Fragile X mental retardation protein interaction with neuronal voltage-gated calcium channels: mechanism and consequences

Fragile X syndrome is the most common form of inherited severe cognitive impairment, and is usually due to the loss, or sometimes the malfunction, of Fragile X mental retardation protein (FMRP).

It is usually caused by a piece of repetitive DNA just before this gene, called the CGG repeat region. This piece of DNA becomes mutated so that it is much longer in Fragile X syndrome, and interferes with the proper function of the gene. The faulty gene is on the X chromosome and is found in 1 in 2500-4000 men and 1 in 6000-8000 women. The prevalence of people carrying this elongation has been estimated to be up to 1 in 130-250 of females. In females carrying pre-mutations (55-200 CGG repeats), the repeat length may expand to become full mutations in their children. However females with a pre-mutation or even a full mutation often do not themselves show symptoms due to a second, normal gene on their other X-chromosome.

Normal brain development requires FMRP, and its absence results in developmental delay, cognitive impairment, behavioural problems and motor incoordination. It also results in peripheral symptoms including hypersensitivity to touch, self injury and altered intestinal motility. The loss of FMRP causes no gross brain abnormalities, but connections between nerve cells in the brain are immature and there is a loss of development of these cells. There are some parallels between the autistic features of Fragile X syndrome and other forms of autism.
In my research group we have recently found that a protein (CaV2.2) that is commonly found in nerve cells, appears to associate with FMRP. This may relate to some of the behavioural deficits that are characteristic of Fragile X syndrome, particularly the changes that are seen outside the brain, in the regulation of pathways controlling touch and pain.

CaV2.2 is a type of calcium ion channel, present in the plasma membrane of nerve cells, which allows these cells to communicate between them. We will probe the interaction between FMRP and CaV2.2 to examine whether how this interaction affects the function of the channels, and the processes in which these channels are involved. We will pay particular attention to communication between pain sensing nerve cells and nerve cells in the spinal cord, which are involved in the transmission of pain signals. It is possible that disruption of this pathway might underlie some of the behavioural problems seen in those with Fragile X syndrome, particularly hypersensitivity to touch. Regulation of CaV2.2 function is highly relevant to touch hypersensitivity in animal models of nerve damage-induced pain, and self-injury behaviour is also one of the behavioural features of in some of those with Fragile X syndrome.

In summary, the newly-identified interaction between FMRP and CaV2.2 may play a role in the behavioural features and pathology of Fragile X syndrome, and may at some point in the future lead to novel avenues for treatment.

We will examine a novel molecular interaction involving Fragile X mental retardation protein (FMRP). This may have a bearing on the behavioural deficits that are characteristic of Fragile X syndrome, including mental retardation and autistic behaviour which results from a loss of the expression of FMRP. People with Fragile X also have peripheral symptoms, including heightened tactile sensitivity, and gastrointestinal motility changes.
Dynamic regulation of CaV2.2 calcium channel trafficking and turnover is important for the functions of these channels in neurons, and may occur on a rapid timescale. CaV2.2 channels are critical for neurotransmission both in central neurons, particularly early in development, and in the autonomic and sensory nervous system. Thus they are the main mediators of neurotransmission between primary sensory neurons involved in nociception and other modalities, and the spinal cord.
Regulation of N-type channel function is highly relevant to neuropathic pain, and self-injurious behaviour is one of the features of Fragile X syndrome. Thus, the newly-identified interaction between FMRP and CaV2.2 may play a role in the pathology and behavioural features of Fragile-X syndrome.
We will probe the interaction between FMRP and CaV2.2 (N-type) voltage-gated calcium channels that we have recently identified, which markedly inhibits the functional expression of CaV2.2 currents. We will define the interaction biochemically and biophysically, make peptide reagents to inhibit the association between the two proteins, and examine the role of the interaction, both in terms of the effect of FMRP on N-type currents in DRGs, and the in the contribution of CaV2.2 channels to synaptic transmission. As a subsidiary Aim we will examine the regulation by N-type channels of FMRP localization and its role in regulation of mRNA translation.

1. Who will benefit from this research?
i) Patient and carer group beneficiaries: We are carrying out this research in order to understand the role of the novel interaction that we have identified between Fragile X mental retardation protein (FMRP) and voltage-gated calcium channels, which may relate to a presynaptic role for FMRP. We expect that, by understanding more fully the complex and interacting functions of FMRP, we can increase the possibilities for early intervention to reduce the neuronal dysfunction in Fragile X syndrome. Therefore this research will be of most direct interest and benefit to those Fragile X syndrome and their carers.
Potential beneficiaries include not only people with Fragile X syndrome, but also their carers and carriers of fragile X mutations or pre-mutations. The potential for genetic screening of at-risk groups will only increase the number of people known to be in this group.
ii) Commercial private sector: potential for application and exploitation: We do not envisage commercial exploitation being feasible within the lifetime of this grant; however it is possible that we will develop tools that can interfere with the interaction between FMRP and CaV2.2. More details are given in the Impact statement.
iii) Public and charity sector beneficiaries: As soon as this work is funded, we will inform the Fragile X society UK and provide them with a summary of the research aims for their website, and participate in meetings. We will do the same for the FRAXA Research Foundation of the USA. Since we have not yet published in this area, we have not yet communicated this work to these societies.
iv) Beneficiaries within the wider public: There are some parallels between the autistic features of Fragile X syndrome and other forms of autism. It is therefore likely that this study will be of interest to the wider public, and will inform other studies on autism research.
v) Research staff beneficiaries The staff working on this grant will develop not only their generic research and professional skills which will be of future use in the UK's knowledge economy, but they will also develop generic skills of data analysis, public speaking, scientific writing that will be of use in future careers in both the public sector/universities and in the private sector.

2. How will these groups benefit from this research?
i) Patient and carer group beneficiaries: The PI (ACD) and the post-doctoral RA Laurent Ferron will undertake the impact activities, will write the material for Fragile X society websites, and communicate our research activities to carer groups associated with the Fragile X societies.
ii) Commercial private sector potential for application and exploitation: Highlighting the role of N-type calcium channels in the self-harming behaviour of patients with Fragile X syndrome, might be important with regard to therapy for this condition. More details are given in the Impact statement.
iii) Public and charity sector beneficiaries: We will make every effort to speak to patient/care groups via the Fragile X society UK and the FRAXA Research Foundation USA on our research, and we will attend meetings of these societies both to present the findings of our research, and to inform ourselves about the latest research, the tools available for Fragile X research, and about potential ways of publicising our work further.
iv) Beneficiaries within the wider public: It is highly likely that this study will be of interest to the wider public, not only to those who are involved as patient or care with Fragile X syndrome, but also for those who have family or friends with autism or those more generally interested in the mechanisms and potential treatments for autism.