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Genetic pain loss includes congenital insensitivity to pain (CIP), hereditary sensory neuropathies and, if autonomic nerves are involved, hereditary sensory and autonomic neuropathy (HSAN). This heterogeneous group of disorders highlights the essential role of nociception in protecting against tissue damage. Patients with genetic pain loss have recurrent injuries, burns and poorly healing wounds as disease hallmarks. CIP and HSAN are caused by pathogenic genetic variants in >20 genes that lead to developmental defects, neurodegeneration or altered neuronal excitability of peripheral damage-sensing neurons. These genetic variants lead to hyperactivity of sodium channels, disturbed haem metabolism, altered clathrin-mediated transport and impaired gene regulatory mechanisms affecting epigenetic marks, long non-coding RNAs and repetitive elements. Therapies for pain loss disorders are mainly symptomatic but the first targeted therapies are being tested. Conversely, chronic pain remains one of the greatest unresolved medical challenges, and the genes and mechanisms associated with pain loss offer new targets for analgesics. Given the progress that has been made, the coming years are promising both in terms of targeted treatments for pain loss disorders and the development of innovative pain medicines based on knowledge of these genetic diseases.

Peripheral sensory neurons are afferent neurons that innervate the skin, joints, bones, muscles, and mucosal tissues. By converting different stimuli into action potentials, they transmit signals for the sensing of temperature, touch, pressure, or pain. This review discusses the known Mendelian disorders which affect pain sensing in humans. For painlessness, these disorders can be classified as developmental, neurodegenerative, or functional, where pain-sensing neurons (nociceptors) are present but cannot be activated or produce action potentials. Affected patients suffer from numbness with recurrent injuries, burns, and poorly healing wounds. For Mendelian disorders of excess pain, aberrant overactivity of nociceptors is a hallmark and leads to paroxysmal or continuous pain states. Again, the effect can be functional or, as in small fiber neuropathies, can be accompanied by degeneration of small unmyelinated nerve fibers in the skin. About 20 different genes are known to cause Mendelian pain disorders and the molecules involved are of general interest for human pain research and as analgesic targets. Comprehensive genetic testing is the key to early diagnosis and adaptation of clinical management.

Summary By studying healthy women who do not request analgesia during their first delivery, we investigate genetic effects on labor pain. Such women have normal sensory and psychometric test results, except for significantly higher cuff pressure pain. We find an excess of heterozygotes carrying the rare allele of SNP rs140124801 in KCNG4. The rare variant KV6.4-Met419 has a dominant-negative effect and cannot modulate the voltage dependence of KV2.1 inactivation because it fails to traffic to the plasma membrane. In vivo, Kcng4 (KV6.4) expression occurs in 40% of retrograde-labeled mouse uterine sensory neurons, all of which express KV2.1, and over 90% express the nociceptor genes Trpv1 and Scn10a. In neurons overexpressing KV6.4-Met419, the voltage dependence of inactivation for KV2.1 is more depolarized compared with neurons overexpressing KV6.4. Finally, KV6.4-Met419-overexpressing neurons have a higher action potential threshold. We conclude that KV6.4 can influence human labor pain by modulating the excitability of uterine nociceptors., Graphical Abstract, Highlights • KCNG4 variant highly prevalent in women requiring no analgesia in childbirth • KCNG4 variant encodes KV6.4Met-419; KV6.4 is a silent subunit modifying KV activity • KV6.4Met-419 is retained in the cytoplasm and acts in a dominant-negative manner • KV6.4Met-419 overexpression results in hypoexcitable sensory neurons, Lee et al. find that the KV6.4 variant KV6.4Met-419 is highly prevalent in women requiring no analgesia in childbirth and that KV6.4Met-419 is retained in the cytoplasm, preventing it from modifying KV2.1 channel activity, which results in hypoexcitable sensory neurons.

In human, many cases of congenital insensitivity to pain (CIP) are caused by mutations of components of the NGF/TrkA signaling pathway, required for survival and specification of nociceptors, and which plays a major role in pain processing. Mutations in PRDM12 have recently been identified in CIP patients indicating a putative role for this epigenetic modifier in pain sensing. Here, we show that Prdm12 expression is restricted to developing and adult nociceptors and that its genetic ablation compromises their viability and maturation. Mechanistically, we found that Prdm12 is required for the initiation and maintenance of the expression of TrkA, by acting as a modulator of Neurogenin1/2 transcription factor activity, both in frogs, mice and humans. Together, our results identify Prdm12 as an evolutionarily conserved epigenetic regulator of nociceptor specification, and as an actionable target for new pain therapeutics., SCOPUS: ar.j, info:eu-repo/semantics/published

Inherited pain disorders are typically rare in the general population. However, in the postgenomic era, single-gene mutations for numerous human Mendelian pain disorders have been described owing to advances in sequencing technology and improvements in pain phenotyping. This article describes the history, phenotype, gene mutations, and molecular/cellular pathology of painless and painful inherited monogenic disorders. The study of these disorders has led to the identification of key genes that are needed for the normal development or function of nociceptive neurons. Genes that are covered include ATL1, ATL3, DNMT1, DST, ELP1, FLVCR1, KIF1A, NGF, NTRK1, PRDM12, RETREG1, SCN9A, SCN10A, SCN11A, SPTLC1, SPTLC2, TRPA1, WNK1, and ZFHX2. The study of some Mendelian disorders of pain sensing has the potential to lead to new classes of analgesic drugs.

We sought genetic effects on labour pain by studying healthy women who did not request analgesia during their first delivery. Extensive sensory and psychometric testing were normal in these women, except for significantly higher cuff-pressure pain. We found an excess of heterozygotes carrying the rare allele of SNP rs140124801 in KCNG4. The rare variant KV6.4-Met419 exerts a dominant negative effect and cannot modulate the voltage-dependence of KV2.1 inactivation because it fails to traffic to the plasma membrane. In vivo, we observed Kcng4 (KV6.4) expression in 40% of retrograde labelled mouse uterine sensory neurones, all of which expressed KV2.1, and over 90% expressed nociceptor genes Trpv1 and Scn10a. In neurones overexpressing KV6.4-Met419, the voltage-dependence of inactivation for KV2.1 is more depolarised compared to neurones overexpressing KV6.4. Finally, KV6.4-Met419 overexpressing neurones have a higher action potential threshold. We conclude KV6.4 can influence human labour pain by modulating the excitability of uterine nociceptors.

BackgroundProgressive encephalopathy, hypsarrhythmia and optic atrophy (PEHO) has been described as a clinically distinct syndrome. It has been postulated that it is an autosomal recessive condition. However, the aetiology is poorly understood, and the genetic basis of the condition has not been fully elucidated. Our objective was to discover if PEHO syndrome is a single gene disorder.MethodChildren with PEHO and PEHO-like syndrome were recruited. Clinical, neurological and dysmorphic features were recorded; EEG reports and MRI scans were reviewed. Where possible, exome sequencing was carried out first to seek mutations in known early infantile developmental and epileptic encephalopathy (DEE) genes and then to use an agnostic approach to seek novel candidate genes. We sought intra–interfamilial phenotypic correlations and genotype–phenotype correlations when pathological mutations were identified.ResultsTwenty-three children were recruited from a diverse ethnic background, 19 of which were suitable for inclusion. They were similar in many of the core and the supporting features of PEHO, but there was significant variation in MRI and ophthalmological findings, even between siblings with the same mutation. A pathogenic genetic variant was identified in 15 of the 19 children. One further girl’s DNA failed analysis, but her two affected sisters shared confirmed variants. Pathogenic variants were identified in seven different genes.ConclusionsWe found significant clinical and genetic heterogeneity. Given the intrafamily variation demonstrated, we question whether the diagnostic criteria for MRI and ophthalmic findings should be altered. We also question whether PEHO and PEHO-like syndrome represent differing points on a clinical spectrum of the DEE. We conclude that PEHO and PEHO-like syndrome are clinically and genetically diverse entities—and are phenotypic endpoints of many severe genetic encephalopathies.

The importance of NaV1.7 (encoded bySCN9A) in the regulation of pain sensing is exemplified by the heterogeneity of clinical phenotypes associated with its mutation. Gain-of-function mutations are typically pain-causing and have been associated with inherited erythromelalgia (IEM) and paroxysmal extreme pain disorder (PEPD). IEM is usually caused by enhanced NaV1.7 channel activation, whereas mutations that alter steady-state fast inactivation often lead to PEPD. In contrast, nonfunctional mutations inSCN9Aare known to underlie congenital insensitivity to pain (CIP). Although well documented, the correlation betweenSCN9Agenotypes and clinical phenotypes is still unclear. Here we report three families with novelSCN9Amutations. In a multiaffected dominant family with IEM, we found the heterozygous change L245 V. Electrophysiological characterization showed that this mutation did not affect channel activation but instead resulted in incomplete fast inactivation and a small hyperpolarizing shift in steady-state slow inactivation, characteristics more commonly associated with PEPD. In two compound heterozygous CIP patients, we found mutations that still retained functionality of the channels, with two C-terminal mutations (W1775R and L1831X) exhibiting a depolarizing shift in channel activation. Two mutations (A1236E and L1831X) resulted in a hyperpolarizing shift in steady-state fast inactivation. To our knowledge, these are the first descriptions of mutations with some retained channel function causing CIP. This study emphasizes the complex genotype–phenotype correlations that exist forSCN9Aand highlights the C-terminal cytoplasmic region of NaV1.7 as a critical region for channel function, potentially facilitating analgesic drug development studies.

Autosomal recessive microcephaly or MicroCephaly Primary Hereditary (MCPH) is a genetically heterogeneous neurodevelopmental disorder characterized by a reduction in brain volume, indirectly measured by an occipitofrontal circumference (OFC) 2 standard deviations or more below the age- and sex-matched mean (-2SD) at birth and -3SD after 6 months, and leading to intellectual disability of variable severity. The Abnormal SPindle-like Microcephaly gene (ASPM), the human ortholog of the Drosophila melanogaster 'abnormal spindle' gene (asp), encodes ASPM, a protein localized at the centrosome of apical neuroprogenitor cells and involved in spindle pole positioning during neurogenesis. Loss-of-function mutations in ASPM cause MCPH5, which affects the majority of all MCPH patients worldwide. Here, we report 47 unpublished patients from 39 families carrying 28 new ASPM mutations, and conduct an exhaustive review of the molecular, clinical, neuroradiological and neuropsychological features of the 282 families previously reported (with 161 distinct ASPM mutations). Furthermore, we show that ASPM-related microcephaly is not systematically associated with intellectual deficiency and discuss the association between the structural brain defects (strong reduction in cortical volume and surface area) that modify the cortical map of these patients and their cognitive abilities. This article is protected by copyright. All rights reserved. ispartof: Human Mutation vol:39 issue:3 pages:319-332 ispartof: location:United States status: published

Inherited microcephaly exposes Zika culprit Microcephaly has been the terrifying hallmark of the recent outbreak of Zika virus (ZIKV) in the Americas. How the virus damages brain development in the fetus is enigmatic. Chavali et al. found that in congenital microcephaly, mutations in a neural precursor protein, Musashi-1 (MSI1), impede RNA binding to neural stem cell targets, resulting in abnormal brain development (see the Perspective by Griffin). MSI1 also binds ZIKV RNA to amplify viral replication in cells. This interaction could put a pregnant woman at risk of giving birth to a microcephalic child. Furthermore, MSI1 is expressed at high levels in the mouse testis, which may explain the sexual transmission of this virus. Science , this issue p. 83 ; see also p. 33

Contains fulltext : 138015.pdf (Publisher’s version ) (Open Access) Ligase IV syndrome is a rare differential diagnosis for Nijmegen breakage syndrome owing to a shared predisposition to lympho-reticular malignancies, significant microcephaly, and radiation hypersensitivity. Only 16 cases with mutations in LIG4 have been described to date with phenotypes varying from malignancy in developmentally normal individuals, to severe combined immunodeficiency and early mortality. Here, we report the identification of biallelic truncating LIG4 mutations in 11 patients with microcephalic primordial dwarfism presenting with restricted prenatal growth and extreme postnatal global growth failure (average OFC -10.1 s.d., height -5.1 s.d.). Subsequently, most patients developed thrombocytopenia and leucopenia later in childhood and many were found to have previously unrecognized immunodeficiency following molecular diagnosis. None have yet developed malignancy, though all patients tested had cellular radiosensitivity. A genotype-phenotype correlation was also noted with position of truncating mutations corresponding to disease severity. This work extends the phenotypic spectrum associated with LIG4 mutations, establishing that extreme growth retardation with microcephaly is a common presentation of bilallelic truncating mutations. Such growth failure is therefore sufficient to consider a diagnosis of LIG4 deficiency and early recognition of such cases is important as bone marrow failure, immunodeficiency, and sometimes malignancy are long term sequelae of this disorder.

Item does not contain fulltext Human studies have firmly implicated voltage-gated sodium channels in human pain disorders, and targeted and massively parallel genomic sequencing is beginning to be used in clinical practice to determine which sodium channel variants are involved. Missense substitutions of SCN9A, the gene encoding sodium channel NaV1.7, SCN10A, the gene encoding sodium channel NaV1.8, and SCN11A, the gene encoding sodium channel NaV1.9, produce gain-of-function changes that contribute to pain in many human painful disorders. Genomic sequencing might help to establish a diagnosis, and in the future might support individualisation of therapeutic approaches. However, in many cases, and especially in sodium channelopathies, the results from genomic sequencing can only be appropriately interpreted in the context of an extensive functional assessment, or family segregation analysis of phenotype and genotype.

Summary The discovery of genetic variants that substantially alter an individual's perception of pain has led to a step-change in our understanding of molecular events underlying the detection and transmission of noxious stimuli by the peripheral nervous system. For example, the voltage-gated sodium ion channel Na v 1.7 is expressed selectively in sensory and autonomic neurons; inactivating mutations in SCN9A , which encodes Na v 1.7, result in congenital insensitivity to pain, whereas gain-of-function mutations in this gene produce distinct pain syndromes such as inherited erythromelalgia, paroxysmal extreme pain disorder, and small-fibre neuropathy. Heterozygous mutations in TRPA1 , which encodes the transient receptor potential cation channel, can cause familial episodic pain syndromes, and variants of genes coding for the voltage-gated sodium channels Na v 1.8 ( SCN10A ) and Na v 1.9 ( SCN11A ) lead to small-fibre neuropathy and congenital insensitivity to pain, respectively. Furthermore, other genetic polymorphisms have been identified that contribute to risk or severity of more complex pain phenotypes. Novel models of sensory disorders are in development—eg, using human sensory neurons differentiated from human induced pluripotent stem cells. Understanding rare heritable pain disorders not only improves diagnosis and treatment of patients but may also reveal new targets for analgesic drug development.

Background Signaling through the T-cell receptor (TCR) is critical for T-cell development and function. Linker for activation of T cells (LAT) is a transmembrane adaptor signaling molecule that is part of the TCR complex and essential for T-cell development, as demonstrated by LAT-deficient mice, which show a complete lack of peripheral T cells. Objective We describe a pedigree affected by a severe combined immunodeficiency phenotype with absent T cells and normal B-cell and natural killer cell numbers. A novel homozygous frameshift mutation in the gene encoding for LAT was identified in this kindred. Methods Genetic, molecular, and functional analyses were used to identify and characterize the LAT defect. Clinical and immunologic analysis of patients was also performed and reported. Results Homozygosity mapping was used to identify potential defective genes. Sanger sequencing of the LAT gene showed a mutation that resulted in a premature stop codon and protein truncation leading to complete loss of function and loss of expression of LAT in the affected family members. We also demonstrate loss of LAT expression and lack of TCR signaling restoration in LAT-deficient cell lines reconstituted with a synthetic LAT gene bearing this severe combined immunodeficiency mutation. Conclusion For the first time, the results of this study show that inherited LAT deficiency should be considered in patients with combined immunodeficiency with T-cell abnormalities.

Autosomal recessive primary microcephaly (MCPH) is characterized by a substantial reduction in prenatal human brain growth without alteration of the cerebral architecture and is caused by biallelic mutations in genes coding for a subset of centrosomal proteins. Although at least three of these proteins have been implicated in centrosome duplication, the nature of the centrosome dysfunction that underlies the neurodevelopmental defect in MCPH is unclear. Here we report a homozygous MCPH-causing mutation in human CEP63. CEP63 forms a complex with another MCPH protein, CEP152, a conserved centrosome duplication factor. Together, these two proteins are essential for maintaining normal centrosome numbers in cells. Using super-resolution microscopy, we found that CEP63 and CEP152 co-localize in a discrete ring around the proximal end of the parental centriole, a pattern specifically disrupted in CEP63-deficient cells derived from patients with MCPH. This work suggests that the CEP152-CEP63 ring-like structure ensures normal neurodevelopment and that its impairment particularly affects human cerebral cortex growth.

3-hydroxyisobutyric aciduria is an organic aciduria with a poorly understood biochemical basis. It has previously been assumed that deficiency of 3-hydroxyisobutyrate dehydrogenase (HIBADH) in the valine catabolic pathway is the underlying enzyme defect, but more recent evidence makes it likely that individuals with 3-hydroxyisobutyryic aciduria represent a heterogeneous group with different underlying mechanisms, including respiratory chain defects or deficiency of methylmalonate semialdehyde dehydrogenase. However, to date methylmalonate semialdehyde dehydrogenase deficiency has only been demonstrated at the gene level for a single individual. We present two unrelated patients who presented with developmental delay and increased urinary concentrations of 3-hydroxyisobutyric acid. Both children were products of consanguineous unions and were of European or Pakistani descent. One patient developed a febrile illness and subsequently died from a hepatoencephalopathy at 2 years of age. Further studies were initiated and included tests of the HIBADH enzyme in fibroblast homogenates, which yielded normal activities. Sequencing of the ALDH6A1 gene (encoding methylmalonate semialdehyde dehydrogenase) suggested homozygosity for the missense mutation c.785 C > A (S262Y) in exon 7 which was not found in 210 control alleles. Mutation analysis of the ALDH6A1 gene of the second patient confirmed the presence of a different missense mutation, c.184 C > T (P62S), which was also identified in 1/530 control chromosomes. Both mutations affect highly evolutionarily conserved amino acids of the methylmalonate semialdehyde dehydrogenase protein. Mutation analysis in the ALDH6A1 gene can reveal a cause of 3-hydroxyisobutyric aciduria, which may present with only slightly increased urinary levels of 3-hydroxyisobutyric acid, if a patient is metabolically stable.

Autosomal recessive primary microcephaly (MCPH) is a disorder of neurodevelopment resulting in a small brain1,2. We identified WDR62 as the second most common cause of MCPH after finding homozygous missense and frame-shifting mutations in seven MCPH families. In human cell lines, we found that WDR62 is a spindle pole protein, as are ASPM and STIL, the MCPH7 and MCHP7 proteins3–5. Mutant WDR62 proteins failed to localize to the mitotic spindle pole. In human and mouse embryonic brain, we found that WDR62 expression was restricted to neural precursors undergoing mitosis. These data lend support to the hypothesis that the exquisite control of the cleavage furrow orientation in mammalian neural precursor cell mitosis, controlled in great part by the centrosomes and spindle poles, is critical both in causing MCPH when perturbed and, when modulated, generating the evolutionarily enlarged human brain6–9.

Summary Human monogenic pain syndromes have provided important insights into the molecular mechanisms that underlie normal and pathological pain states. We describe an autosomal-dominant familial episodic pain syndrome characterized by episodes of debilitating upper body pain, triggered by fasting and physical stress. Linkage and haplotype analysis mapped this phenotype to a 25 cM region on chromosome 8q12–8q13. Candidate gene sequencing identified a point mutation (N855S) in the S4 transmembrane segment of TRPA1, a key sensor for environmental irritants. The mutant channel showed a normal pharmacological profile but altered biophysical properties, with a 5-fold increase in inward current on activation at normal resting potentials. Quantitative sensory testing demonstrated normal baseline sensory thresholds but an enhanced secondary hyperalgesia to punctate stimuli on treatment with mustard oil. TRPA1 antagonists inhibit the mutant channel, promising a useful therapy for this disorder. Our findings provide evidence that variation in the TRPA1 gene can alter pain perception in humans. Video Abstract, Highlights ► We describe a novel human autosomal-dominant pain syndrome (FEPS) ► We demonstrate linkage of FEPS to a TRPA1 N855S point mutation ► FEPS patients show debilitating pain on fasting and physical stress ► N855S TRPA1 channels show enhanced inward currents but unaltered pharmacology

Item does not contain fulltext Dominant gain-of-function mutations that hyperpolarize activation of the Na(v)1.7 sodium channel have been linked to inherited erythromelalgia (IEM), a disorder characterized by severe pain and redness in the feet and hands in response to mild warmth. Pharmacotherapy remains largely ineffective for IEM patients with cooling and avoidance of triggers being the most reliable methods to relieve pain. We now report a 5 year old patient with pain precipitated by warmth, together with redness in her hands and feet. Her pain episodes were first reported at 12 months, and by the age of 15-16 months were triggered by sitting as well as heat. Pain has been severe, inducing self-mutilation, with limited relief from drug treatment. Our analysis of the patient's genomic DNA identified a novel Na(v)1.7 mutation which replaces isoleucine 234 by threonine (I234T) within domain I/S4-S5 linker. Whole-cell voltage-clamp analysis shows a I234T-induced shift of -18 mV in the voltage-dependence of activation, accelerated time-to-peak, slowed deactivation and enhanced responses to slow ramp depolarizations, together with a -21 mV shift in the voltage-dependence of slow-inactivation. Our data show that I234T induces the largest activation shift for Na(v)1.7 mutations reported thus far. Although enhanced slow-inactivation may attenuate the gain-of-function of the I234T mutation, the shift in activation appears to be dominant, and is consistent with the severe pain symptoms reported in this patient. 01 oktober 2010

P rogressive E ncephalopathy with oedema, H ypsarrhythmia and O ptic atrophy (PEHO) is a rare, neurodegenerative disorder of unknown aetiology. Nahorski et al . identify the first causative recessive mutation in CCDC88A , which encodes the actin-binding protein Girdin. The phenotype and brain anatomy of the Ccdc88a knockout mouse resemble those of human PEHO syndrome., Progressive encephalopathy with oedema, hypsarrhythmia and optic atrophy (PEHO) syndrome is a rare Mendelian phenotype comprising severe retardation, early onset epileptic seizures, optic nerve/cerebellar atrophy, pedal oedema, and early death. Atypical cases are often known as PEHO-like, and there is an overlap with ‘early infantile epileptic encephalopathy’. PEHO is considered to be recessive, but surprisingly since initial description in 1991, no causative recessive gene(s) have been described. Hence, we report a multiplex consanguineous family with the PEHO phenotype where affected individuals had a homozygous frame-shift deletion in CCDC88A (c.2313delT, p.Leu772*ter). Analysis of cDNA extracted from patient lymphocytes unexpectedly failed to show non-sense mediated decay, and we demonstrate that the mutation produces a truncated protein lacking the crucial C-terminal half of CCDC88A (girdin). To further investigate the possible role of CCDC88A in human neurodevelopment we re-examined the behaviour and neuroanatomy of Ccdc88a knockout pups. These mice had mesial-temporal lobe epilepsy, microcephaly and corpus callosum deficiency, and by postnatal Day 21, microcephaly; the mice died at an early age. As the mouse knockout phenotype mimics the human PEHO phenotype this suggests that loss of CCDC88A is a cause of the PEHO phenotype, and that CCDC88A is essential for multiple aspects of normal human neurodevelopment.

Numerical centrosome aberrations underlie certain developmental abnormalities and may promote cancer. A cell maintains normal centrosome numbers by coupling centrosome duplication with segregation, which is achieved through sustained association of each centrosome with a mitotic spindle pole. Although the microcephaly- and primordial dwarfism-linked centrosomal protein CEP215 has been implicated in this process, the molecular mechanism responsible remains unclear. Here, using proteomic profiling, we identify the minus end-directed microtubule motor protein HSET as a direct binding partner of CEP215. Targeted deletion of the HSET-binding domain of CEP215 in vertebrate cells causes centrosome detachment and results in HSET depletion at centrosomes, a phenotype also observed in CEP215-deficient patient-derived cells. Moreover, in cancer cells with centrosome amplification, the CEP215–HSET complex promotes the clustering of extra centrosomes into pseudo-bipolar spindles, thereby ensuring viable cell division. Therefore, stabilization of the centrosome–spindle pole interface by the CEP215–HSET complex could promote survival of cancer cells containing supernumerary centrosomes., Centrosome clustering allows survival of cells with amplified centrosomes at the cost of chromosome instability. Here, Chavali et al. show that the centrosome component CEP215 collaborates with the kinesin motor HSET both to maintain spindle poles connections and to cluster centrosomes.

Erroneous activation of the pain-sensing system, as in chronic or neuropathic pain, represents a major health burden with insufficient treatment options. However, the study of genetic disorders rendering individuals completely unable to feel pain offers hope. All causes of congenital painlessness affect nociceptors, evolutionarily conserved specialist neurons able to sense all type of tissue damage. The discovery of new genes essential for sensing pain (SCN11A, PRDM12, and CLTCL1) has provided unexpected insights into the biological mechanisms that drive distinct stages of nociception. Drugs targeting two previously discovered painlessness genes, NGF and SCN9A, are currently in late-stage clinical trials; thus, characterization of these new painlessness genes has significant potential for the generation of new classes of analgesics.

International audience; The Na v 1.7 voltage-gated sodium channel, encoded by SCN9A, is critical for human pain perception yet the transcriptional and post-transcriptional mechanisms that regulate this gene are still incompletely understood. Here, we describe a novel natural antisense transcript (NAT) for SCN9A that is conserved in humans and mice. The NAT has a similar tissue expression pattern to the sense gene and is alternatively spliced within dorsal root ganglia. The human and mouse NATs exist in cis with the sense gene in a tail-to-tail orientation and both share sequences that are complementary to the terminal exon of SCN9A/Scn9a. Overexpression analyses of the human NAT in human embryonic kidney (HEK293A) and human neuroblastoma (SH-SY5Y) cell lines show that it can function to downregulate Na v 1.7 mRNA, protein levels and currents. The NAT may play an important role in regulating human pain thresholds and is a potential candidate gene for individuals with chronic pain disorders that map to the SCN9A locus, such as Inherited Primary Erythromelalgia, Paroxysmal Extreme Pain Disorder and Painful Small Fibre Neuropathy, but who do not contain mutations in the sense gene. Our results strongly suggest the SCN9A NAT as a prime candidate for new therapies based upon augmentation of existing antisense RNAs in the treatment of chronic pain conditions in man.

Pain perception has evolved as a warning mechanism to alert organisms to tissue damage and dangerous environments. In humans, however, undesirable, excessive or chronic pain is a common and major societal burden for which available medical treatments are currently suboptimal. New therapeutic options have recently been derived from studies of individuals with congenital insensitivity to pain (CIP). Here we identified 10 different homozygous mutations in PRDM12 (encoding PRDI-BF1 and RIZ homology domain-containing protein 12) in subjects with CIP from 11 families. Prdm proteins are a family of epigenetic regulators that control neural specification and neurogenesis. We determined that Prdm12 is expressed in nociceptors and their progenitors and participates in the development of sensory neurons in Xenopus embryos. Moreover, CIP-associated mutants abrogate the histone-modifying potential associated with wild-type Prdm12. Prdm12 emerges as a key factor in the orchestration of sensory neurogenesis and may hold promise as a target for new pain therapeutics. © 2015 Nature America, Inc. All rights reserved.

Individuals born of consanguineous union have segments of their genomes that are homozygous as a result of inheriting identical ancestral genomic segments through both parents. One consequence of this is an increased incidence of recessive disease within these sibships. Theoretical calculations predict that 6% (1/16) of the genome of a child of first cousins will be homozygous and that the average homozygous segment will be 20 cM in size. We assessed whether these predictions held true in populations that have preferred consanguineous marriage for many generations. We found that in individuals with a recessive disease whose parents were first cousins, on average, 11% of their genomes were homozygous (n = 38; range 5%-20%), with each individual bearing 20 homozygous segments exceeding 3 cM (n = 38; range of number of homozygous segments 7-32), and that the size of the homozygous segment associated with recessive disease was 26 cM (n = 100; range 5-70 cM). These data imply that prolonged parental inbreeding has led to a background level of homozygosity increased approximately 5% over and above that predicted by simple models of consanguinity. This has important clinical and research implications.

One of the most notable trends in human evolution is the dramatic increase in brain size that has occurred in the great ape clade, culminating in humans. Of particular interest is the vast expanse of the cerebral cortex, which is believed to have resulted in our ability to perform higher cognitive functions. Recent investigations of congenital microcephaly in humans have resulted in the identification of several genes that non-redundantly and specifically influence mammalian brain size. These genes appear to affect neural progenitor cell number through microtubular organisation at the centrosome.

Meckel-Gruber syndrome is a severe autosomal, recessively inherited disorder characterized by bilateral renal cystic dysplasia, developmental defects of the central nervous system (most commonly occipital encephalocele), hepatic ductal dysplasia and cysts and polydactyly1,2,3. MKS is genetically heterogeneous, with three loci mapped: MKS1, 17q21-24 (ref. 4); MKS2, 11q13 (ref. 5) and MKS3 (ref. 6). We have refined MKS3 mapping to a 12.67-Mb interval (8q21.13-q22.1) that is syntenic to the Wpk locus in rat, which is a model with polycystic kidney disease, agenesis of the corpus callosum and hydrocephalus7,8. Positional cloning of the Wpk gene suggested a MKS3 candidate gene, TMEM67, for which we identified pathogenic mutations for five MKS3-linked consanguineous families. MKS3 is a previously uncharacterized, evolutionarily conserved gene that is expressed at moderate levels in fetal brain, liver and kidney but has widespread, low levels of expression. It encodes a 995–amino acid seven-transmembrane receptor protein of unknown function that we have called meckelin.

Autosomal recessive primary microcephaly (MCPH) is a neurodevelopmental disorder. It is characterized by two principal features, microcephaly present at birth and nonprogressive mental retardation. The microcephaly is the consequence of a small but architecturally normal brain, and it is the cerebral cortex that shows the greatest size reduction. There are at least seven MCPH loci, and four of the genes have been identified: MCPH1, encoding Microcephalin; MCPH3, encoding CDK5RAP2; MCPH5, encoding ASPM; and MCPH6, encoding CENPJ. These findings are starting to have an impact on the clinical management of families affected with MCPH. Present data suggest that MCPH is the consequence of deficient neurogenesis within the neurogenic epithelium. Evolutionary interest in MCPH has been sparked by the suggestion that changes in the MCPH genes might also be responsible for the increase in brain size during human evolution. Indeed, evolutionary analyses of Microcephalin and ASPM reveal evidence for positive selection during human and great ape evolution. So an understanding of this rare genetic disorder may offer us significant insights into neurogenic mitosis and the evolution of the most striking differences between us and our closest living relatives: brain size and cognitive ability.

Seckel syndrome (OMIM 210600) is an autosomal recessive disorder characterized by intrauterine growth retardation, dwarfism, microcephaly and mental retardation. Clinically, Seckel syndrome shares features in common with disorders involving impaired DNA-damage responses, such as Nijmegen breakage syndrome (OMIM 251260) and LIG4 syndrome (OMIM 606593). We previously mapped a locus associated with Seckel syndrome to chromosome 3q22.1–q24 in two consanguineous Pakistani families1. Further marker analysis in the families, including a recently born unaffected child with a recombination in the critical region, narrowed the region to an interval of 5 Mbp between markers D3S1316 and D3S1557 (145.29 Mbp and 150.37 Mbp). The gene encoding ataxia–telangiectasia and Rad3–related protein (ATR) maps to this region2,3. A fibroblast cell line derived from an affected individual displays a defective DNA damage response caused by impaired ATR function. We identified a synonymous mutation in affected individuals that alters ATR splicing. The mutation confers a phenotype including marked microcephaly (head circumference 12 s.d. below the mean) and dwarfism (5 s.d. below the mean). Our analysis shows that UV-induced ATR activation can occur in non-replicating cells following processing by nucleotide excision repair.

Mutations in the ASPM gene at the MCPH5 locus are expected to be the most common cause of human autosomal recessive primary microcephaly (MCPH), a condition in which there is a failure of normal fetal brain development, resulting in congenital microcephaly and mental retardation. We have performed the first comprehensive mutation screen of the 10.4-kb ASPM gene, identifying all 19 mutations in a cohort of 23 consanguineous families. Mutations occurred throughout the ASPM gene and were all predicted to be protein truncating. Phenotypic variation in the 51 affected individuals occurred in the degree of microcephaly (5-11 SDs below normal) and of mental retardation (mild to severe) but appeared independent of mutation position.

One of the most notable trends in mammalian evolution is the massive increase in size of the cerebral cortex, especially in primates. Humans with autosomal recessive primary microcephaly (MCPH) show a small but otherwise grossly normal cerebral cortex associated with mild to moderate mental retardation. Genes linked to this condition offer potential insights into the development and evolution of the cerebral cortex. Here we show that the most common cause of MCPH is homozygous mutation of ASPM, the human ortholog of the Drosophila melanogaster abnormal spindle gene (asp), which is essential for normal mitotic spindle function in embryonic neuroblasts. The mouse gene Aspm is expressed specifically in the primary sites of prenatal cerebral cortical neurogenesis. Notably, the predicted ASPM proteins encode systematically larger numbers of repeated 'IQ' domains between flies, mice and humans, with the predominant difference between Aspm and ASPM being a single large insertion coding for IQ domains. Our results and evolutionary considerations suggest that brain size is controlled in part through modulation of mitotic spindle activity in neuronal progenitor cells.

Pain is a sense that has two purposes in humans: first, to warn of impending tissue damage and detect actual tissue damage – a process known as nociception; and second to guide childhood and adolescent development by setting upper limits to activity. A complete absence of pain sensing is found in a small number of Mendelian disorders. These disorders of painlessness can be categorised into three categories: defects in peripheral nervous system development, disorders in pain sensing and progressive nociceptor-specific neuropathies. The study of these conditions has helped to sculpt our understanding of the human pain sensing systems and provided insights into the uniqueness of nociceptive neurons, with each supporting an axon/dendrite of up to 50 cm in length (which lacks a myelin sheath) and lasting a lifetime. Furthermore, these painlessness genes are starting to provide new therapeutic avenues for pain control, for example, NGF and NaV1.7 antagonists. Key Concepts: Pain is the only one of the senses ubiquitously present in all multicellular organisms. Pain has roles in both normal development and also in the prolongation of each individual through injury avoidance and management. Mendelian disorders of painlessness exist and are individually very rare. The study of Mendelian disorders of painlessness has led to significant advances in our understanding of pain processing and defined a subset of genes/proteins, which are nonredundantly necessary for the generation, function and survival of nociceptors. Biallelic mutations in either NTRK1 or NGF lead to a failure of nociception system development and congenital inability to perceive pain. Mutations in the voltage-gated sodium channels SCN9A (presumably SCN10A) and SCN11A cause defective integrative pain sensing and congenital inability to perceive pain. Degenerative disorders of nociception neurons are associated with mutations in SPTLC1, SPTLC2, WNK1, FAM134B, IKBKAP and DST and generally lead to a progressive loss of pain sensing. New analgesic treatments are being generated from the study of Mendelian disorders of painlessness. Keywords: nociception; pain insensitivity; Mendelian disorders of painlessness; pain sensing; hereditary sensory and autonomic neuropathy (HSAN)

Primary autosomal recessive microcephaly is a clinical diagnosis of exclusion in an individual with a head circumference/=4 SDs below the expected age-and-sex mean. There is associated moderate mental retardation, and neuroimaging shows a small but structurally normal cerebral cortex. The inheritance pattern in the majority of cases is considered to be autosomal recessive. Although genetic heterogeneity for this clinical phenotype had been expected, this has only recently been demonstrated, with the mapping of two loci for autosomal recessive primary microcephaly: MCPH1 at 8p and MCPH2 at 19q. We have studied a large multiaffected consanguineous pedigree, using a whole-genome search, and have identified a third locus, MCPH3 at 9q34. The minimal critical region is approximately 12 cM, being defined by the markers cen-D9S1872-D9S159-tel, with a maximum two-point LOD score of 3.76 (recombination fraction 0) observed for the marker D9S290.

SummaryPrimary (or “true”) microcephaly is inherited as an autosomal recessive trait and is thought to be genetically heterogeneous. Using autozygosity mapping, we have identified a genetic locus (MCPH1) for primary microcephaly, at chromosome 8p22-pter, in two consanguineous families of Pakistani origin. Our results indicate that the gene lies within a 13-cM region between the markers D8S1824 and D8S1825 (maximum multipoint LOD score of 8.1 at D8S277). In addition, we have demonstrated the genetic heterogeneity of this condition by analyzing a total of nine consanguineous families with primary microcephaly.

Neurodegenerative disorders with high brain iron include Parkinson disease, Alzheimer disease and several childhood genetic disorders categorized as neuroaxonal dystrophies. We mapped a locus for infantile neuroaxonal dystrophy ( INAD) and neurodegeneration with brain iron accumulation (NBIA) to chromosome 22q12-q13 and identified mutations in PLA2G6, encoding a calcium-independent group VI phospholipase A(2), in NBIA, INAD and the related Karak syndrome. This discovery implicates phospholipases in the pathogenesis of neurodegenerative disorders with iron dyshomeostasis. United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)-R01HD050832 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Center for Research Resources (NCRR)-M01RR000334 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Eye Institute (NEI)-R01EY012353 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Human Genome Research Institute (NHGRI)-N01HG065403 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Institute on Aging (NIA)-P30AG008017 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Center for Research Resources (NCRR)-M01 RR000334 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Eye Institute (NEI)-R01 EY012353-07, R01 EY012353 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Human Genome Research Institute (NHGRI)-N01HG65403 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Institute on Aging (NIA)-P30 AG008017 United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)-R01 HD050832-01A1, R01 HD050832 Fondazione Telethon-GTF04002

Two mutation detection methods based on the cleavage of mismatched heteroduplexes were compared and evaluated. These techniques, chemical cleavage of mismatch (CCM) and enzyme mismatch cleavage (EMC), have the advantages over other available methods of being able to detect and localize mutations in relatively large fragments of DNA (or = 1 kb). We have constructed clones that enable us to create heteroduplexes of 500 bp, 1 kb, and 1.5 kb and have assessed each of the methods over a range of criteria. Both were able to detect and localize all four types of single-base mismatch and insertion/deletions of 1-5 bp. Whereas EMC was efficient at detection of insertion/deletions in a broad size range of fragments and has the advantage over CCM of using no hazardous chemicals, in our hands it has not been sufficiently robust that we felt confident to consider it for diagnostic use in its present form. CCM using hydroxylamine was efficient over the entire range of fragment sizes tested and using potassium permanganate with tetraethylammonium chloride was efficient up to 1 kb.

Autosomal recessive primary microcephaly is a potential model in which to research genes involved in human brain growth. We show that two forms of the disorder result from homozygous mutations in the genes CDK5RAP2 and CENPJ. We found neuroepithelial expression of the genes during prenatal neurogenesis and protein localization to the spindle poles of mitotic cells, suggesting that a centrosomal mechanism controls neuron number in the developing mammalian brain.

Genes encoding the α subunits of neuronal sodium channels have evolutionarily conserved sites of alternative splicing but no functional differences have been attributed to the splice variants. Here, using Na(V)1.7 as an exemplar, we show that the sodium channel isoforms are functionally distinct when co-expressed with β subunits. The gene, SCN9A, encodes the α subunit of the Na(V)1.7 channel, and contains both sites of alternative splicing that are highly conserved. In conditions where the intrinsic properties of the Na(V)1.7 splice variants were similar when expressed alone, co-expression of β1 subunits had different effects on channel availability that were determined by splicing at either site in the α subunit. While the identity of exon 5 determined the degree to which β1 subunits altered voltage-dependence of activation (P = 0.027), the length of exon 11 regulated how far β1 subunits depolarised voltage-dependence of inactivation (P = 0.00012). The results could have a significant impact on channel availability, for example with the long version of exon 11, the co-expression of β1 subunits could lead to nearly twice as large an increase in channel availability compared to channels containing the short version. Our data suggest that splicing can change the way that Na(V) channels interact with β subunits. Because splicing is conserved, its unexpected role in regulating the functional impact of β subunits may apply to multiple voltage-gated sodium channels, and the full repertoire of β subunit function may depend on splicing in α subunits.

Distinguishing Ciliopathy Cilia were once thought to be evolutionary remnants, but structural defects reveal their importance in signaling pathways and human disease, such as Joubert syndrome. Either of the genes TMEM138 and TMEM216 can be found mutated in phenotypically indistinguishable ciliopathy patients. Interestingly, despite their lack of sequence homology, these genes have always been aligned in head-to-tail configuration during vertebrate evolution. The proteins expressed by these genes mark distinct tethered vesicles, which differentially carry ciliary proteins for assembly. Lee et al. (p. 966 , published online 26 January; see the Perspective by Chakravarti and Kapoor ) show that the coordinated expression of these adjacent genes depends upon a coevolved regulatory element in the noncoding intergenic region, which thus integrates the roles of both gene products. This discovery explains not only the indistinguishable pathogenesis of the patients' genotypes but also how the evolutionary clustering of genes unrelated in sequence may correlate with coordinated control of expression and function.

We investigated three families whose offspring had extreme microcephaly at birth and profound mental retardation. Brain scans and postmortem data showed that affected individuals had brains less than 10\% of expected size (T (p.Ala29GlnfsX114) in a Turkish family and c.684\_685del (p.Pro229TrpfsX85) in two families of Pakistani origin. Using patient cells, we found that c.83+1G>T led to the use of a novel splice site and to a frameshift after NDE1 exon 2. Transfection of tagged NDE1 constructs showed that the c.684\_685del mutation resulted in a NDE1 that was unable to localize to the centrosome. By staining a patient-derived cell line that carried the c.83+1G>T mutation, we found that this endogeneously expressed mutated protein equally failed to localize to the centrosome. By examining human and mouse embryonic brains, we determined that NDE1 is highly expressed in neuroepithelial cells of the developing cerebral cortex, particularly at the centrosome. We show that NDE1 accumulates on the mitotic spindle of apical neural precursors in early neurogenesis. Thus, NDE1 deficiency causes both a severe failure of neurogenesis and a deficiency in cortical lamination. Our data further highlight the importance of the centrosome in multiple aspects of neurodevelopment.