinase
portion of the signaling pathway (next
two slides).
Ras Activation of MAP Kinase
Ras activates MAP kinase via a
phosphorylation cascade that
proceeds from Ras to Raf kinase, to
MEK kinase, and finally to MAP
kinase (Fig. 16.20). MAP kinase then
dimerizes and enters the nucleus
(next slide).
MAP Kinase Activation of Transcription
In the final steps of RTK-Ras/MAP kinase signaling, MAP
kinase phosphorylates and activates the p90RSK kinase in the
cytoplasm (Fig. 16.22). Both kinases enter the nucleus
where they phosphorylate ternary complex factor (TCF) and
serum response factor (SRF), respectively. The
phosphorylated forms
of these TFs bind to
serum response
element (SRE)
enhancer sequences
that control genes
such as c-fos. c-fos
activates the
expression of genes
that propel cells
through the cell
cycle. SREs occur in a
number of genes that
are regulated by
growth factors
present in serum.
Signaling via Phosphatidylinositol 3-phosphates
RTKs also can signal via formation of
phosphoinositide compounds. Like
GPCRs, they signal via the IP3/DAG
pathway. However, RTKs activate the
PLCg isoform of phospholipase C, not
PLCß as occurs with GPCRs. PLCg binds
to activated RTKs via SH2 domains.
In addition, RTKs can signal via PI
3,4-bisphosphate and PI 3,4,5trisphosphate formed by the enzyme
PI-3 kinase (Fig. 16.25). PI-3 kinase
is recruited to the membrane by SH2
domain-mediated binding to activated
RTK phosphotyrosine residues. The PI
3-phosphate compounds synthesized by
PI-3 kinase activate protein kinase B
(PKB) (next slide).
Activation of Protein Kinase B
Signaling downstream of PI 3-phosphates is conducted by PKB
(Fig. 16.26). PKB is recruited to the membrane via binding to PI
3-phosphates via its PH domain. There it is phosphorylated and
activated by the PDK1 & PDK2 kinases. PDK1 also is recruited to
the membrane via binding to PI 3-phosphates. Activated PKB then
enters the cytosol, where it phosphorylates target proteins. In
insulin receptor signaling, PKB phosphorylates and inactivates
glycogen synthase kinase, stimulating glycogen synthesis. PKB also
is a potent inhibitor of apoptosis. PI 3-phosphate signaling
ultimately is terminated by cleavage of 3-phosphates from
phosphoinositides by the PTEN phosphatase. PTEN is inactive in
many advanced cancers.
Biological Roles of TGFß
Growth factors are proteins that play important roles in
regulating cell differentiation, division, and movement.
Activating mutations in growth factor receptors or their
signaling pathways commonly are associated with cancers.
Transforming growth factor ß (TGFß) plays widespread roles in
regulating development in both vertebrates and invertebrates.
Despite their names, all three types of human TGFßs exert
anti-proliferative effects on target cells. Therefore, the loss
of a TGFß receptor can lead to transformation of a cell to a
cancerous state. TGFß is secreted from cells as an inactive
precursor. It subsequently undergoes proteolytic processing and
attaches to the extracellular matrix. It is released from the
matrix after the receipt of an appropriate signal, and then
carries out paracrine signaling on neighboring cells.
TGFß/Smad Pathway
The signal transduction pathway by
which TGFß regulates TF activity
is illustrated in Fig. 16.28. The
points to know for this pathway
are 1) TGFß binds to cell surface
receptors causing phosphorylation
of these receptors, 2) the
activated receptors phosphorylate
certain Smad TFs exposing their
nuclear localization signals, and 3)
Smad TFs enter the nucleus where
they combine with other TFs and
activate the transcription of
target genes. Smad TF activity
ultimately is shut down via the
action of transcription repressors,
whose activity is induced by TGFß
via a feedback loop. These
repressors bind to the Smad TFs,
and recruit a histone deacetylase
to the activated promoter, which
then causes chromatin
condensation and gene silencing.
Because TGFß has antiproliferative
effects on cells, overexpression of
the repressors results in cellular
transformation and cancer.