Scientists from the Francis Crick Institute and the Leibniz
Institute of Molecular Pharmacology in Berlin have turned the field
of protein phosporylation on its head by showing that
phosphorylation of multiple protein sites by the same enzyme does
not, as previously thought, act in the same direction.
They propose a new mechanism whereby these modifications trigger
opposing effects in being able to activate as well as inhibit the
function of a protein substrate.
Reversible protein phosphorylation in response to cell signaling
is implicated in many human diseases including many cancers. It is
the focus of extensive research into ways to treat these
diseases.
Phosphorylation is a process where a chemical modification
called a phosphate group is added to a protein, activating or
deactivating it or otherwise changing its function.
A number of different enzymes are involved in phosphorylation.
Enyzmes called kinases add phosphates and enzymes called
phosphatases remove them, resetting their function to
prestimulation level. Until now it was thought that kinases and
phosphatases act together and unidirectionally, relying on each
other's antagonistic activities to adjust protein activity.
Anastasia Mylona, a postdoc in Richard Treisman's group at the
Crick, led the work. The researchers studied a protein called
Elk-1, that plays an important role in turning the genes on when a
cell receives a signal to proliferate. Such signals activate a
kinase called ERK which adds phosphate groups at multiple sites
on
Elk-1.
Dr Treisman explains: "Our work discovered a phosphorylation
switch in which the action of ERK turns Elk-1 activity both on and
off. Sites that become modified fast act positively, whereas sites
that are phosphorylated more slowly act negatively. The kinase thus
first acts positively, but with time it also phosphorylates the
slow sites, and limits Elk-1 activity. Importantly, this
'resetting' does not require the action of a phosphatase - that is,
the active removal of phosphorylation marks."
With their collaborators in Berlin, led by Philip Selenko, the
research team tracked the phosphorylation process of Elk-1 using
time-resolved NMR spectroscopy, a technique that enabled them to
look at both the sites of phosphorylation and simultaneously
measure their rates of modification.
They then used mathematical modeling to understand how these
rates depended on and affected each other, leading to the
classification of fast, intermediate and slow Elk-1 phosphorylation
sites.
Dr Treisman says: "Our results propose a new way in which cells
can respond to a signal that is non-linear - that is, not strictly
proportional to its strength. This may be particularly important in
the Elk-1 system, which controls a gene network that determines
cells' sensitivity to growth promoting signals."
The paper, Opposing effects of Elk-1 multisite phosphorylation
shape its response to ERK activation, is published in Science.