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The P212L mutation in CaMKIIalpha, an enzyme important for learning and memory, is linked to intellectual disability. However, the exact process by which the mutation affects the enzyme’s activity was unclear, until now. A newly developed method of protein analysis has shown that the P212L mutation causes dramatically more activation of CaMKIIalpha.

“So far, there has been no effective medical treatment for children with genetically based intellectual disability. This study can offer the possibility of treatment to patients with intellectual disability who have this rare variant of CaMKII,” said Assistant Professor Hiroyuki Kidokoro from Nagoya University, a co-author of the study and pediatrician.

The team’s paper was just published in Frontiers in Molecular Neuroscience.

About 1% of the global population lives with an intellectual disability. There are several commonly known causes including infection, injury, or genetic conditions. CaMKIIalpha is an enzyme that mediates biochemical reactions in the brain and is important for our ability to learn.

Typical learning requires that CaMKIIalpha activity is regulated at appropriate levels and at the appropriate timing, and irregularities with it have previously been linked to a variety of neurological disorders. A P212L mutation in CaMKIIalpha is one cause of intellectual disability. although exactly how the enzyme’s mutation alters its function was not previously understood.

The mutation of P212L is very rare, but a patient was identified in Japan who then took part in a study with researchers from the University of Tokyo and Nagoya University.

“This is only the fourth known case of this P212L mutation in the world. However, the relationship between this single gene mutation and symptoms is relatively clear, making it important in the study of intellectual disability,” said Hajime Fujii, lecturer from the Graduate School of Medicine at the University of Tokyo.

Fujii noted that, “It is difficult to process many samples in parallel and not possible to measure enzyme activity in physiological conditions, such as living cells or synapses. We wanted something more simple, scalable, sensitive and quantitative. So, we developed a method to measure enzyme activity by fluorescent probe.”

He explained that, “This can tell us the progress of a biochemical reaction by its brightness or color. In order to create a fluorescent probe, we had to couple the biochemical reaction, such as molecular binding or changes in protein shape (that occur at the scale of a nanometer), with fluorescence brightness or color. So, we used a physical phenomenon called FRET (Förster resonance energy transfer), with which the probe can change relative brightness between two colors according to changes of the CaMKIIalpha that occur when it is activated. We call our approach a FRET-based kinase phenotyping strategy.”

This new method enabled the team to rapidly and accurately analyze nearly a hundred cell extracts and study their biological activity. They found that CaMKIIalpha with the P212L mutation exhibited enhanced activation compared to wild type. This suggests rewiring or changes in the brain that usually occur during learning may possibly be irregular in people with this mutation, compared to people without it. The researchers also found that in neurons, taken from rats in this study, the CaMKIIalpha response to stimulation was increased. The activation response of the enzyme rose faster and fell slower, again demonstrating an unusually enhanced response.

The team hopes that its research will help to identify treatment options for genetically based intellectual disabilities.

Looking to the future, Fujii said, “Mutations in CaMKIIalpha have been related to other neurodevelopmental disorders, so we may be able to clarify the development of and treatment strategies for these mutations in the same way, by applying this FRET-based kinase phenotyping platform.”

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