Mutational analysis reveals the FUS homolog TAF15 as a candidate gene for familial amyotrophic lateral sclerosis.

FUS, EWS, and TAF15 belong to the TET family of structurally similar DNA/RNA-binding proteins. Mutations in the FUS gene have recently been discovered as a cause of familial amyotrophic lateral sclerosis (FALS). Given the structural and functional similarities between the three genes, we screened TAF15 and EWS in 263 and 94 index FALS cases, respectively. No coding variants were found in EWS, while we identified six novel changes in TAF15. Of these, two 24 bp deletions and a R388H missense variant were also found in healthy controls. A D386N substitution was shown not to segregate with the disease in the affected pedigree. A single A31T and two R395Q changes were identified in FALS cases but not in over 1,100 controls. Interestingly, one of the R395Q FALS cases also harbors a TARDBP mutation (G384R). Altogether, these results suggest that additional studies are needed to determine whether mutations in the TAF15 gene represent a cause of FALS.

Analysis of FUS gene mutation in familial amyotrophic lateral sclerosis within an Italian cohort.

OBJECTIVE: Mutations in the FUS gene on chromosome 16 have been recently discovered as a cause of familial amyotrophic lateral sclerosis (FALS). This study determined the frequency and identities of FUS gene mutations in a cohort of Italian patients with FALS. METHODS: We screened all 15 coding exons of FUS for mutations in 94 Italian patients with FALS. RESULTS: We identified 4 distinct missense mutations in 5 patients; 2 were novel. The mutations were not present in 376 healthy Italian controls and thus are likely to be pathogenic. CONCLUSIONS: Our results demonstrate that FUS mutations cause approximately 4% of familial amyotrophic lateral sclerosis cases in the Italian population.

Mutations in the FUS/TLS gene on chromosome 16 cause familial amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal degenerative motor neuron disorder. Ten percent of cases are inherited; most involve unidentified genes. We report here 13 mutations in the fused in sarcoma/translated in liposarcoma (FUS/TLS) gene on chromosome 16 that were specific for familial ALS. The FUS/TLS protein binds to RNA, functions in diverse processes, and is normally located predominantly in the nucleus. In contrast, the mutant forms of FUS/TLS accumulated in the cytoplasm of neurons, a pathology that is similar to that of the gene TAR DNA-binding protein 43 (TDP43), whose mutations also cause ALS. Neuronal cytoplasmic protein aggregation and defective RNA metabolism thus appear to be common pathogenic mechanisms involved in ALS and possibly in other neurodegenerative disorders.

An Unusual Occurrence of Southern Rust, Caused by Rpp9-virulent Puccinia polysora, on Corn in Southwestern Georgia.

Southern rust, caused by Puccinia polysora Underw., occurs frequently on corn (Zea mays) grown in subtropical or tropical regions. When conditions are favorable, southern rust also occurs in temperate climates of the central and southern United States although the fungus does not survive on corn crop residue and must be introduced to temperate regions each growing season. Several single, dominant, resistance genes, designated as Rpp genes, convey hypersensitive, chlorotic fleck reactions when challenged with avirulent isolates of P. polysora (1). Rpp resistance prevents or limits the formation of uredinia. The Rpp9 gene has been used successfully in North America in the past 20 years to control southern rust even though the gene has been ineffective in other parts of the world (e.g., Africa and Hawaii) because of the prevalence of virulent races. During the past 3 years, Rpp9 virulence has occurred in the western hemisphere (e.g., Brazil, Mexico, Nebraska, and Texas), but prior to 2008, uredinia were not observed east of the Mississippi River on corn with the Rpp9 gene. A few uredinia were observed on corn with the Rpp9 gene in eastern Nebraska in 2006 and near Victoria, TX in 2007 (W. Dolezal, personal observation). In July of 2008, a virulent isolate of P. polysora was confirmed from Grady County, GA on corn lines with the Rpp9 gene including the original source of this resistance gene, Boesman yellow flint, which is PI 186208 (3). In August of 2008, isolates of P. polysora were collected from severely infected corn hybrids with Rpp9 grown in Macon County, GA. Rust samples from hybrids without Rpp genes also were collected in Burke County, GA where Rpp-resistant corn was asymptomatic. In greenhouse trials, corn lines with and without the Rpp9 gene were inoculated with urediniospores from collections from Burke and Macon counties and Illinois. Rust infection types (1) were scored 18 to 25 days after inoculation. The Macon County isolate produced type 1 and 2 infections (small uredinia surrounded by necrotic or chlorotic tissue) on Oh43Rpp9 and W64aRpp9 and type 4 infections (large, sporulating uredinia) on two versions of a commercial hybrid with and without the Rpp9 gene and on Va59 (which carries an Rpp gene different from Rpp9). The Burke County isolate and an isolate from Illinois collected in 2001 produced type 0 infections (chlorotic flecks) on all of these lines except the non-Rpp version of the commercial hybrid which had a type 4 reaction. To our knowledge, Rpp9-virulent isolates of P. polysora have not been reported from the continental United States for nearly 50 years. In the late 1950s and early 1960s, A. L. Robert (2) collected isolates of P. polysora from throughout the world and observed multiple races on a set of host differentials that is no longer available. A. L. Robert's collection included an isolate from Georgia that was virulent on PI 186208. Commercial hybrids containing the Rpp9 gene may continue to be resistant throughout most of North America if previously common Rpp9-avirulent isolates of P. polysora are prevalent, but those hybrids should be carefully monitored for infection by newly introduced Rpp9-virulent isolates. References: (1) A. L. Hooker. Page 207 in: The Cereal Rusts. Vol. II. Academic Press, San Diego, 1985. (2) A. L. Robert. Phytopathology 52:1010, 1962. (3) A. J. Ullstrup. Phytopathology 55:425, 1965.

New VAPB deletion variant and exclusion of VAPB mutations in familial ALS.

OBJECTIVE: Amyotrophic lateral sclerosis (ALS) is a progressive, neurodegenerative disorder involving upper and lower motor neurons. The vesicle-associated membrane protein B (VAPB) gene has been genetically linked to ALS in several large Brazilian families in which the disorder is caused by a proline to serine mutation at codon 56 (P56S). No additional mutations have been identified. METHODS: To establish the prevalence of VAPB mutations, we screened 80 familial ALS samples by DNA sequencing. RESULTS: Our study failed to identify any novel VAPB gene mutations but identified a single Brazilian family harboring the P56S mutation. In a second familial ALS case, we identified a three-base pair deletion within exon 5 of the VAPB gene that deleted the serine residue at position 160 (Delta S160). This variant is detected in a normal population at low frequency (0.45%). Analyses of homology alignment and secondary structure predict that this deletion significantly alters the structure of VAPB, although a GFP-Delta S160 VAPB fusion protein demonstrates a wild-type subcellular localization. This contrasts the aberrant localization observed in a GFP-P56S VAPB fusion protein. The allele frequency of Delta S160 in patients with sporadic ALS does not differ significantly from that in the normal population. CONCLUSIONS: Mutations in the VAPB gene are rare and the Delta S160 variant does not contribute to the development of amyotrophic lateral sclerosis.

SOD1A4V-mediated ALS: absence of a closely linked modifier gene and origination in Asia.

Familial amyotrophic lateral sclerosis (ALS) accounts for 10% of all ALS. Approximately 20% of cases are due to mutations in the Cu/Zn superoxide dismutase gene (SOD1). In North America, SOD1(A4V) is the most common SOD1 mutation. Carriers of the SOD1(A4V) mutation share a common phenotype with rapid disease progression and death on average occurring at 1.4 years (versus 3-5 years with other dominant SOD1 mutations). Previous studies of SOD1(A4V) carriers identified a common haplotype around the SOD1 locus, suggesting a common founder for most SOD1(A4V) patients. In the current study we sequenced the entire common haplotypic region around SOD1 to test the hypothesis that polymorphisms in either previously undescribed coding regions or non-coding regions around SOD1 are responsible for the more aggressive phenotype in SOD1(A4V)-mediated ALS. We narrowed the conserved region around the SOD1 gene in SOD1(A4V) ALS to 2.8Kb and identified five novel SNPs therein. None of these variants was specifically found in all SOD1(A4V) patients. It therefore appears likely that the aggressive nature of the SOD1(A4V) mutation is not a result of a modifying factor within the region around the SOD1 gene. Founder analysis estimates that the A4V mutation occurred 540 generations (approximately 12,000 years) ago (95% CI 480-700). The conserved minimal haplotype is statistically more similar to Asian than European population DNA sets, suggesting that the A4V mutation arose in native Asian-Americans who reached the Americas through the Bering Strait.

Motoneuron-specific NR3B gene: no association with ALS and evidence for a common null allele.

OBJECTIVE: The GRIN3B gene encodes NR3B, a motoneuron-specific member of the NMDA type of ionotropic glutamate receptors. NR3B reduces the Ca(2+)-permeability as well as the overall current of the receptor response and may thereby protect motoneurons against glutamate-mediated excitotoxicity. We tested whether genetic dysfunction of GRIN3B is implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS). METHODS: We searched for mutations in the GRIN3B coding region (3.1 kb) in 117 individuals with familial ALS and in 46 individuals with sporadic ALS. We genotyped the newly identified GRIN3B null allele and four "tag single nucleotide polymorphisms (SNPs)" at the GRIN3B locus in 342 individuals with sporadic ALS and in 374 matched controls. The GRIN3B null allele frequency was determined in 2,128 individuals from a worldwide panel of 42 populations. We furthermore compared the GRIN3B coding sequence in primates (human-macaque) and rodents (rat-mouse) to evaluate the molecular evolution of GRIN3B. RESULTS: Thirty-two SNPs, including 16 previously unreported SNPs, one 27-bp deletion, a polymorphic CAG repeat, and a 4-bp insertion (insCGTT), were identified. Mutational and case-control studies did not reveal variants that cause or modify disease in ALS. Intriguing is an insCGTT variant that truncates the protein at its amino terminus and results in a GRIN3B null allele. We demonstrated a global distribution of the null allele with allele frequencies ranging between 0 and 0.38, and we delineated a null allele specific haplotype of 9.89 kb. Comparative genomic analysis across four taxa demonstrated accelerated evolution of NR3B in primates. CONCLUSIONS: Our study supports the conclusions that 1) GRIN3B does not seem to be associated with familial or sporadic ALS, 2) the GRIN3B null allele is a common polymorphism, 3) the GRIN3B null allele has arisen once and early in human evolution, and 4) the GRIN3B gene belongs to a group of nervous system-related genes that have been subjected to faster evolution during evolution.

A locus on chromosome 9p confers susceptibility to ALS and frontotemporal dementia.

OBJECTIVE: To perform genetic linkage analysis in a family affected with ALS and frontotemporal dementia (FTD). METHODS: The authors performed a genome-wide linkage analysis of a four-generation, 50-member Scandinavian family in which five individuals were diagnosed with ALS and nine with FTD. Linkage calculations assuming autosomal dominant inheritance of a single neurodegenerative disease manifesting as either ALS or FTD with age-dependent penetrance were performed. Further analyses for ALS alone and FTD alone were performed. A parametric logarithm of odds (lod) score of 2.0 or greater was required for further study of a potential locus and crossover (haplotype) analysis. RESULTS: A new ALS-FTD locus was identified between markers D9s1870 and D9s1791 on human chromosome 9p21.3-p13.3. A maximum multipoint lod score of 3.00 was obtained between markers D9s1121 and D9s2154. Crossover analysis indicates this region covers approximately 21.8 cM, or 14Mb. CONCLUSIONS: A locus on chromosome 9p21.3-p13.3 is linked to ALS-FTD.

Survival in transgenic ALS mice does not vary with CNS glutathione peroxidase activity.

OBJECTIVE: Transgenic mice that overexpress a human gene encoding mutant cytosolic superoxide dismutase (SOD1) develop a progressive motor neuron loss that resembles human ALS. Why mutant SOD1 initiates motor neuron death is unknown. One hypothesis proposes that the mutant molecule has enhanced peroxidase activity, reducing hydrogen peroxide (H2O2) to form toxic hydroxyl adducts on critical targets. To test this hypothesis, the authors generated transgenic ALS mice with altered levels of glutathione peroxidase (GSHPx), the major soluble enzyme that detoxifies H2O2. METHODS: SOD1(G93A) ALS mice were bred with mice bearing a murine GSHPx transgene that have a four-fold elevation in brain GSHPx levels and with mice having targeted inactivation of the GSHPx gene and reduced brain GSHPx activity. RESULTS: Survival was not prolonged in ALS mice with elevated brain GSHPx activity (p = 0.09). ALS mice with decreased GSHPx brain activity (20% of normal) showed no acceleration of the disease course (p = 0.89). The age at disease onset in the ALS mice was unaffected by brain GSHPx activity. CONCLUSION: The level of GSHPx activity in the CNS of transgenic ALS mice does not play a critical role in the development of motor neuron disease.

Elevated expression of hTERT is associated with dysplastic cell transformation during human oral carcinogenesis in situ.

PURPOSE: Telomerase is a ribonucleoprotein complex composed of the catalytic protein subunit (human telomerase reverse transcriptase or hTERT) and the RNA template. This enzyme activity is a necessary and rate-limiting step of cellular immortalization and could provide a unique marker of aberrant cells, which may selectively be targeted. The current study was undertaken to quantitatively determine the degree of telomerase activation during multistage oral carcinogenesis using paraffin-embedded tissue samples. EXPERIMENTAL DESIGN: hTERT expression level was quantitatively compared between normal and cancerous oral tissues by real-time reverse-transcription-PCR (RT-PCR). Also, the presence of hTERT transcript in individual cells was surveyed in the biopsy specimens with varying degrees of histopathology by in situ RT-PCR. RESULTS: Low level of hTERT amplification was detected by real-time RT-PCR in most (11/13) normal human oral mucosa. hTERT expression was also detected in the majority (11/12) of squamous cell carcinoma tissues, and the level was significantly (P < 0.05) elevated, on the average, by a factor >6.9. By in situ RT-PCR, hTERT expression was not noted in normal epithelium (0/10) nor in mild dysplasia (0/7) but was detected in moderate dysplasia (4/5) and in those tissues with a higher grade of histopathology: severe dysplasia (3/3) and invasive carcinoma (4/4). CONCLUSIONS: These results indicate that enhanced expression of telomerase activity occurs early during human oral carcinogenesis and support the critical role of telomerase in the development of human oral cancer.

Linkage of familial amyotrophic lateral sclerosis with frontotemporal dementia to chromosome 9q21-q22.

CONTEXT: Occasionally, 2 or more major neurodegenerative diseases arise simultaneously. An understanding of the genetic bases of combined disorders, such as amyotrophic lateral sclerosis (ALS) with frontotemporal dementia (FTD), will likely provide insight into mechanisms of these and related neurodegenerative diseases. OBJECTIVE: To identify loci that contain genes whose defects cause ALS. DESIGN: A genome-wide linkage analysis of 2 data sets from an ongoing study begun in the mid-1980s at 4 university research centers. SUBJECTS: An initial subset of 16 families (549 people) potentially informative for genetic analysis, in which 2 or more individuals were diagnosed as having ALS, identified from a Boston data set of 400 families and 4 families potentially informative (244 people) subsequently identified from a Chicago data set of more than 300 families to test a hypothesis based on findings from the Boston families. MAIN OUTCOME MEASURES: Linkage calculations assuming autosomal dominant inheritance with age-dependent penetrance (a parametric logarithm-of-odds [lod] score of 1.0 or greater required for further study of a potential locus); crossover analysis involving the ALS-FTD locus. RESULTS: In a set of families in which persons develop both ALS and FTD or either ALS or FTD alone, a genetic locus that is linked to ALS with FTD located between markers D9S301 and D9S167 was identified on human chromosome 9q21-q22. Families with ALS alone did not show linkage to this locus. Crossover analysis indicates this region covers approximately 17 cM. CONCLUSION: These data suggest that a defective gene located in the chromosome 9q21-q22 region may be linked to ALS with FTD. JAMA. 2000;284:1664-1669.

Refined mapping and characterization of the recessive familial amyotrophic lateral sclerosis locus (ALS2) on chromosome 2q33.

Amyotrophic lateral sclerosis (ALS) is a progressive degenerative neuromuscular disease that shows familial, autosomal dominant inheritance in 10%-15% of cases. Previous genetic analysis of one large family linked a recessive form of familial ALS (FALS-AR type 3) to the chromosome 2q33-35 region. Using additional polymorphic markers, we have narrowed the size of the linked region to approximately 1.7 cM by linkage and haplotype analysis. We have also established a yeast artificial chromosome contig across the locus that covers an approximate physical distance of 3 million bases. Based on this contig, genes and expressed sequences that map near the 2q33 region have been examined to determine whether they are located within this ALS2 candidate locus. Five identified genes and 34 expressed sequence tags map within the region defined by crossover analysis and merit further consideration as candidate genes for this disease.

Lack of association between apolipoprotein E genotype and sporadic amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neuro-degenerative disorder with both sporadic and familial forms. Approximately 20% of autosomal dominant ALS is caused by mutations in the Cu/Zn superoxide dismutase (SOD1) gene. The causes of the remaining forms of ALS are unknown. The apolipoprotein E (APOE) gene is a known genetic risk factor for Alzheimer disease (AD), another neuro-degenerative disease. The APOE-4 allele increases risk and decreases age at onset in AD. Studies examining ALS and APOE have failed to show a significant effect of APOE on overall risk in ALS. Studies examining the effect of APOE-4 on site of onset in ALS (bulbar or limb) have been contradictory, with some studies showing an APOE association with bulbar onset and others showing no effect. Sample size was limited in these previous reports, particularly with respect to the number of bulbar onset cases (n = 33, 34 and 53). The present study examines a large collaborative data set of ALS patients (n = 363; 95 with bulbar onset) and age-matched neurologically normal controls. The results for these data showed no significant differences in the percentage of subjects with the APOE-4/4 and APOE-4/X genotypes (X = APOE-2 or APOE-3) when comparing cases and controls in both the overall data set or in the data set stratified by site of onset. Similarly, logistic regression analysis in the overall and stratified data set while controlling for sex showed no increase or decrease in risk of ALS associated with the APOE-4 allele. In addition, there were no significant differences in age at onset between patients with APOE-X/X, and APOE-4/4 or APOE-4/X genotypes, overall or stratified by site of onset. We conclude based on these data that the APOE gene is not a major genetic risk factor for site of onset in ALS.

Epidemiology of mutations in superoxide dismutase in amyotrophic lateral sclerosis.

We registered 366 families in a study of dominantly inherited amyotrophic lateral sclerosis. Two hundred ninety families were screened for mutations in the gene encoding copper-zinc cytosolic superoxide dismutase (SOD1). Mutations were detected in 68 families. The most common SOD1 mutation is an alanine for valine substitution in codon 4 (50%). We present clinical and genetic data concerning 112 families with 395 affected individuals. The clinical characteristics of patients with familial amyotrophic lateral sclerosis arising from SOD1 mutations are similar to those lacking SOD1 defects. Mean age at onset was earlier (Wilcoxon test, p = 0.004) in the SOD1 group (46.9 years [standard deviation, 12.5] vs 50.5 years [11.5] in the non-SOD1 group). Bulbar onset was associated with a later onset age. The presence of either of two mutations, G37R and L38V, predicted an earlier age at onset. Kaplan-Meier plots demonstrated shorter survival in the SOD1 group compared with the non-SOD1 group at early survival times (Wilcoxon test, p = 0.0007). The presence of one mutation, A4V, correlated with shorter survival. G37R, G41D, and G93C mutations predicted longer survival. This information suggests it will be productive to investigate other genetic determinants in amyotrophic lateral sclerosis and to use epidemiological characteristics of the disease to help discern molecular mechanisms of motor neuron cell death.

Three novel mutations and two variants in the gene for Cu/Zn superoxide dismutase in familial amyotrophic lateral sclerosis.

Autosomal dominant inheritance is exhibited by about 10% of cases of amyotrophic lateral sclerosis (ALS), a paralytic disorder characterized by the death of motor neurons in the brain and spinal cord. A subgroup of these familial cases are linked to mutations in the gene which codes for Cu/Zn superoxide dismutase (SOD1). We report three additional mutations occurring in the SOD1 gene in ALS patients and two single base pair variant changes. The single base pair change in an ALS family causes a glycine 93 to valine substitution, which is the fifth distinct amino acid change reported for the glycine 93 residue. One missense mutation in exon 5 would substitute neutral valine for the negatively-charged aspartate 124 (aspartate 124 to valine). An individual with an apparently sporadic case of ALS carries a three base pair deletion in exon 5 of the SOD1 gene. These three mutations bring to 38 the total number of distinct SOD1 mutations associated with familial ALS.

Identification of three novel mutations in the gene for Cu/Zn superoxide dismutase in patients with familial amyotrophic lateral sclerosis.

About 10% of cases of amyotrophic lateral sclerosis (ALS), a paralytic disorder characterized by death of motor neurons in the brain and spinal cord, exhibit autosomal dominant inheritance. A subgroup of these familial cases are caused by mutations in the gene encoding Cu/Zn superoxide dismutase (SOD1). We report here three additional mutations occurring in the SOD1 gene in three families with ALS. Two of these changes are missense mutations in exon 5 of the SOD1 gene, resulting in leucine 144 to serine and alanine 145 to threonine substitutions. The third, a single base pair change in intron 4 immediately upstream of exon 5, results in an alternatively spliced mRNA. The alternate transcript conserves the open reading frame of exon 5, producing an SOD1 protein with three amino acids inserted between exons 4 and 5 (following residue 118). These three mutations bring to 29 the total number of distinct SOD1 mutations associated with familial ALS.

Superoxide dismutase concentration and activity in familial amyotrophic lateral sclerosis.

Some cases of autosomal-dominant familial amyotrophic lateral sclerosis (FALS) have been associated with mutations in SOD1, the gene that encodes Cu/Zn superoxide dismutase (Cu/Zn SOD). We determined the concentrations (microgram of Cu/Zn SOD/mg of total protein), specific activities (U/microgram of total protein), and apparent turnover numbers (U/mumol of Cu/Zn SOD) of Cu/Zn SOD in erythrocyte lysates from patients with known SOD1 mutations. We also measured the concentrations and activities of Cu/Zn SOD in FALS patients with no identifiable SOD1 mutations, sporadic ALS (SALS) patients, and patients with other neurologic disorders. The concentration and specific activity of Cu/Zn SOD were decreased in all patients with SOD1 mutations, with mean reductions of 51 and 46%, respectively, relative to controls. In contrast, the apparent turnover number of the enzyme was not altered in these patients. For the six mutations studied, there was no correlation between enzyme concentration or specific activity and disease severity, expressed as either duration of disease or age of onset. No significant alterations in the concentration, specific activity, or apparent turnover number of Cu/Zn SOD were detected in the FALS patients with no identifiable SOD1 mutations, SALS patients, or patients with other neurologic disorders. That Cu/Zn SOD concentration and specific activity are equivalently reduced in erythrocytes from patients with SOD1 mutations suggests that mutant Cu/Zn SOD is unstable in these cells. That concentration and specific activity do not correlate with disease severity suggests that an altered, novel function of the enzyme, rather than reduction of its dismutase activity, may be responsible for the pathogenesis of FALS.

A frequent ala 4 to val superoxide dismutase-1 mutation is associated with a rapidly progressive familial amyotrophic lateral sclerosis.

Familial amyotrophic lateral sclerosis (FALS), a degenerative disorder of motor neurons, is associated with mutations in the Cu/Zn superoxide dismutase gene SOD1 in some affected families. We confirm a recently reported ala4-->val mutation in exon 1 of the SOD1 gene and report that this mutation is both the most commonly detected of all SOD1 mutations and among the most clinically severe. By comparison with our other FALS families, the exon 1 mutation is associated with reduced survival time after onset: 1.2 years, as compared to 2.5 years for all other FALS patients. We also demonstrate that SOD1 is prominently expressed in normal motor neurons and that neural expression of SOD1 is not prevented by this exon 1 mutation. Assays of SOD1 enzymatic activity in extracts from red blood cells, lymphoblastoid cells, and brain tissues revealed an approximately 50% reduction in activity of cytosolic SOD1 in patients with this mutation compared to normal individuals. By contrast, patients with sporadic ALS had normal levels of SOD1 enzymatic activity. Why this SOD1 mutation causes motor neuron death in FALS remains to be established. While it may be that FALS is a consequence of loss of SOD1 function, it is also possible that motor neuron death in this dominantly inherited disease occurs because the mutations confer an additional, cytotoxic function on the SOD1 protein.

Genetic linkage analysis of familial amyotrophic lateral sclerosis using human chromosome 21 microsatellite DNA markers.

Amyotrophic lateral sclerosis (ALS: Lou Gehrig's Disease) is a lethal neurodegenerative disease of upper and lower motorneurons in the brain and spinal cord. We previously reported linkage of a gene for familial ALS (FALS) to human chromosome 21 using 4 restriction fragment length polymorphism DNA markers [Siddique et al.: N Engl J Med 324:1381-1384, 1991] and identified disease-associated mutations in the superoxide dismutase (SOD)-1 gene in some ALS families [Rosen et al.: Nature 362:59-62, 1993]. We report here the genetic linkage data that led us to examine the SOD-1 gene for mutations. We also report a new microsatellite DNA marker for D21S63, derived from the cosmid PW517 [VanKeuren et al.: Am J Hum Genet 38:793-804, 1986]. Ten microsatellite DNA markers, including the new marker D21S63, were used to reinvestigate linkage of FALS to chromosome 21. Genetic linkage analysis performed with 13 ALS families for these 10 DNA markers confirmed the presence of a FALS gene on chromosome 21. The highest total 2-point LOD score for all families was 4.33, obtained at a distance of 10 cM from the marker D21S223. For 5 ALS families linked to chromosome 21, a peak 2-point LOD score of 5.94 was obtained at the DNA marker D21S223. A multipoint score of 6.50 was obtained with the markers D21S213, D21S223, D21S167, and FALS for 5 chromosome 21-linked ALS families. The haplotypes of these families for the 10 DNA markers revealed recombination events that further refined the location of the FALS gene to a segment of approximately 5 megabases (Mb) between D21S213 and D21S219.(ABSTRACT TRUNCATED AT 250 WORDS)