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Connexin 32 (Cx32) is a gap junction protein expressed in the peripheral nervous system (PNS), central nervous system (CNS), and in many other tissues.1 Mutations in the Cx32 gene are associated with X-linked Charcot-Marie-Tooth disease (CMTX) , and account for about 10% of the patients with hereditary motor and sensory neuropathy (HMSN).
At least 130 different mutations have been reported in the Cx32 gene causing peripheral neuropathy. Classically, distal weakness and atrophy initially involving the lower limbs, as well as sensory abnormalities, depressed tendon reflexes, and pes cavus are usually found in males by the second decade, whereas in carrier females clinical manifestations, if present, are in most instances milder than in affected males. Nerve conduction studies in affected males are usually, but not always, suggestive of a demyelinating process, although they are not quite as slow as in patients with CMT1A. In females, conduction velocities (CVs) may be in the normal range or only mildly reduced, as seen in axonal neuropathies.
We describe a new Cx32 point mutation (Ala39 to Val) in genetically established identical twins with similar CMT phenotypes and extensor plantar reflexes.
The probands were first seen at the age of 20. Their principal complaint was cramps in the legs, “going over” on the ankles, and mild weakness in the hands. On examination, Twin 1 could not stand on his heels and had a mild intrinsic hand muscle weakness. There was a mild distal atrophy in both upper and lower limbs. Pinprick and tactile sensations were diminished up to the knees and vibration was impaired distally in the lower limbs. Tendon reflexes were absent or depressed, but both plantar responses were extensor. His median, ulnar, and peroneal motor CVs were 33.0 m/s, 33.0 m/s, and 31.0 m/s, respectively, and the distal amplitudes were 0.7 mV, 5.0 mV, and 3.3 mV. The sensory potentials were all absent. Twin 2 had identical clinical manifestations, except that the left plantar reflex was flexor whereas the right was clearly extensor. His motor CVs and amplitudes of the same nerves described above were 32.0 m/s and 1.7 mV, 34.0 m/s and 6.0 mV, and 33.0 m/s and 4.0 mV, respectively. No sensory response was obtained. Their mother had minimal neuropathic features and both plantar reflexes were extensor. Her median and peroneal motor CVs were 43.0 m/s and 37.0 m/s, and the median sensory CV was 40.0 m/s. Their sister and the mother’s brother were clinically and electrophysiologically normal. The maternal grandfather was not examined, but had a long history of a slowly progressive neuropathy.
The presence of the 17p11.2-p12 duplication was excluded with fluorescent quantitative polymerase chain reaction with five microsatellite markers contained within the involved segment.
Sequencing Cx32 with the ABITM Dye Primer Cycle Sequencing Ready Reaction detected a C179 to T transition (figure) at amino acid 39 causing an alanine to valine substitution in the first extracellular loop. This mutation abolishes a restriction site for the enzyme Bbv I and on screening 200 control chromosomes and the father’s DNA, no mutation was found. The mother was shown to harbour the mutation.
The monozygosity status of the twins was confirmed by the demonstration of the same alleles at each of the 13 highly polymorphic microsatellite markers tested. The possibility of this occurring by chance is >0.01%.
CMTX is now recognised as a frequent cause of HMSN.1Mutations have been detected in all domains of the protein and are postulated to be either non-functional or exert a dominant-negative effect. The clinical manifestations detected in this family with a novel point mutation leading to an Ala39 to Val amino acid substitution are clearly on the mild side of the classic CMT phenotype spectrum. This amino acid is conserved in other species, not found in 200 control chromosomes and segregates with the disease.
A second notable feature in this family is the presence of extensor plantar responses in all three people shown to carry the mutation. Involvement of the CNS in patients with Cx32 mutations have been demonstrated by slowing of the central conduction time in their brain stem auditory evoked potentials,2 but clinical manifestations secondary to central dysfunction does not seem to be a frequent finding. Paulson et al 3 reported a patient who developed dysarthria and incoordination after high altitude skiing. His MRI showed confluent, symmetric, white matter changes. Another member of the family carrying the mutation had normal MRI4 and other non-related patients with the same mutation did not show any clinical signs of CNS involvement, raising the possibility of a casual association. Bellet al 5 presented a family with a mutation on code 93 whose clinical manifestations included tremor, brisk reflexes, and spasticity. On MRI there was atrophy of the cerebral cortex and cerebellum. The presence of a Babinski’s sign in our family strongly suggests that in this novel mutation there is involvement of the corticospinal tract. Unfortunately no imaging or evoked potential studies were possible.
Cx32 is a gap junction protein expressed in the paranodal region and Schmidt-Lanterman incisures in the PNS, and in cell bodies and oligodendrocytes processes in the CNS. Why mutations in Cx32 usually lead only to PNS dysfunction is still an open question. Presumably, there is a unique relation between Cx32 and the structural organisation or metabolic requirements of the PNS. Another possibility is that other connexin proteins might compensate for Cx32 dysfunction in the CNS and other tissues, but not in the PNS.
Although the clinical manifestations are extremely similar in most of the Cx32 neuropathies suggesting that different mutations do not cause different phenotypes, different degrees of severity and the presence of unusual signs, like the one we present here, have already been described to occur with some mutations.
There are only two previous reports relating to three pairs of identical twins with CMT and known genetic defects. In the two pairs with the 17p11.2 duplication there was remarkable clinical variability.6 We have also seen a pair of identical twins with a P0 mutation in whom there was marked variability in early ages (unpublished data). Apart from the asymmetry of toe responses in one of the probands, the genetically identical twins described here are phenotypically very similar, suggesting that the expression of this mutation was not influenced by other non-genetic factors.
Codon 39 seems to be of particular importance to Cx32 protein function as changing of the wild type amino acid has caused CNS dysfunction in addition to the peripheral neuropathy. Moreover its expression does not seem to depend on non-genetic factors, as might be expected in a hemizygous condition.
WMJr was supported by grants from CAPES, FAPESP, and FAEPA (Brazil).
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