In 2006, Okada and co-workers identified Dok-7 as a muscle-intrinsic activator of MuSK required for synaptogenesis.22
Dok-7 harbors N-terminal pleck-strin homology and phosphotyrosine-binding domains and is strongly expressed at the postsynaptic region of skeletal muscle and in heart. Subsequently, Beeson and co-workers detected recessive DOK7
mutations in CMS patients whose weakness was mostly in a limbgirdle distribution.23
The majority of patients were heterozygous or homozygous for a common 1124-1127dupTGCC mutation, but in two patients only a single mutation was detected, and only the common mutation was functionally characterized. In six patients eventually shown to carry DOK7
mutations by Beeson, Slater and co-workers described nonfatigable weakness largely confined to the proximal limb muscles. These patients had small EPs relative to muscle fiber size with few and simple junctional folds, which the authors considered a constitutive feature of the disease; AChR counts per EP were also reduced, but were deemed appropriate for small EPs.24
The amplitude of the miniature EP potentials (MEPPs) was decreased; this was largely attributed to the decreased input resistance of the simplified postsynaptic architecture. The quantal content (m
) and the amplitude of the EP potential (EPP) were decreased but the amplitude of the miniature EP currents (MEPCs) was not.
After Okada’s discovery, we considered DOK7
a good candidate gene in patients who had small EPs, or a reduced number of AChRs per EP, but no mutations in AChR or rapsyn. Using these criteria, we first identified 1325
and then three additional patients who carry DOK7
mutations. Fourteen of these had had intercostal muscle biopsies with detailed in vitro
electrophysiology and electron microscopy studies of the neuromuscular junction.25
All 16 patients had a significant EMG decrement, limb-girdle weakness, and short-term fatigability. In eight patients the disease presented in the neonatal period. Eyelid ptosis was present in 14 patients; it was frequently asymmetric but was severe only in one patient. Bulbar symptoms occurred in 11, oculoparesis in six, respiratory muscle involvement in 12 with evidence of ventilatory failure in six. Eleven patients experienced intermittent worsenings, and 12 had a progressive course with disability ranging from mild to severe at the time of the last evaluation.
Type-1 fiber preponderance was noted in 13 patients, type-2 fiber atrophy in eight, isolated necrotic or regenerating fibers in four, and pleomorphic oxidative-enzyme decreases or target formations in 12. All EPs consisted of one to multiple small synaptic contacts.
The mean MEPP, the MEPC amplitude, and the quantal content of the EPP were reduced by ~25%, but the distribution of patient and control values over-lapped. Electron microscopy of 613 EP regions of 409 EPs revealed disintegrating junctional folds in 33% (), denuded postsynaptic regions with nearby nerve sprouts in 18%, and degenerating subsynaptic organelles in 10% (). Importantly, however, some EPs in each patient had a normal postsynaptic architecture (), and the density and distribution of AChR on the crests of the preserved junctional folds was normal (); therefore, a simplified postsynaptic architecture is not a constitutive feature of Dok-7 myasthenia, and the defects in Dok-7 do not cause a decreased concentration of AChR on the preserved junctional folds of the innervated human neuromuscular junction.
FIGURE 9 Dok-7 myasthenia. (A) AChR localization at a structurally normal EP region with peroxidase-labeled α-bungarotoxin. The density and distribution of AChR on the junctional folds are normal. (B) A structurally abnormal EP region. Several junctional (more ...)
Dok-7 myasthenia. A normal EP region displays elongated, narrow, and complex junctional folds predicting a high input resistance. A number of EP regions with normal postsynaptic architecture were present in each patient. Bar, 1 μm.
Each patient harbored hetero-allelic pathogenic mutations: 11 of these were detected in genomic DNA and six in cDNA isolated from EP-enriched muscle specimens. Four mutations in genomic DNA were previously reported.23
No consistent phenotype—genotype correlations emerged.
We conclude that (1) the clinical features of Dok-7 myasthenia are highly variable, (2) the morphologicfeatures bespeak of ongoing destruction and remodeling of the EPs which likely contribute importantly to defective neuromuscular transmission, and (3) some mutations are complex and can only be identified and analyzed in cDNA or cloned cDNA.25