Phosphorylation of WAVE1 protein remodels neuronal connections

Paper marks 50th anniversary of Paul Greengard’s first publication in journal Nature. 

Within the brain, branched nerve cell extensions called dendrites play a key role in how cells communicate with one another by sending electrical signals from the tip of one neuron to the tip of the next – a process known as synaptic transmission. Scientists have recently begun to appreciate how the shapes of dendrites affect memory and learning, but the molecular events that underlie these changes remain largely unknown.

Now, new research from Paul Greengard’s laboratory at Rockefeller University explains how a protein called WAVE1 regulates the polymerization of actin, a major cytoskeleton protein in spines that protrude from dendrites, and thereby alters the number of spines. The finding, published in the August 17 issue of the journal Nature, also marks a milestone in Greengard’s career: 2006 is the 50th anniversary of Greengard’s first published paper in Nature.

In the new study, Greengard and a group of collaborators from the United States, South Korea and France, were interested in the role of a brain protein called CDK5, which Greengard and others had previously shown to be important in both normal and abnormal brain functions. Using an assay that searches for proteins that interact with CDK5, Kim and colleagues found that WAVE1 had a strong affinity for P35, the regulatory subunit of CDK5, and that P35 mediates this interaction.

Scientists have known that WAVE1 regulates actin polymerization and actin rearrangement is important for dendritic spine morphology, but the Greengard lab shows for the first time that WAVE1 regulates actin polymerization and dendritic spine morphology through phosphorylation, a process by which a phosphate chemical group is attached to certain amino acid sites on a protein. Proteins are switched on and off, respectively, through phosphorylation and dephosphorylation.

Greengard, Kim and their colleagues found that CDK5 phosphorylates WAVE1 at three specific sites, which inhibits WAVE1’s ability to regulate actin polymerization. In experiments in which roscovitine, an inhibitor of CDK5, was added to mouse brain slices, phosphorylation of WAVE1 decreased. In addition, phosphorylation was reduced in cortical neurons cultured from CDK5-deficient mice.

The researchers also found that WAVE1 purified from rat brain can stimulate actin polymerization in the test tube, and that phosphorylation of WAVE1 by CDK5, particularly at Serine 310, WAVE1’s primary phosphorylation site, interferes with WAVE1’s ability to stimulate actin polymerization. Furthermore, WAVE1-deficient neurons have fewer dendritic spines, and this phenotype can be rescued by expression of a dephosphorylation-mimic mutant of WAVE1 but not by expression of a phosphorylation-mimic mutant.

A recent study by other researchers showed that WAVE-1-deficient mice have sensorimotor retardation and impairments in learning and memory, which Greengard and his colleagues suggest could be related to the abnormal spine morphology they describe in this study.

“Spines contain both bundled and branched forms of filamentous actin and it is likely that WAVE1 contributes to the formation of spines, as well as to remodelling of the actin cytoskeleton in spines during development and in processes affecting synaptic plasticity,” Greengard says.

The researchers also found that stimulation of a signaling cascade called the cAMP pathway in brain slices or in hippocampal neurons results in decreased WAVE1 phosphorylation and in increased spine density in hippocampal neurons. Because WAVE1 is largely phosphorylated in brain, Greengard and his colleagues hypothesize that WAVE1 remains inactive until triggered by neurotransmitters, such as dopamine, which increase levels of cAMP and decrease phosphorylation of WAVE1.

“Further clarification of the mechanisms by which CDK5, cAMP and other signaling pathways regulate WAVE1 phosphorylation and dephosphorylation should lead to an increased understanding of the regulation of spine morphogenesis and synaptic function in developing and adult brain,” says Greengard.