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Biophysics of Proteins at Surfaces: Assembly, Activation, Signaling
Tuesday Speaker Abstracts
Membrane Surface Activation of Protein Kinases C
Juan Gomez-Fernandez
, Senena Corbalan-Garcia.
University of Murcia, Murcia, Spain.
Classical protein kinases C are known to be important in cell physiology both in terms of health
and disease. They are activated by triggering signals that induce their translocation to
membranes. The consensus view is that several secondary messengers are involved in this
activation, such as cytosolic Ca
2+
and diacylglycerol. The former bridges the C2 domain to
anionic phospholipids as phosphatidylserine in the membrane and diacylglycerol binds to the C1
domain. Both diacylglycerol and the increase in Ca
2+
concentration are assumed to arise from the
extracellular signal that triggers the hydrolysis of phosphatidylinositol-4,5-bisphosphate,
however results obtained during the last decade indicate that this phosphoinositide itself is also
responsible for modulating classical PKC activity and its localization in the plasma membrane.
Novel protein kinases C, on the other hand, are known to be activated in a Ca
2+
-independent
way, with diascylglycerol/phorbol esters, playing a very important role. Recent results indicate
that the C2 domain may also play an important role in this activation and, furthermore,
negatively charged phospholipids are also very important in the binding of C1B domains to the
membrane, participating in the activation of these isoenzymes. A picture emerges in which there
is a concerted interplay of activators modulating the translocation of PKCs to the membrane,
triggering conformational changes that give place to a strictly regulated activation of these
enzymes.
Protein Kinase B (PKB) and Phosphoinositide dependent Kinase 1 (PDK1) regulation at
membranous compartments.
Banafshe LARIJANI
1,2,1
, Gloria De Las Heras-Martinez
2
, Jose Requejo-Isidro
2
, Veronique
Calleja
3
.
1
Ikerbasque Basque Foundation for Science and Universidad del País Vasco, Leioa,
Spain,
2
Biophysics Unit - CSIC, Leioa, Spain,
3
CRICK INSTITUTE, LONDON, United
Kingdom.
Protein Kinase B (PKB)/Akt and Phosphoinositide dependent Kinase 1 (PDK1) are members of
the AGC kinase superfamily and are activated downstream of many growth factor and hormone
receptors as a result of phosphoinositide 3-kinase activation. PKB and PDK1 phosphorylate a
diverse set of substrates involved in many fundamental aspects of cell biology, including growth,
survival, proliferation, angiogenesis, migration, and metabolism.
The most prominent site of PKB recruitment and activation is at the plasma membrane; however,
this may not be the only site. Therefore we have investigated the potential for intracellular PKB
activation in response to growth factor stimulation using the rapalogue dimerisation tool to
inducibly and acutely recruit Akt to the endosomes or to the nuclear envelope.
We have also investigated, in cells, the mechanism of regulation of PDK1 in response to the
level of negatively charged phospholipids at the plasma membrane using time resolved Forster
resonance energy transfer.
Our cross-disciplinary approach has resulted in determining a refined model for the in situ,
regulation of both these master regulators.