Congenital fibrosis of the extraocular muscles (autosomal dominant congenital external ophthalmoplegia): genetic homogeneity, linkage refinement, and physical mapping on chromosome 12.

Congenital fibrosis of the extraocular muscles (CFEOM) is an autosomal dominant syndrome of congenital external ophthalmoplegia and bilateral ptosis. We previously reported linkage of this disorder in two unrelated families to an 8-cM region near the centromere of human chromosome 12. We now present refinement of linkage in the original two families, linkage analysis of five additional families, and a physical map of the critical region for the CFEOM gene. In each of the seven families the disease gene is linked to the pericentromeric region of chromosome 12. D12S345, D12S59, D12S331, and D12S1048 do not recombine with the disease gene and have combined lod scores of 35.7, 35.6, 16.0, and 31.4, respectively. AFM136xf6 and AFMb320wd9 flank the CFEOM locus, defining a critical region of 3 cM spanning the centromere of chromosome 12. These data support the concept that this may be a genetically homogeneous disorder. We also describe the generation of a YAC contig encompassing the critical region of the CFEOM locus. This interval has been assigned cytogenetically to 12p11.2-q12 and spans the centromere of chromosome 12. These results provide the basis for further molecular analyses of the structure and organization of the CFEOM locus and will help in the identification of candidate genes.     

Congenital fibrosis of the vertically acting extraocular muscles maps to the <Emphasis Type="Italic">FEOM3</Emphasis> locus

The diagnosis of congenital fibrosis of the extraocular muscles (CFEOM) encompasses several different inherited strabismus syndromes characterized by congenital restrictive ophthalmoplegia affecting extraocular muscles innervated by the oculomotor and/or trochlear nerves. The OMIM database (http://www.ncbi.nlm.nih.gov/Omim/) currently contains four familial CFEOM phenotypes: CFEOM1-3, which map to the FEOM1-3 loci (MIM 135600, 602078, 604361), respectively, and congenital fibrosis of the vertically acting extraocular muscles (MIM 600638), reported in a single family without a corresponding genotype. We have had the opportunity to study the reported family with this fourth phenotype and now demonstrate that their phenotype can be reclassified as CFEOM3 and that it maps to FEOM3, flanked by D16S498 to 16qter, with a maximum lod score of 6.0.     

KIF21A基因的p.Arg954Trp突变引起中国人先天性眼外肌纤维化

一型先天性眼外肌纤维化（Congenital fibrosis of the extraocular muscles, CFEOM）是一种罕见的常染色体显性遗传的眼肌疾病,临床上主要表现为动眼神经缺陷而引起的斜视。本研究鉴定了具有四代病人的一个呈常染色体显性遗传的CFEOM1家系,连锁分析表明致病基因与染色体12q处的微卫星标记D12S85紧密连锁,最大LOD值为2．1。对D12S85附近的CFEOM1基因K1F21A进行突变检测,在K1F21A基因第21个外显子发现有一C→T的碱基替换,该变化引起K1F21A基因的第954位密码子由精氨酸突变为色氨酸,SSCP结果表明该家系中的所有患者都具有这一突变,而在家系中的所有正常人以及150个正常汉人对照中则不能检测到这一改变。我们的研究表明,K1F21A的p．Arg954Trp突变是引起这一先天性眼外肌纤维化家系病人患病的致病原因。     

Genomic organization and single-nucleotide polymorphism map of desmuslin, a novel intermediate filament protein on chromosome 15q26.3

Background

Congenital fibrosis of the extraocular muscles type 1 (CFEOM1) is an autosomal dominant eye movement disorder linked to the pericentromere of chromosome 12 (12p11.2 - q12). Sarcospan is a member of the dystrophin associated protein complex in skeletal and extraocular muscle and maps to human chromosome 12p11.2. Mutations in the genes encoding each of the other components of the skeletal muscle sarcospan-sarcoglycan complex (α - δ sarcoglycan) have been shown to cause limb girdle muscular dystrophy (LGMD2C-F). To determine whether mutations in the sarcospan gene are responsible for CFEOM1 we: (1) attempted to map sarcospan to the CFEOM1 critical region; (2) developed a genomic primer set to directly sequence the sarcospan gene in CFEOM1 patients; and (3) generated an anti-sarcospan antibody to examine extraocular muscle biopsies from CFEOM1 patients.

Results

When tested by polymerase chain reaction, sarcospan sequence was not detected on yeast or bacterial artificial chromosomes from the CFEOM1 critical region. Sequencing of the sarcospan gene in CFEOM1 patients from 6 families revealed no mutations. Immunohistochemical studies of CFEOM1 extraocular muscles showed normal levels of sarcospan at the membrane. Finally, sarcospan was electronically mapped to bacterial artificial chromosomes that are considered to be outside of the CFEOM1 critical region.

Conclusions

In this report we evaluate sarcospan as a candidate gene for CFEOM1. We have found that it is highly unlikely that sarcospan is involved in the pathogenesis of this disease. As of yet no sarcospan gene mutations have been found to cause muscular abnormalities.     

Mutation p.Arg954Trp of KIF21A Causes Congenital Fibrosis of the Extraocular Muscles in a Chinese Family

Congenital fibrosis of the extraocular muscles type 1 (CFEOM1) is an autosomal dominant strabismus disorder associated with defects of the oculomotor nerve. In this study, we identified a Chinese family with CFEOM1 for four generations. Linkage analysis mapped the causative gene of the family to 12q with a Lod score 2.1 for polymorphic marker D12S85, where KIF21A is located. Direct DNA sequence analysis identified a 2860C→T change in exon 21, resulting in a tryptophan substitution for arginine in codon 954 of KIF21A. SSCP (single-stranded conformational polymorphism) analysis showed that mutation p.Arg954Trp of KIF21A co-segregated with the affected members, but was absent in the unaffected individuals in the family and 150 normal controls. Our results indicate that mutation p.Arg954Trp of the KIF21A is the genetic basis of the Chinese family with CFEOM1.     

Spatiotemporal expression pattern of KIF21A during normal embryonic development and in congenital fibrosis of the extraocular muscles type 1 (CFEOM1)

Congenital fibrosis of the extraocular muscles type 1 (CFEOM1) is a rare inherited strabismus syndrome characterized by non-progressive ophthalmoplegia. We previously identified that CFEOM1 results from heterozygous missense mutations in KIF21A, which encodes a kinesin motor protein. Here we evaluate the expression pattern of KIF21A in human brain and muscles of control and CFEOM1 patients, and during human and mouse embryonic development. KIF21A is expressed in the cell bodies, axons, and dendrites of many neuronal populations including those in the hippocampus, cerebral cortex, cerebellum, striatum, and motor neurons of the oculomotor, trochlear, and abducens nuclei from early development into maturity, and its spatial distribution is not altered in the CFEOM1 tissues available for study. Multiple splice isoforms of KIF21A are identified in human fetal brain, but none of the reported CFEOM1 mutations are located in or near the alternatively spliced exons. KIF21A immunoreactivity is also observed in extraocular and skeletal muscle biopsies of control and CFEOM1 patients, where it co-localizes with triadin, a marker of the excitation-contractile coupling system. The diffuse and widespread expression of KIF21A in the developing human and mouse central and peripheral nervous system as well as in extraocular muscle does not account for the restricted ocular phenotype observed in CFEOM1, nor does it permit the formal exclusion of a myogenic etiology based on expression patterns alone.     

一个常染色体显性遗传先天性眼外肌 纤维化家系A

为寻找疾病相关基因,通过随访调查、体检、病理检查等手段,发现了一眼外肌纤维化家系4代中有15人患有眼外肌纤维化综合征,主要表现先天性上眼睑下垂、下颌上举、头后仰、双眼固定下转位和被动牵拉试验阳性,眼外肌病理检查结果为肌纤维化和玻璃样变性,所有阳性体征者除眼球运动限制程度有区别外,其他眼部症状基本相同。遗传分析表明,该疾病属常染色体显性遗传。该家系可作为寻找眼外肌纤维化疾病相关基因的宝贵资源。Abstract: To discover novel disease genes, a family with congenital fibrosis of the extraocular muscle was studied by a follow-up investigation, eye examinations and histo-pathological examination. There were fifteen cases suffering from congenital general fibrosis syndrome in four generations. They have congenital blepharoptosis, head tilt, chin lift, primary gaze fixed in a hypo- and exotropic position. The diagnosis is confirmed with positive forced duction testing in the affected eye. Furthermore, fibrosis of the extraocular muscles and hyaline degeneration was confirmed by histo-pathological examination. Except for different levels of restriction of the eyeball movements , other eye symptoms in positive patients are substantially identical. The genetic analysis showed that this disease was caused by autosomal dominant inheritance. The pedigree may be precious resource candidate for discovering disease gene related with congenital fibrosis of the extraocular muscle.     

Mitochondrial neurogastrointestinal encephalomyopathy syndrome maps to chromosome 22q13.32-qter.

Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) syndrome is a rare, multisystem disorder characterized clinically by ptosis, progressive external ophthalmoplegia, gastrointestinal dysmotility, leukoencephalopathy, thin body habitus, and myopathy. Laboratory studies reveal defects of oxidative-phosphorylation and multiple mtDNA deletions frequently in skeletal muscle. We studied four ethnically distinct families affected with this apparently autosomal recessive disorder. Probands from each family were shown, by Southern blot, to have multiple mtDNA deletions in skeletal muscle. We mapped the MNGIE locus to 22q13.32-qter, distal to D22S1161, with a maximum two-point LOD score of 6.80 at locus D22S526. Cosegregation of MNGIE with a single chromosomal region in families with diverse ethnic backgrounds suggests that we have mapped an important locus for this disorder. We found no evidence to implicate three candidate genes in this region, by using direct sequence analysis for DNA helicase II and by assaying enzyme activities for arylsulfatase A and carnitine palmitoyltransferase.     

The contribution of the DFNB1 locus to neurosensory deafness in a Caucasian population.

Classical studies have demonstrated genetic heterogeneity for nonsyndromic autosomal recessive congenital neurosensory deafness, with at least six loci postulated. Linkage analysis in two consanguineous Tunisian kindreds has demonstrated that one such deafness locus, DFNB1, maps near chromosome 13 markers D13S175, D13S143, and D13S115. We tested these markers for cosegregation with deafness in 18 New Zealand and 1 Australian nonconsanguineous kindreds, each of which included at least two siblings with nonsyndromic presumed congenital sensorineural deafness and that had a pedigree structure consistent with autosomal recessive inheritance. When all families were combined, a peak two-point lod score of 2.547 (theta = .1) was obtained for D13S175, 0.780 (theta = .2) for D13S143, and 0.664 (theta = .3) for D13S115. While there was no statistically significant evidence for heterogeneity at any of the three loci tested, nine families showed cosegregation of marker haplotypes with deafness. These observations suggest that the DFNB1 locus may make an important contribution to autosomal recessive neurosensory deafness in a Caucasian population. In the nine cosegregating families, phenotypic variation was observed both within sibships (in four families), which indicates that variable expressivity characterizes some genotypes at the DFNB1 locus, and between generations (in two families), which suggests allelic heterogeneity.     

Linkage of Wolfram syndrome to chromosome 4p16.1 and evidence for heterogeneity.

Wolfram syndrome (DIDMOAD syndrome; MIM 222300) is an autosomal recessive neurodegenerative disorder characterized by juvenile-onset diabetes mellitus and bilateral optic atrophy. Previous linkage analysis of multiply affected families indicated that the gene for Wolfram syndrome is on chromosome 4p, and it produced no evidence for locus heterogeneity. We have investigated 12 U.K. families with Wolfram syndrome, and we report confirmation of linkage to chromosome 4p, with a maximum two-point LOD score of 4.6 with DRD5, assuming homogeneity, and of 5.1, assuming heterogeneity. Overlapping multipoint analysis using six markers at a time produced definite evidence for locus heterogeneity: the maximum multipoint LOD score under homogeneity was <2, whereas when heterogeneity was allowed for an admixture a LOD of 6.2 was obtained in the interval between D4S432 and D4S431, with the peak close to the marker D4S3023. One family with an atypical phenotype was definitely unlinked to the region. Haplotype inspection of the remaining 11 families, which appear linked to chromosome 4p and had typical phenotypes, revealed crossover events during meiosis, which also placed the gene in the interval D4S432 and D4S431. In these families no recombinants were detected with the marker D4S3023, which maps within the same interval.     

Homozygosity mapping, to chromosome 11p, of the gene for familial persistent hyperinsulinemic hypoglycemia of infancy.

Familial persistent hyperinsulinemic hypoglycemia of infancy (PHHI) is a rare, autosomal recessive disease of unregulated insulin secretion, defined by elevations in serum insulin despite severe hypoglycemia. We used the homozygosity gene-mapping strategy to localize this disorder to the region of chromosome 11p between markers D11S1334 and D11S899 (maximum LOD score 5.02 [theta = 0] at marker D11S926) in five consanguineous families of Saudi Arabian origin. These results extend those of a recent report that also placed PHHI on chromosome 11p, between markers D11S926 and D11S928. Comparison of the boundaries of these two overlapping regions allows the PHHI locus to be assigned to the 4-cM region between the markers D11S926 and D11S899. Identification of this gene may allow a better understanding of other disorders of glucose homeostasis, by providing insight into the regulation of insulin release.     

Tightly linked flanking microsatellite markers for the Usher syndrome type I locus on the short arm of chromosome 11.

Usher syndrome type I is an autosomal recessive disease characterized by profound congenital hearing impairment and vestibular dysfunction followed by the onset of progressive pigmentary retinopathy in childhood or early adolescence. A locus (USH1C) for one form of this disease was previously assigned to the short arm of chromosome 11 through linkage studies in the Acadian population of southwestern Louisiana. Linkage analyses of a set of microsatellite markers in 27 Acadian families provide evidence that USH1C lies between D11S861 and D11S928. Three markers (D11S419, D11S921, and D11S899) that lie between the flanking markers show no recombination with USH1C, and all 54 chromosomes with the abnormal allele at the disease locus have identical alleles for D11S419 and D11S921. This haplotype was found on only 10 of 50 chromosomes with the normal allele at the disease locus, suggesting a strong founder effect. Of the 54 chromosomes with the abnormal allele, 12 had a divergent allele at D11S899. These results suggest that USH1C is in the 2-3-cM interval between D11S861 and D11S899.     

Coinheritance of chromosomes 3 and 11 in the patients with autosomal recessive polycythemia from Chuvachia

Inheritance of chromosomes 3 and 11 in the families with Chuvash autosomal recessive polycythemia and in control group with no disease symptoms was examined using polymorphic dinucleotide markers D3S1597 and D3S1263, mapped to region 3p25, and D11S4111, D11S4127, and D11S1356, mapped to region 11q23. All patients were homozygous for the C598T mutation in the VHL gene (3p25). The analysis showed that in 75% of the cases, chromosome 3 carrying C598T mutation was coinherited with certain chromosome 11, which differed from 50%, expected upon independent inheritance of each chromosome. In case of chromosome 3 without C598T mutation, this pattern was observed neither in healthy sibs form the families with autosomal recessive polycythemia (44%), nor in the control group (43%). These results suggest that in case of the C598T mutation in the VHL gene, chromosomal loci 3p25 and 11q23 are inherited not independently, compared to the inheritance of these loci in the absence of the mutation in healthy sibs from the affected families (chi2 = 16.14; P < 0.001), and also in the control family sample (chi2 = 17.91; P < 0.001).     

Osteoporosis-pseudoglioma syndrome, a disorder affecting skeletal strength and vision, is assigned to chromosome region 11q12-13.

Osteoporosis-pseudoglioma syndrome (OPS) is an autosomal recessive disorder characterized by severe juvenile-onset osteoporosis and congenital or juvenile-onset blindness. The pathogenic mechanism is not known. Clinical, biochemical, and microscopic analyses suggest that OPS may be a disorder of matrix homeostasis rather than a disorder of matrix structure. Consequently, identification of the OPS gene and its protein product could provide insights regarding common osteoporotic conditions, such as postmenopausal and senile osteoporosis. As a first step toward determining the cause of OPS, we utilized a combination of traditional linkage analysis and homozygosity mapping to assign the OPS locus to chromosome region 11q12-13. Mapping was accomplished by analyzing 16 DNA samples (seven affected individuals) from three different consanguineous kindreds. Studies in 10 additional families narrowed the candidate region, supported locus homogeneity, and did not detect founder effects. The OPS locus maps to a 13-cM interval between D11S1298 and D11S971 and most likely lies in a 3-cM region between GSTP1 and D11S1296. At present, no strong candidate genes colocalize with OPS.     

Autosomal recessive familial exudative vitreoretinopathy is associated with mutations in LRP5

Familial exudative vitreoretinopathy (FEVR) is a hereditary eye disorder that affects both the retina and vitreous body. Autosomal recessive FEVR was diagnosed in multiple individuals from three consanguineous families of European descent. A candidate-locus-directed genome scan shows linkage to the region on chromosome 11q flanked by markers D11S905 and D11S1314. The maximum LOD score of 3.6 at theta =0 is obtained with marker D11S987. Haplotype analysis confirms that the critical region is the 22-cM (311-Mb) interval flanked by markers D11S905 and D11S1314. This region contains LRP5 but not FZD4; mutations in both of these genes cause autosomal dominant FEVR. Sequencing of LRP5 shows, in all three families, homozygous mutations R570Q, R752G, and E1367K. This suggests that mutations in this gene can cause autosomal recessive as well as autosomal dominant FEVR.     

Localization of two genes for Usher syndrome type I to chromosome 11.

The Usher syndromes (USH) are autosomal recessive diseases characterized by congenital sensorineural hearing loss and progressive pigmentary retinopathy. While relatively rare in the general population, collectively they account for approximately 6% of the congenitally deaf population. Usher syndrome type II (USH2) has been mapped to chromosome 1q (W. J. Kimberling, M. D. Weston, C. M?ller, et al., 1990, Genomics 7: 245-249; R. A. Lewis, B. Otterud, D. Stauffer, et al., 1990, Genomics 7: 250-256), and one form of Usher syndrome type I (USH1) has been mapped to chromosome 14q (J. Kaplan, S. Gerber, D. Bonneau, J. Rozet, M. Briord, J. Dufier, A. Munnich, and J. Frezal, 1990. Cytogenet. Cell Genet. 58: 1988). These loci have been excluded as regions of USH genes in our data set, which is composed of 8 French-Acadian USH1 families and 11 British USH1 families. Both of these sets of families show linkage to loci on chromosome 11. Linkage analysis demonstrates locus heterogeneity between these sets of families, with the French-Acadian families showing linkage to D11S419 (Z = 4.20, theta = 0) and the British families showing linkage to D11S527 (Z = 6.03, theta = 0). Genetic heterogeneity of the data set was confirmed using HOMOG and the M test (log likelihood ratio > 10(5)). These results confirm the presence of two distinct USH1 loci on chromosome 11.     

Mutations in CYP1B1, the gene for cytochrome P4501B1, are the predominant cause of primary congenital glaucoma in Saudi Arabia.

The autosomal recessive disorder primary congenital glaucoma (PCG) is caused by unknown developmental defect(s) of the trabecular meshwork and anterior chamber angle of the eye. Homozygosity mapping with a DNA pooling strategy in three large consanguineous Saudi PCG families identified the GLC3A locus on chromosome 2p21 in a region tightly linked to PCG in another population. Formal linkage analysis in 25 Saudi PCG families confirmed both significant linkage to polymorphic markers in this region and incomplete penetrance, but it showed no evidence of genetic heterogeneity. For these 25 families, the maximum combined two-point LOD score was 15.76 at a recombination fraction of .021, with the polymorphic marker D2S177. Both haplotype analysis and homozygosity mapping in these families localized GLC3A to a 5-cM critical interval delineated by markers D2S2186 and D2S1356. Sequence analysis of the coding exons for cytochrome P4501B1 (CYP1B1) in these 25 families revealed three distinctive mutations that segregate with the phenotype in 24 families. Additional clinical and molecular data on some mildly affected relatives showed variable expressivity of PCG in this population. These results should stimulate a study of the genetic and environmental events that modify the effects of CYP1B1 mutations in ocular development. Furthermore, the small number of PCG mutations identified in this Saudi population makes both neonatal and population screening attractive public health measures.     

Localization of the Homolog of a Mouse Craniofacial Mutant to Human Chromosome 18q11 and Evaluation of Linkage to Human CLP and CPO

The transgene-induced mutation 9257 and the spontaneous mutation twirler cause craniofacial and inner ear malformations and are located on mouse chromosome 18 near the ataxia locusax.To map the human homolog of 9257, a probe from the transgene insertion site was used to screen a human genomic library. Analysis of a cross-hybridizing human clone identified a 3-kb conserved sequence block that does not appear to contain protein coding sequence. Analysis of somatic cell hybrid panels assigned the human locus to 18q11. The polymorphic microsatellite markers D18S1001 and D18S1002 were isolated from the human locus and mapped by linkage analysis using the CEPH pedigrees. The 9257 locus maps close to the centromeres of human chromosome 18q and mouse chromosome 18 at the proximal end of a conserved linkage group. To evaluate the role of this locus in human craniofacial disorders, linkage to D18S1002 was tested in 11 families with autosomal dominant nonsyndromic cleft lip and palate and 3 families with autosomal dominant cleft palate only. Obligatory recombinants were observed in 8 of the families, and negative lod scores from the other families indicated that these disorders are not linked to the chromosome 18 loci.     

Linkage analysis in spinal muscular atrophy, by six closely flanking markers on chromosome 5

The proximal spinal muscular atrophies (SMA) represent the second most common autosomal recessive disorder, after cystic fibrosis. The gene responsible for chronic SMA has recently been mapped to chromosome 5q by using genetic linkage studies. Among six markers mapping to this region, five were shown to be linked with the SMA locus in 39 chronic SMA families each containing at least two affected individuals. Multilocus analysis by the method of location score was used to establish the best estimate of the SMA gene location. Our data suggest that the most likely location for SMA is between loci D5S6 and D5S39. The genetic distances between these two markers are estimated to be 6.4 cM in males and 11.9 cM in females. Since meiosis were informative with D5S39 and D5S6 in 92% and 87% of SMA families, respectively, it is hoped that the present study will contribute to the calculation of genetic risk in SMA families.