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Journal Reviews

Posted in Journal Reviews on 29th Sep 2014

Flying in the face of RNA foci: RAN peptide toxicity in ALS

Reviewer: Dr Jemeen Sreedharan, Dept of Neurobiology/ Neurology, University of Massachusetts Medical School, Worcester, USA.

The commonest known cause of ALS (and a common cause of FTD) is a huge intronic hexanucleotide (GGGGCC) repeat expansion in the gene C9orf72. The normal role of C9orf72 is unknown, though knockout mice (so far) seem normal, suggesting that the repeat expansion causes a toxic gain of function rather than a loss of function. But how does an expansion of hundreds to thousands of GGGGCC motifs cause neurodegeneration? Two main hypotheses have been proposed. Firstly, the RNA transcript composed of repeats forms foci within the nuclei of neurons, glia and even peripheral cells such as fibroblasts. Similar foci in myotonic dystrophy are known to sequester muscleblind, a critical RNA binding protein, whose subsequent loss of function leads to the disease. The hunt for the key proteins sequestered in C9orf72 RNA foci has so far led to inconsistent results between labs. Furthermore, it has not been convincingly shown that these RNA foci are actually directly toxic.

The second model proposes that protein toxicity underlies C9orf72 ALS and FTD. Last year it was shown that the RNA expansions, despite being intronic, are actually translated into dipeptide repeat (DPR) proteins through an unusual mechanism termed Repeat Associated Non-ATG (RAN) translation. DPR proteins were found sequestered in the brains of patients with ALS and FTD and marked C9orf72 expansion carriers out from sporadic patients. However, as with the RNA foci, it was not clear if these peptides are toxic. The work of Adrian Isaacs’ group (Mizielinska et al Science 2014) has helped to shed light on this question.

Isaacs’ team created artificial C9orf72 RNA repeat expansions (not an easy thing to do as the GGGGCC repeat is very unstable) and then expressed these in cells in vitro and also in vivo using flies. They found that these expansions formed RNA foci and were also translated into DPR proteins. These animals died prematurely. What they also did was to express modified RNA expansions in which the repeats are sparsely interrupted such that RNA foci still form, but the RNA is not translated into DPR proteins. These animals, strikingly, appeared normal. They also did the inverse, expressing DPR proteins using RNA sequences that were unable to form foci, and found that these animals died. This data collectively implicates protein rather than RNA in C9orf72-ALS-FTD.

Although there is likely to be a rush towards understanding DPR toxicity following this and another recent paper (Kwon et al Science 2014), RNA toxicity remains a possibility. One should also be aware that the fly does not have an obvious orthologue of C9orf72 and validation of dipeptide toxicity in a mouse is eagerly awaited. And, aside from the RNA and DPR proteins, what on earth does C9orf72 actually do under normal circumstances? Its biology remains unexplored.

Kwon I, Xiang S, Kato M et al. Poly-dipeptides encoded by the C9ORF72 repeats bind nucleoli, impede RNA biogenesis, and kill cells. Science. 2014 Jul 31. [Epub ahead of print]

Mizielinska S, Grönke S, Niccoli T et al. C9orf72 repeat expansions cause neurodegeneration in Drosophila through arginine-rich proteins. Science. 2014 Aug 7. [Epub ahead of print]

Good with faces. Treating prosopagnosia

Reviewer: Dr Aidan Neligan, UCL Institute of Neurology, Queen Square, London, UK.

This month sees the publication of a large multicentre collaborative meta-analysis of data from 12 previously published or unpublished genetic cohort studies to identify potential variants that increase the risk of epilepsy.

Seizures and epilepsy was classified according to the International League Against Epilepsy (ILAE) terminology into genetic generalised epilepsy, focal epilepsy or unclassified epilepsy (where there was neither electro-clinical evidence for a focal or generalised onset or alternatively if there was evidence for both focal and generalised epilepsy). In total 40,789 people comprising 10,064 people with epilepsy and 30,725 controls from 12 cohorts were studied. In the all-epilepsy analysis two loci with genome-wide significance were identified (p<1.66 x 10-8). The first signal was located at 2q24.3, which is centred on the voltage-gated sodium channel gene SCN1A which has previously been associated with genetic epilepsy with febrile seizures plus (GEFS+) and the severe epileptic encephalopathy Dravet syndrome. A second signal for the all-epilepsy phenotype was located at 4p15.1 including the 3’ end of the protocadherin gene PCDH7. This is a novel association not previously described with epilepsy. The PCDHT gene is a member of the cadherin gene family, which encodes for a calcium-dependent adhesion protein and is specifically expressed in the hippocampus and the thalmocortical pathways. Whilst the PCDH7 signal only achieved genome-wide significance for the all-epilepsy phenotype, it was more strongly associated with generalised rather than the focal epilepsies.

Analysis of the generalised epilepsies included 2,606 people with epilepsy and 18,990 controls across eight cohorts. A single signal achieved the threshold of genome wide significance; this was located at 2p16.1, the interval containing the genes encoding the vaccine-related kinase 2 (VRK2) and Fanconi anaemia, complementation group L (FANCL). Variation in VRK2 has previously been postulated as a risk factor for schizophrenia and epilepsy. VRK2 is a serine-threonine protein kinase, which is involved in cell apoptosis and signal transduction. The other gene in the interval FANCL, which encodes for a RING-type E3 ubiquitin ligase in the Fanconi anaemia pathway, has not previously been associated with a seizure phenotype.

In the meta-analysis of focal epilepsy involving 28,916 individuals no single signal achieved genome-wide significance. Finally, the authors found that none of the identified susceptibility loci with nominal genome-wide significance were associated with prognosis or outcome of newly treated epilepsy.

The importance of such work is the demonstration that with cohorts of a large enough sample size, susceptibility loci for common epilepsies can be identified through the analysis of common variation. The authors correctly conclude that whilst this work may not have immediate clinical application, it provides useful pointers to the genetic architecture of the epilepsies, which may, in time, lead to clinically applicable biomarkers of seizure prognosis and outcome.

International League Against Epilepsy Consortium on Complex Epilepsies. Genetic determinants of common epilepsies: a meta-analysis of genome-wide association studies.

Lancet Neurol 2014;13:893-903.


Reviewer: Dr Lloyd Bradley, Consultant in Rehabilitation Medicine, Western Sussex Hospitals NHS Foundation Trust, UK

Chronic regional pain syndrome (CRPS) is a poorly defined disorder that involves a range of clinical features including swelling, changes in peripheral temperature, dysasthaesia and weakness. It often occurs in a limb that has suffered some sort of trauma or other noxious insult, and may cause substantial impairment for an individual in terms of functional loss of activity through pain or weakness. It is now increasingly recognised that as well as CRPS occurring within the realm of mechanical injury, acute neurological insults (such as strokes) can precipitate a similar constellation of symptoms. Although in the past these may have been lumped within the broad category of “central neuropathic pain,” this conceptualisation of the problem as a disorder of central pain perception probably oversimplifies the aetiology and means that opportunities for thinking about different clinical interventions are lost. The interaction between somatosensation, weakness, swelling and pain in an affected limb following a stroke are very complex and may be driven by the interaction between both mechanical and primarily neurological factors.

This study retrospectively reviews a large cohort of patients admitted following a stroke who went on to develop CRPS. CRPS was defined by diffuse limitations in uptake on a bone scan. While this is the diagnostic gold standard for the diagnosis, the possible contribution of motor weakness to bone density loss following a stroke is not considered. The cohort assessed all had somatosensory evoked potentials (SEPs) performed shortly following admission post-stroke, which allowed the authors to evaluate how the measured SEPs correlated with the later presentation of CRPS. The glenohumeral distance (on x-ray) was also measured in the acute stage which allowed a similar assessment of correlation of shoulder subluxation with the development of CRPS.

The researchers identified that absence of a median SEP was strongly correlated with the later development of CRPS as well as with shoulder subluxation. There is great difficulty in defining the role of shoulder subluxation in the development of CRPS per se. The researchers postulate that it is the severity of the motor deficit that predicts onset of CRPS, however it is never really defined how this suggestion could be supported.

Given the negative impact that the development of CRPS has on stroke rehabilitation, there is a need to consider its presence as soon as possible. Early and aggressive desensitisation and analgesia are associated with better outcomes, although there is obviously a need to firstly positively identify it. Despite the fact that this study does not adequately explain how absent SEPs may encourage the development of CRPS, it does remind the clinician of the importance of maintaining a high index of suspicion for this syndrome, particularly in patients where a stroke produces sensory impairment in a distal limb.

Han EY, Jung HY, Kim MO. Absent Median Somatosensory Evoked Potential is a Predictor of Type 1 Complex Regional Pain Syndrome. Disability and rehabilitation.



ACNR 2014: 14(4). Published online 29/9/14