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Receptor Coupled Cell Signaling

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Receptor Coupled Cell Signaling Powered By Docstoc
					Lecture Notes Series I. 2008
Cell Signaling
Biology 4407B, Biochemistry 4806B, Neuroscience 4376B
Pharmacology 5409B




   .
       G protein Coupled Receptors
              and G proteins

             Dr. Melanie Kelly, Department of Pharmacology,
             Dalhousie University.
             mkelly@dal.ca



                                                              1
  G proteins Couple Receptors to
             Effectors

Receptors are Seven Transmembrane Serpentine
receptors

 Receptors are specialized for interaction with G
proteins

Effectors are enzymes (Phospholipases, Kinases,
Synthases) and other target molecules such as ion
channels.

                                                     2
             GPCR Superfamily
 1983 rhodopsin was the first G-protein-coupled
  receptor to be cloned and was found to have seven
  hydrophobic segments, constituting 7
  transmembrane segments.

 7TM receptors now known to be the largest
  superfamily of proteins in eucaryotes.

 GPCRs signal via coupling to guanine nucleotide
  binding proteins.

 7 TM receptors may exist as oligomeric protein
  structures and linked to multiple G-proteins to initiate
  pleiotropic effects.

                                                         3
G-Protein-Coupled Receptors Have
 Evolved Through Divergent and
      Convergent Evolution



               20%   70%
        70%




                                   4
There are Several Families of
G-Protein-Coupled Receptors




                                5
Rhodopsin- The Founding
  member of Family A!
                Members of Family A
               constitute aprox. 90% of all
               GPCRs.
                Rhodopsins are activated
               by light and turn on signaling
               pathways that leads to
               vision.




                                         6
Rhodopsin Cycle




                  7
   Structural Characteristics of
        Rhodopsin Family




 Conserved residues largely located in transmembrane segments are a
“finger-print signature” of the Rhdopsin-like GPCR family              8
       7TM Receptor Family B




 Family B is a 7 TM family of peptide and neurohormone receptors that
do not have the identifying conserved residues of the rhodopsin-like
receptors.
 Family B have a large N-terminal extracellular domain containing 6
conserved cysteine residues.
 All receptors in this family stimulate AC and couple to Gs proteins.
 Some members of family B interact with chaperone proteins called
RAMPs (receptor activity modifying proteins).
                                                                         9
         7TM Receptor Family C




 Contains receptors for metabotropic glutamate receptors, Ca2+-sensing
receptor (CaSR), the -aminobutyric acid (GABA)B receptor, some
pheromone receptors and taste receptors.
 Most family 3 GPCRs have a large extracellular domain responsible for
ligand recognition and all Family C receptors function as dimers.         10
    Guanine Nucleotide Binding
       Regulatory Proteins
 GTP binding proteins act as transducers between receptors and
effectors

 Two major classes of G proteins have been described in the
literature: Heterotrimeric G proteins consisting of ,  subunits and
low molecular weight monomeric G proteins.

*  subunits of trimeric G proteins (39-52 kDa) characterizes the
individual G protein and have a high affinity Mg2+ -modulated
nucleotide binding site.
*  subunits are 35-36 kDa and the -subunit is 7-16 kDa.

* Low-molecular weight monomeric G proteins are 18-32 kDa and
include ras-related proteins and many oncogene products.
                                                                         11
Heterotrimeric G Protein
        Diversity
           G proteins derived from large gene family
          at least 16 genes encode -subunit, 4 genes
          encode -subunits and multiple genes encode
          -subunit.

           A given subtype may have more than one
          isotype:
          Gs includes 4 isotypes one of which is Golf
          Gi consists of Gi1,G i2, G i3
          Gq consists of G15, G16, G14, G11,
          Gq
          G12 consists of G12 and G13

          In mammals 5 distinct -subunits and
          several different -subunits have been
          identified.

                                               12
Interactions between G proteins
         and Receptors
                  (a) Agonist binds to receptor

                  (b) G protein interacts with
                  activated receptor and GDP
                  exchanged for GTP on  subunit
                  and subunits dissociate (?)

                  (c) Activated  subunit interacts
                  with other signaling molecules. 
                  subunits also signal.

                  (d)  -subunit possess slowly
                  hydrolysing intrinsic GTPase
                  activity. RGS proteins enhance
                  hydrolysis of GTP to GDP.

                  (e) -subunit inactivated and
                  recycled back to associate with 
                  subunits.
                                                   13
The Life-cycle of GPCR




                         14
     Agonists and Antagonists Act by
      Stabilizing Active and Inactive
        Receptor Conformations
 7TM Receptors are in equilibrium between active and inactive
conformations. -Dynamic allosteric proteins that interchange between
different conformations that are stabilized by ligands

 An agonist -stimulates the receptor by stabilizing an active
conformation
 by various mechanisms including binding between transmembrane
segments

 An antagonist is a compound that stabilizes one or more of the
many different inactive conformations of the receptor and thus
prevents the receptor from going into the active signaling
conformation.
                                                                   15
    Desensitization Mechanisms Turn
     Signaling Off During Prolonged
               Stimulation
 G protein-coupled receptor signalling is attenuated by
phosphorylation by protein kinases and interaction with an intracellular
protein, arrestin, which binds to the phosphorylated receptor and
promotes dissociation of G protein

 Ligand-occupied receptors are then sequestered into endocytic
intracellular vesicles where the ligand is degraded and the receptor
dephosphorylated and shunted back to the membrane.

 After repeated or prolonged activation downregulation, receptors
can be delivered to lysosomes and degraded after internalization. This
results in a decreased response which is only recoverable with
synthesis of new receptor molecules.
                                                                       16
Desensitization Mechanisms




                             17
Kinases and Arrestins involved in
       desensitization and
    internalization of GPCRs
 GRKs – G protein receptor kinases –
homologous desensitization.

 Protein Kinase C and Protein Kinase A –
heterologous desensitization.


                                        18
Diversity of GPCRs, G proteins and
     their signaling pathways




                                     19
Some G protein-coupled Receptors
Activate cAMP-dependent Signaling
            Pathways




                                    20
   Some G protein-coupled Receptors
Activate Phospholipid Signaling Pathways




                                       21
G Protein Receptor Cross-
Talk for Signal Amplification




                Selbie and Hill, 1998   22
                TIPS 19: 87-93
     Bacterial Toxins Can Regulate The
   Activity of Specific G protein  Subunits

 Several bacterial toxins modify G protein activity by a process
called ADP-ribosylation. This process involves the addition of an
ADP-ribose group from nicotinamide adenine dinucleotide (NAD+)
to an amino acid residue in the  subunit.

 Cholera Toxin – ADP-ribosylates and irreversibly activates Gs
by inhibiting its GTPase activity. This leads to prolonged pathway
activation.

 Pertussis Toxin - ADP-ribosylates and inactivates Gi and Go
by stabilizing their association with  subunits. This leads to
inhibition of GPCR pathways that couple to Gi/o G proteins.


                                                                     23

				
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posted:11/14/2011
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