Luigi Citrigno

Mutation analysis of the SPG4 gene in Italian patients with pure and complicated forms of spastic paraplegia

Mutations in the SPG4 gene are the most common causes of hereditary spastic paraplegia (HSP) accounting for up to 40% of autosomal dominant (AD) forms and 12-18% of sporadic cases. The phenotype associated with HSP due to mutations in the SPG4 gene tends to be pure. There is increasing evidence, however, of patients with complicated forms of spastic paraplegia in which SPG4 mutations were identified. A cohort of 38 unrelated Italian patients with spastic paraplegia, of which 24 had a clear dominant inheritance and 14 were apparently sporadic, were screened for mutations in the SPG4 gene. We identified 11 different mutations, six of which were novel (p.Glu143GlyfsX8, p.Tyr415X, p.Asp548Asn, c.1656_1664delinsTGACCT, c.1688-3C>G and c.*2G>T) and two exon deletions previously reported. The overall rate of SPG4 gene mutation in our patients was 36.8% (14/38); in AD-HSP we observed a mutation frequency of 45.8% (11/24), in sporadic cases the frequency was 21.4% (3/14). Furthermore, we found a mutational rate of 22.2% (2/9) and 41.4% (12/29) in the complicated and pure forms, respectively. The results underlie the importance of genetic testing in all affected individuals.

CADASIL: Extended polymorphisms and mutational analysis of the NOTCH3 gene

CADASIL is a cerebrovascular disease caused by mutations in the NOTCH3 gene. Most mutations result in a gain or loss of cysteine residue in one of the 34 epidermal growth factor‐like repeats in the extracellular domain of the Notch3 protein, thus sparing the number of cysteine residues. To date, more than 130 different mutations in the NOTCH3 gene have been reported in CADASIL patients, of which 95% are missense point mutations. Many polymorphisms have also been identified in the NOTCH3 coding sequence, some of them leading to amino acid substitutions. The aim of the present study was to analyze the NOTCH3 gene in a large group of patients affected by leukoencephalopathy and to investigate the presence of genetic variants. The molecular analysis revealed several nucleotide alterations. In particular, we identified 20 different mutations, 22 polymorphisms, and 8 genetic variants of unknown pathological significance never reported previously. We hope that this NOTCH3 gene mutational analysis, performed in such a significant number of unrelated and related patients affected by leukoencephalopathy, will help in molecular screening for the NOTCH3 gene, thus contributing to enlargement of the NOTCH3 gene variation database.

A novel mutation in the X‐linked cyclin‐dependent kinase‐like 5 (CDKL5) gene associated with a severe Rett phenotype

Mutations in the X‐linked cyclin‐dependent kinase‐like 5 (CDKL5) gene have recently been reported in patients with severe neurodevelopmental disorder characterized by early‐onset seizures, infantile spasms, severe psychomotor impairment and very recently, in patients with Rett syndrome (RTT)‐like phenotype. Although the involvement of CDKL5 in specific biological pathways and its neurodevelopmental role have not been completely elucidated, the CDKL5 appears to be physiologically related to the MECP2 gene. Here we report on the clinical and CDKL5 molecular investigation in a very unusual RTT case, with severe, early‐neurological involvement in which we have shown in a previous report, a novel P388S MECP2 mutation [Conforti et al. (2003); Am J Med Genet A 117A: 184–187]. The patient has had severe psychomotor delay since the first month of life and infantile spasms since age 5 months. Moreover, at age 5 years the patient suddenly presented with renal failure. The severe pattern of symptoms in our patient, similar to a CDKL5 phenotype, prompted us to perform an analysis of the CDKL5, which revealed a novel missense mutation never previously described. The X‐inactivation assay was non‐informative. In conclusion, this report reinforces the observation that the CDKL5 phenotype overlaps with RTT and that CDKL5 analysis is recommended in patients with a seizure disorder commencing during the first months of life.

TARDBP gene mutations in south Italian patients with amyotrophic lateral sclerosis

TAR-DNA-binding protein 43 (TDP-43) has recently been identified as the major pathological protein in abnormal inclusions in neurons and glial cells in sporadic amyotrophic lateral sclerosis (SALS) and SOD1 negative familial cases with amyotrophic lateral sclerosis (FALS). TDP-43 is evolutionarily conserved, consisting of two RNA recognition motifs and a glycine-rich C-terminal domain. It is involved in the regulation of expression and splicing, and in other cellular processes such as microRNA biogenesis, apoptosis and cell division.1 2 Starting in early 2008, dominant mutations in TARDBP gene have been reported by several groups as a primary cause of ALS. To date, a total of 30 mutations of TARDBP have been reported not only in SOD1 -negative FALS cases (∼3%) but also in SALS cases (∼1.5%). All but one of the mutations identified (D169G) reside in exon 6 of the TARDBP gene, which encodes for the C-terminal glycine-rich domain of TDP-43. All of these mutations are dominantly inherited missense changes with the exception of a truncating mutation (Y374X) at the extreme C terminus of the protein.3 In this study, in order to investigate the presence and frequency of TARDBP mutations in a cohort of 310 south Italian patients affected by ALS, we performed a mutational screening of the exon 4 and exon 6 of the gene in SOD1 -negative FALS and SALS patients. …

Spastic paraplegia with thinning of the corpus callosum and white matter abnormalities: Further mutations and relative frequency in ZFYVE26/SPG15 in the Italian population

Spastic paraplegia with thinning of the corpus callosum (ARHSP-TCC) is a relatively frequent form of complicated hereditary spastic paraplegia in which mental retardation and muscle stiffness at onset are followed by slowly progressive paraparesis and cognitive deterioration. Although genetically heterogeneous, ARHSP-TCC is frequently associated with mutations in the SPG11 gene, on chromosome 15q. However, it is becoming evident that ARHSP-TCC can also be the clinical presentation of mutations in ZFYVE26 (SPG15), as shown by the recent identification of eight families with a variable phenotype. Here, we present an additional Italian ARHSP-TCC patient harboring two new, probably loss-of-function mutations in ZFYVE26. This finding, together with the report of a mutation in another Italian family, provides confirmation that ZFYVE26 is the second gene responsible for ARHSP-TCC in the Italian population.

First evidence of a pathogenic insertion in the NOTCH3 gene causing CADASIL

CADASIL (OMIM 125310) is an increasingly recognised adult-onset autosomal-dominant vascular disease that is characterised by recurrent transient ischaemic attacks and strokes (43% of patients), vascular dementia (6%), migraine with aura (40% of patients) and psychiatric disturbances (9% of patients); epilepsy has been reported in 2–10% of subjects.1 All patients revealed prominent signal abnormalities on brain magnetic resonance imaging (MRI)—leukoencephalopathy on T2- and small subcortical infarcts on T1-weighted images.2 The pathological hallmark of CADASIL is a non-amyloid and non-arteriosclerotic angiopathy, which predominantly affects the small penetrating brain arteries. Vascular lesions are characterised by degeneration and loss of smooth-muscle cells and by the presence of granular osmiophilic material (GOM) accumulating within the smooth-muscle-cell basement membrane and the surrounding extracellular matrix. Examination of several peripheral organs revealed vessel changes, including the presence of GOM deposits, providing evidence that CADASIL is a systemic arteriopathy.3 It has been reported that CADASIL is caused by single missense mutations, small in-frame deletions or splice-site mutations in the NOTCH3 gene encoding a transmembrane receptor (http://www.hgmd.cf.ac.uk/ac/gene.php?gene = NOTCH3). Almost all previously reported mutations resulted in an odd number of cysteine residues within one of the 34 epidermal growth factor (EGF)-like repeats in the …

A novel locus for dHMN with pyramidal features maps to chromosome 4q34.3-q35.2

The distal hereditary motor neuropathy (dHMN) is a rare genetically and clinically heterogeneous disorder characterized by weakness and wasting of distal limb muscles in absence of overt sensory abnormalities. Recently, pyramidal signs have been also described in some patients with dominant or recessive dHMN, and two different loci have been identified in families affected by dHMN complicated with pyramidal dysfunction. We investigated an Italian family affected by an autosomal dominant dHMN complicated by pyramidal signs in order to map a new gene locus. The disease maps to a novel locus in a 26‐cM region flanked by D4S1552 and D4S2930 on chromosome 4q34.3‐35.2. Three candidate genes (SNX25, CASP3 and TUBB4Q) located in the critical region were screened for the presence of mutations by heteroduplex analysis. No mutations have been detected in the analyzed genes. In conclusion, the new private genetic locus we reported further confirms the wide heterogeneity of dHMN.

Further evidence that DDHD2 gene mutations cause autosomal recessive hereditary spastic paraplegia with thin corpus callosum

A. Magariello*, L. Citrigno*, S. Zuchner, M. Gonzalez, A. Patitucci, V. Sofia, F. L. Conforti, I. Pappalardo, R. Mazzei, C. Ungaro, M. Zappia and M. Muglia Institute of Neurological Sciences, National Research Council, Mangone (CS), Italy; Department of Human Genetics and Hussman Institute for Human Genomics, Miller School of Medicine, University of Miami, Miami, FL, USA; and Dipartimento G.F. Ingrassia Sezione di Neuroscienze Universit a di Catania, Catania, Italy

A novel KIF5A mutation in an Italian family marked by spastic paraparesis and congenital deafness

Hereditary spastic paraplegia (HSP) includes a group of diseases characterized by progressive spastic weakness of the lower limbs (pure forms) with possible additional signs (complicated forms). The SPG10 form is due to alteration in the kinesin1A gene (KIF5A) that encodes the neuronal kinesin heavy chain, a protein required for the anterograde axonal transport. We performed clinical, neurophysiological and molecular studies in two siblings affected by AD-HSP complicated by deafness. The screening of the KIF5A gene revealed the novel mutation p.Leu259Gln in two affected siblings and in their father with a pure form of HSP.

Further evidence that D90A-SOD1 mutation is recessively inherited in ALS patients in Italy.

Mutations in the Cu/Zn superoxide dismutase 1 (SOD1) gene have been reported to cause adult-onset autosomal dominant amyotrophic lateral sclerosis (FALS). In sporadic cases (SALS), de novo mutations in the SOD1 gene have occasionally been observed. All the SOD1 mutations are autosomal dominantly inherited with the exception of D90A. To date, in Italy, only two sporadic ALS cases carrying the D90A mutation have been reported in a homozygous state. We investigated for the presence of this mutation in 169 unrelated ALS patients from southern Italy. The genetic analysis revealed three ALS patients (1.8%) with mild phenotype carrying the homozygous D90A mutation.