Cell migration: Rho GTPases lead the way

1
Cell migrationRho GTPases lead the way

• Xia Fan

2
How migration occurs?

• The major driving force of migration is the
  extension of a leading edge protrusion or
  lamellipodium, the establishment of new adhesion
  sites at the front, cell body contraction, and
  detachment of adhesions at the cell rear.

3
The Rho GTPase switch(Rho,Rac,Cdc42)

• Cycle between a GDP-bound, inactive form
  GTP-bound, active form.
• The cycle is regulated by
• GEFpromote the exchange of GDP for GTP
• GAPenchance the intrinsic GTPase activity
• GDIblock the cycle by sequestering and
  solubilizing he GDP-bound form
• Predominantly through GEF.
• How they regulate migration
• Rhoassembly of contractile actinmyosin
  filaments
• Racpolymerization of actin to form lamellipodial
• Cdc42polymerization of actin to form filopodial
  protrusion
• all promotes the assembly of integrin-based,
  matrix adhesion complex

4
Regulation of Rho GTPases during cell migration

• Observation Rac can be visualized with highest
  concentrations at the leading edge.
• Focus Rac activation-two pathways

5
PI-3-kinase and PI(3,4,5)P3

• Extracellular signal activated receptor
  protein activated PI-3-kinase PI(3,4,5)P3
  docked Rac GEF
• How Rac GEF promotes GTP loading
• PI(3,4,5)P3 binding relieves interaction between
  PH and DH domain to stimulate activity
• Loss of activity due to deletion of PH domain is
  restored by addition of a CAAX motif to stable
  the binding.(The DH domain induces Rac to
  displace GDP and bind to GTP. The DH domain is
  invariably proceeded by a PH domain, which
  greatly increase catalytic efficiency.)
• PI(3,4,5)P3 regulates PIX, which activates Rac
  and interacts with PAK
• Rac activation stimulates PI 3-kinase, resulting
  a positive feedback loop

6
p130Cas/CrkII/DOCK80

• Extracellular signal activated receptor
  protein activated PI-3-kinase PI(3,4,5)P3
  docked DOCK180
• How DOCK180 promotes GTP loading
• Bind to the adaptor protein Crk, which in turn
  associates with another adapor p130Cas. The
  Crk/p130Cas/DOCK180 can lead to Rac activation.
• Two plausible explanation of the biochemical
  mechanism
• DOCK180 recruits a DH-containing GEF to the
  complex.
• The C-terminus of DOCK180 can stimulate GTP
  loading on Rac.
• The basic region of DOCK180 seems to gunction in
  the same way as the PH domain of
  DH-domain-containing GEF

7
Downstream effects of Rho GTPases during cell
migration

• Regulation of the actin cytoskeleton
• Rac generates a protrusive force through the
  localized polymerization of actin.
• Cdc42 generates filopodia through the localized
  polymerization of actin.
• Rho generates focal adhesion and cell
  contractility by across-linking actinmyosin
  filaments.
• Regulation of the microtubule and polarity
• Efficient and persistent long-range migration
  requires stabilizeion of cell polarity and is
  achieved through reorganization of the
  microtubule cytoskeleton.
• Cdc42 plays a crucial role in polarized migration
  by reorientation of microtubules and centrosome.

8
Regulation of the actin cytoskeleton

• Either Rac or Cdc42 can activate p65PAK Cdc42
  activates WASp and N-WASpRac activates the
  Scar/WAVE.
• p65PAK
• 1.regulates focal adhesion turnover with
  help of PIX and GIT1(mechanism is unkonwn)
• 2.phosphorlates and activates LIMK, which in
  turn phosphorylates and inactivates
  cofilin.(confilin faciliates subunit dissociation
  from the pointed end of actin filaments and
  induces filament severing and is essential fro
  promoting filament treadmilling at the front)
• WASP/SCAR/WAVE
• 1.stimulates the Arp2/3 complex, which can
  initiate actin polymerization.
• 2.WASP/WAVE can bind to profilin, which acts
  synergistically with Arp2/3 to speed up actin
  polymerization.

9
Regulation of the actin cytoskeleton

• Rho can activates p160ROCK and mDia
• p160ROCK
• 1.works like p65PAK,leading to stabilization
  of actin filaments within actinmyosin filament
  bundles.
• 2.interacts with and phosphorylates the
  myosin binding subunit of myosin light chain
  phosphatase and inactivates it,leading to
  increased levels of myosin phosphorylation, which
  then can cross-link actin filaments.
• mDia is linked to actin filament assembly.
  (mechanism is unknown)

10
Regulation of the actin cytoskeleton
11
The microtubule cytoskeleton and polarity

• Rhoactivates mDia, which directly interacts with
  microtubules and promotes their capping, leading
  the stabilization of microtubules.
• Racpromotes microtubule elongation through
• 1. p65PAK-dependent phosphorylation by
  activating p65PAK
• 2.inactivating stathmin, the microtubule
  destabilizing protein.
• Cdc42 can activates the complex Par6/PKC.
• PKC phosphorylates and inactivates GSK-3.
• This induces the association of APC with the
  plus ends of microtubules specifically at the
  leading edge.
• Through a dynein-or dynactin-dependent
  mechanism, this results in microtubule
  reorganization and centrosome reorientation.
• (APC could also serve to localize Asef, a
  Rac-specific GEF, to sites of Rac-dependent actin
  polymerization)

12
The microtubule cytoskeleton and polarity