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Am J Hum Genet. Aug 2000; 67(2): 357–368.
Published online Jul 5, 2000. doi:  10.1086/303011
PMCID: PMC1287211

The SPCH1 Region on Human 7q31: Genomic Characterization of the Critical Interval and Localization of Translocations Associated with Speech and Language Disorder


The KE family is a large three-generation pedigree in which half the members are affected with a severe speech and language disorder that is transmitted as an autosomal dominant monogenic trait. In previously published work, we localized the gene responsible (SPCH1) to a 5.6-cM region of 7q31 between D7S2459 and D7S643. In the present study, we have employed bioinformatic analyses to assemble a detailed BAC-/PAC-based sequence map of this interval, containing 152 sequence tagged sites (STSs), 20 known genes, and >7.75 Mb of completed genomic sequence. We screened the affected chromosome 7 from the KE family with 120 of these STSs (average spacing <100 kb), but we did not detect any evidence of a microdeletion. Novel polymorphic markers were generated from the sequence and were used to further localize critical recombination breakpoints in the KE family. This allowed refinement of the SPCH1 interval to a region between new markers 013A and 330B, containing ~6.1 Mb of completed sequence. In addition, we have studied two unrelated patients with a similar speech and language disorder, who have de novo translocations involving 7q31. Fluorescence in situ hybridization analyses with BACs/PACs from the sequence map localized the t(5;7)(q22;q31.2) breakpoint in the first patient (CS) to a single clone within the newly refined SPCH1 interval. This clone contains the CAGH44 gene, which encodes a brain-expressed protein containing a large polyglutamine stretch. However, we found that the t(2;7)(p23;q31.3) breakpoint in the second patient (BRD) resides within a BAC clone mapping >3.7 Mb distal to this, outside the current SPCH1 critical interval. Finally, we investigated the CAGH44 gene in affected individuals of the KE family, but we found no mutations in the currently known coding sequence. These studies represent further steps toward the isolation of the first gene to be implicated in the development of speech and language.


Between 2% and 5% of children who are otherwise normal have significant difficulties in acquiring expressive and/or receptive language, despite adequate intelligence and opportunity (Bishop et al. 1995). Strong evidence for genetic influences in developmental disorders of speech and language was found in a twin study that showed significant heritability for expressive subtypes of language impairment, both with and without articulation disorder (Bishop et al. 1995). The vast majority of families segregating such disorders do not follow a simple Mendelian inheritance pattern, so that the results of conventional parametric linkage analysis should usually be viewed with caution.

However, in 1990, Hurst et al. described a rare case of a large extended family (known as KE) in which the speech and language disorder is clearly inherited with an autosomal dominant, monogenic mode of transmission. Some reports have suggested that the core deficit of the disorder in this family is an inability to use grammatical suffixation rules, such as those for tense, number, and gender (Gopnik 1990; Gopnik and Crago 1991). Other studies have shown that the phenotype is not as selective and is characterized by difficulties with many aspects of grammar and expressive language (Hurst et al. 1990; Vargha-Khadem and Passingham 1990; Vargha-Khadem et al. 1995, 1998). Furthermore, the phenotype involves grossly defective articulation (verbal dyspraxia), such that the speech of affected individuals is largely incomprehensible to the naive listener (Hurst et al. 1990; Vargha-Khadem et al. 1995). In addition, there is evidence of moderate nonverbal cognitive impairment in some affected family members (Vargha-Khadem et al. 1995). Recent brain-imaging studies of affected individuals from this family revealed functional and structural abnormalities in both cortical and subcortical motor-related areas of the frontal lobe, particularly the basal ganglia (Vargha-Khadem et al. 1998). The findings of Vargha-Khadem et al. (1995, 1998) suggest that the central aspect of the disorder may be the disruption of selection and sequencing of fine orofacial movements, leading to deficits in the development of language skills.

We previously identified a region on chromosome 7 which cosegregates with the speech and language disorder in the KE family (maximum LOD score 6.62 at recombination fraction [θ] 0), confirming autosomal dominant inheritance with full penetrance (Fisher et al. 1998). Fine mapping using all the available Généthon microsatellites from the region allowed us to localize the gene responsible (termed SPCH1) to a ~5.6-cM interval of 7q31, flanked by markers D7S2459 and D7S643 (Fisher et al. 1998). The 7q31 band is well characterized at the physical level, with extensive coverage in YAC, BAC, and PAC clones (Bouffard et al. 1997). In addition, numerous BAC and PAC clones from this region are being actively sequenced, as part of a broader effort to sequence human chromosome 7, at the Washington University Genome Sequencing Center (WU-GSC). Several known genes and a large number of anonymous expressed-sequence tags (ESTs) have been mapped to 7q31 by YAC-based or radiation-hybrid mapping (Schuler et al. 1996; Bouffard et al. 1997).

In the present study, bioinformatic analyses have been employed to assemble a detailed BAC-/PAC-based sequence map of 7q31, which includes 152 sequence tagged sites (STSs), 20 known genes, and 50 anonymous transcripts, thus providing a framework for the positional cloning of the SPCH1 gene. We have used 120 of these STSs to screen the affected chromosome 7 from the KE family in a search for microdeletions. In addition, we have generated novel polymorphic markers from the sequence and have used these to extract additional linkage information from the KE family, in order to refine the SPCH1 interval. Finally, we report the localization of translocation breakpoints associated with speech and language disorder in two individuals, relative to the newly refined SPCH1 critical interval. We have demonstrated that one of these translocations maps to the same BAC as the CAGH44 gene, which encodes a polyglutamine-repeat protein present in brain, and, therefore, we have investigated this gene further in the KE family.

Subjects and Methods

Family KE

In 1987, the KE family (see fig. 1) was referred for genetic counseling by the director of a school for children with speech and language problems, at which many family members had been pupils. The condition was characterized as a developmental verbal dyspraxia (Hurst et al. 1990).

Figure  1
Pedigree of family KE, affected by speech and language disorder. Blackened symbols indicate affected individuals. Asterisks indicate those individuals who were unavailable for linkage analysis.

Patients Bearing Chromosome Translocations

CS is a 5.5-year-old boy with language impairment and verbal dyspraxia. He has a de novo balanced reciprocal translocation t(5;7)(q22;q31.2) which was identified prior to his birth via amniocentesis. Examination at birth showed no abnormalities, but he was referred back to the genetics team at age 2 years because of concerns of delayed speech and mild motor delay. Subsequent assessment at age 3 years 6 mo, performed by means of the Bailey scale, gave an overall mental development in the mildly delayed range. Although nonverbal skills were in the normal range, there was impairment in both understanding and expression of speech. A diagnosis was made of oral dyspraxia. By 4 years of age, CS was able to put two words together, and his understanding had progressed. Fine and gross motor development had also improved, although there was still evidence of mild impairment. There is no history of speech and language disorder in the family of CS, and none of his siblings (one full sibling and three half-siblings) have any language problems. His mother reports that he has never been able to laugh spontaneously or to sneeze.

BRD is an 8-year-old boy with a history of receptive and expressive language problems, accompanied by behavioral difficulties and low-range intellectual abilities, despite normal physical/motor development. He continues to show difficulties following verbal instructions in school, and has word-finding and sequencing problems accompanied by poor articulation. An MRI scan at age 6 years 11 months detected a small dysembryoplastic neuroepithelial tumor in his right temporal lobe. A more detailed description of the clinical phenotype of BRD is given in Warburton et al. (2000). Cytogenetic analysis previously revealed a de novo balanced reciprocal translocation, t(2;7)(p23;q31.3) (Warburton et al. 2000).

Construction of Human-Hamster Somatic Cell Hybrids

The Chinese hamster mutant cell line a23, deficient in thymidine kinase, was cultured in 5% CO2 as a monolayer in Dulbecco's modification of Eagle medium (DMEM, Life Technologies) supplemented with 10% fetal calf serum (FCS). White cells were separated from peripheral blood from three affected members of the KE family (II-2, II-9, and III-20) using Histopaque (Sigma). Each sample was combined in equal proportions with trypsinized a23 cells and was placed in serum-free medium. After the cells were spun down, the medium was aspirated. The cell mixture was resuspended in 50% polyethylene glycol (PEG, molecular weight 1450, Sigma), was washed, and was set up in culture with DMEM and 10% FCS at low cell density. After 24 h, the culture medium was replaced with the same medium containing hypoxanthine, aminopterin, and thymidine (HAT, Life Technologies). Once hybrid colonies reached 2–4 mm in diameter (14 d after fusion), they were picked into 24-well plates. DNA prepared from samples of 1×105–5×105 cells was tested by PCR for chromosome 7–specific markers.

Genotyping and Linkage Analysis

Primers flanking novel polymorphic repeats were designed using the PRIMER program, accessed through the United Kingdom Human Genome Mapping Project resource center. Fluorescence-based semiautomated genotyping was performed as described (Fisher et al. 1998). Linkage analyses were run under the assumption that the disorder in the KE pedigree is due to a single autosomal dominant locus with full penetrance, as described (Fisher et al. 1998).

Fluorescence In Situ Hybridization (FISH)

PHA-stimulated T lymphocytes or lymphoblastoid cells lines were harvested by conventional techniques, and fixed suspensions were dropped onto slides. Slides were denatured at 70°C in 70% formamide/2×SSC for 2 min 30 s, were incubated in cold 2×SSC, and were serially dehydrated in 70%, 90%, and 100% (twice) ethanol at room temperature. Probe DNA was labeled by nick translation with biotin (Gibco BRL BioNick Labeling System) or Digoxigenin (DIG; Roche) following manufacturers’ protocols. FISH of BACs and PACs was performed as described (Millwood et al. 1997). Biotinylated probes were visualized with two layers of FITC-conjugated streptavidin (green; Vector Labs) and biotinylated goat anti-streptavidin (Vector Labs). DIG-labeled probes were visualized with mouse anti-DIG antibodies (Roche), followed by Cy5-conjugated rabbit anti-mouse and goat anti-rabbit antibodies (pseudocolored blue). Chromosomes were counterstained with Vectorshield containing propidium iodide (Vector Labs). The slides were viewed on a Nikon Optiphot, and images were captured with a Bio-Rad MRC 1024 laser-scanning confocal microscope and Lasersharp software.

Determination of Genomic Organization of CAGH44

Exon/intron boundaries for exons 1–2 of CAGH44 were identified by comparison of the reported mRNA sequence (accession U80741) to completed BAC sequence. Boundaries for exons 3–6 were determined using either long-range PCR or vectorette (Munroe et al. 1994). For the former, we amplified products from human genomic or BAC DNA, using the Expand Long Template System (Boehringer Mannheim), with primers designed from CAGH44 mRNA. Products were sequenced using BigDye Terminator Cycle Sequencing kits (PE Applied Biosystems). For the vectorette method, libraries were made by complete digestion of BAC DNA using frequent-cutting restriction enzymes followed by ligation to annealed vectorette bubble anchors. Fragments containing exon/intron boundary sequences were amplified from vectorette libraries by PCR with the NotI-A primer in combination with specific primers derived from available CAGH44 sequences.


Construction of a BAC-/PAC-Based Sequence Map Spanning the SPCH1 Interval of 7q31

Preliminary analysis of chromosome 7 physical-mapping data indicated that 152 STSs map to the original SPCH1 interval, between D7S2459 and D7S643, specifically within four YAC contigs, designated N, O, P, and Q (Bouffard et al. 1997). Although many BAC and PAC clones from this region have been sequenced and deposited as separate records in the public databases, much of the sequence information is not fully cross-referenced and integrated with other related sequences or with other mapping data. Therefore, we compared the sequence for all 152 STSs mapping between D7S2459 and D7S643 to the “non-redundant,” “high-throughput genome sequence,” and WU-GSC human sequencing project-specific databases using the BLAST algorithm (Altschul et al. 1997). At the time of the most recent analyses (March 2000), 132 of these STSs were found in fully sequenced BACs/PACs, whereas an additional 4 were detected in clones for which partial sequence was available. Thus, only 16 STSs were not found in a sequenced clone.

BLAST analysis of the insert ends of sequenced clones facilitated the establishment of overlaps among adjacent clones and orientation of various sequence blocks. These electronic analyses allowed us to assemble a detailed BAC-/PAC-based sequence map of 7q31 consisting of several blocks of contiguous sequence, the largest of which exceeds 1.5 Mb in length (fig. 2). Within these sequenced regions, we have been able to accurately determine marker order and intermarker distances. In a number of cases, the marker order differs from that previously established in physical-mapping studies. We estimate that the D7S2459–D7S643 interval currently contains >7.75 Mb of completed sequence. No sequence has been obtained, as yet, from the region covered by YAC contig P (sWSS1095-sWSS3263), and this region is estimated from physical mapping studies to span ~1.25 Mb (Bouffard et al. 1997). Therefore, these analyses indicate that the D7S2459–D7S643 interval is likely to be [gt-or-equal, slanted]9 Mb in size. Of note, all pairs of adjacent STSs within assembled sequence blocks are separated by <220 kb.

Figure  2
Sequence map of the D7S2459–D7S643 interval. Previously ordered using YACs, 152 STSs provide a framework for the depicted BAC-/PAC-based sequence map. The most likely marker order was established by analysis of available genomic sequence data, ...

Transcript Map of 7q31

GeneMap ’99 includes 147 ESTs that have been mapped to the interval between anchor markers D7S2459 and D7S480 using radiation-hybrid panels G3 and GB4. Analysis of these ESTs suggested that they cluster into <96 transcripts, including 18 genes that have been fully sequenced and/or encode proteins of known function. Electronic PCR and BLAST searches revealed that the majority (>85%) of the ESTs map to sequenced BACs/PACs. A total of 50 of the anonymous transcripts and 13 of the known genes from GeneMap ’99 could be localized precisely within the D7S2459–D7S643 sequence contigs. (Eleven anonymous transcripts and three known genes are contained in sequenced clones that map proximal or distal to the D7S2459–D7S643 interval; the remaining 19 transcripts were not found in BAC/PAC sequences.) An additional six genes not present in the GeneMap ’99 interval (PDS, LAMB1R, MDG1, CAGH44, TSA806, and KCND2) were identified and were localized within the D7S2459–D7S643 contigs via annotations in BAC/PAC sequence records. Finally, one of the STSs (sWSS3217) residing within an as-yet-unsequenced region was found, by homology screening, to correspond to the HF.12 gene. Figure 2 shows the relative positions of all the known genes detected, with additional summary information provided in table 1.

Table 1
Known Genes Residing between D7S2459 and D7S643

Search for a 7q31 Microdeletion in Family KE

Previously, cytogenetic analyses of affected members of the KE family failed to detect any chromosomal abnormality. However, these studies did not rule out a microdeletion within the SPCH1 interval. We therefore chose to screen affected individuals for the absence of STSs within the critical region. This analysis required the prior separation of the chromosome 7 harboring the mutation from the normal homologue. Somatic hybrid cell lines containing human chromosome 7 on a hamster background were derived from three affected individuals of the KE family (see Methods). DNA from these cell lines was genotyped with the Généthon markers D7S692 and D7S522, previously shown to be polymorphic in the affected individuals. In each case, one allele precisely cosegregates with SPCH1 (Fisher et al. 1998). This allowed identification of hybrid cell lines containing only an affected chromosome 7, those containing only a normal chromosome 7, and those containing both copies of chromosome 7. Hybrid cell lines harboring only the affected chromosome 7 were tested for the presence of 120 STSs mapping within the D7S2459–D7S643 interval (see fig. 2). The average distance between these STSs is <100 kb. In all cases, the expected PCR product was generated from the hybrid cell lines but not from hamster DNA controls, indicating that all 120 STSs are present on the affected chromosome 7 in the KE family.

Generation of Novel Polymorphic Markers in 7q31 and Fine Mapping of the SPCH1 Locus

Our previous linkage study of the KE family with all the Généthon markers from 7q31 identified critical recombinations in affected female III-12 and unaffected male III-3 that defined D7S2459 and D7S643 as proximal and distal limits for SPCH1, respectively (Fisher et al. 1998) (see fig. 3). Within this ~5.6-cM region, we demonstrated that the ~3.6-cM interval between D7S692/D7S2425 and CFTR cosegregates perfectly with the disorder (see figs.figs. 2 2 and and3).3). We therefore used data from the BAC/PAC sequence contigs to develop novel polymorphic markers from the D7S2459–D7S692 and CFTR–D7S643 intervals, in order to refine further the positions of the III-12 proximal and III-3 distal recombination breakpoints, respectively. Note that the only Généthon markers from these intervals (D7S2456, D7S655, and D7S2487) were uninformative with respect to the disorder in the KE family.

Figure  3
Haplotype analysis of new markers from the D7S2459–D7S643 region limits the SPCH1 interval between 013A and 330B. This figure shows critical recombinants in unaffected individual III-3 and affected individual III-12. In addition, haplotypes are ...

Finished sequence data from these intervals was searched for stretches of unbroken tandem dinucleotide repeats with copy numbers of [gt-or-equal, slanted]16, which would be predicted to be polymorphic (Weber 1990). PCR primers flanking these repeats were used for semiautomated genotyping of the KE family. The results were analyzed with parametric linkage programs and haplotypes were determined as in our previous linkage study. Positions of new markers relative to the STS map could be accurately established from the sequence data (see fig. 2).

Genotyping with four novel markers at the proximal end (table 2) revealed that the recombination breakpoint in individual III-12 maps between 013A and 062B-369B-369C. This refines the SPCH1 interval by a few hundred kb at the proximal end, and excludes three known genes as candidates: PDS, DRA, and DLD (fig. 2). At the distal end of the SPCH1 interval, physical mapping data indicated that a polymorphic tetranucleotide repeat, D7S2847, lies between CFTR and D7S643. Genotyping of the KE family with D7S2847 demonstrated that the recombination in individual III-3 maps proximal to this marker. Investigation of four new markers generated from the sequence data between CFTR and D7S2847 (table 2) revealed that the III-3 recombination breakpoint is localized between 363B-084A and 330B. (084B was uninformative with respect to the disorder.) This result excludes a region of >2.65 Mb, containing the KCND2 gene, from the distal end of the SPCH1 interval. In addition, it limits the SPCH1 locus to within YAC contigs N and O, with only one uncloned gap in the region. Haplotypes of the relevant markers for critical individuals from the KE family are shown in figure 3.

Table 2
Novel Polymorphic Markers Generated from Sequence Contigs

Two Translocation Breakpoints in 7q31 Associated with Speech and Language Disorder

We have investigated de novo balanced translocations involving 7q31 in two unrelated patients with speech and language disorder, CS 46,XY t(5;7)(q22;q31.2) and BRD 46,XY t(2;7)(p23;q31.3) (see Subjects and Methods for clinical descriptions). The 7q31 breakpoints were localized using two-color FISH to metaphase spreads, with BACs/PACs selected from proximal and distal ends of the 7q31 contigs. The breakpoints of both patients were found to map in the D7S2459–D7S643 interval (fig. 4), suggesting that the translocations might be relevant to the etiology of speech and language disorder.

Figure  4
Two-color FISH analyses of translocations in patients with speech and language disorder, using BACs from 7q31. A, Patient CS; RG250D13 (blue) hybridizes to normal 7 and der(7) and is proximal to the translocation breakpoint, while NH0563O05 (green) hybridizes ...

Metaphase FISH with a series of further clones identified from the contigs mapped the CS breakpoint between RG250D13, which consistently hybridized to the derivative 7 [der(7)] and RG308B22, which consistently hybridized to the derivative 5 [der(5)]. Two BACs have been identified on the basis of fingerprint data to span the gap between these clones (fig. 2), and these are in the process of being sequenced. Whereas clone NH0208M04 mapped to der (5), NH0563O05 gave signals on both der(7) and der(5), suggesting that this BAC crosses the 7q31 breakpoint (fig. 4).

A recent study using FISH with YAC clones reported that the BRD translocation mapped between CFTR and D7S643 (Warburton et al. 2000). We have been able to confirm this and to localize the breakpoint at higher resolution using BAC/PAC clones. These analyses indicate that the BRD breakpoint maps within GS180J15, which gives signals on both der(7) and der(2) (fig. 4). Therefore, the 7q31 breakpoints of CS and BRD are separated by [gt-or-equal, slanted]3.75 Mb of DNA, as estimated from completed sequence and the size of YAC contig P (fig. 2). Furthermore, the BRD translocation maps [gt-or-equal, slanted]1.45 Mb distal to the newly refined SPCH1 interval, as established from the most recent linkage analyses of the KE family reported above (fig. 2).

Candidate Gene CAGH44, in the Vicinity of the CS Translocation

CAGH44 mRNA was originally isolated from human brain in an attempt to identify novel CAG-repeat-containing cDNAs whose expansion might be implicated in the etiology of neuropsychiatric disorders (Margolis et al. 1997). The CAGH44 product contains a stretch of 40 consecutive glutamine residues, which is the longest polyglutamine tract found thus far in an unexpanded protein. This is encoded by a combination of CAG and CAA codons, such that there are never more than five consecutive CAGs. There is a second polyglutamine stretch, containing only 10 glutamines, encoded by (CAG)7(CAA)(CAG)(CAA), which is separated from the first stretch by eight amino acids. Currently, there is only partial cDNA sequence reported for this gene, covering 912 bases of the coding region, and no information about its genomic structure.

Although Margolis et al. (1997) previously localized CAGH44 to 6q14-15 by radiation-hybrid mapping, the chromosome 7 physical map and sequence data indicates that, in fact, it resides in 7q31, close to the CS translocation breakpoint (fig. 2). Specifically, bioinformatic analyses found that RG250D13, the BAC hybridizing to der(7) adjacent to the CS translocation breakpoint, includes the first 258 bases of the partial CAGH44 coding sequence, organized into two exons. Analysis of sequence data from clones linking RG250D13 to RG308B22 enabled us to orient the completed RG250D13 sequence (including the two 5′ CAGH44 exons) relative to the sequence contigs. This indicated that the more 3′ exons of CAGH44 should lie on the distal side of RG250D13. Using long-range PCR and the vectorette method on BAC DNA, we were able to confirm this and to determine the genomic organization of the CAGH44 3′ coding region (table 3), thereby demonstrating the presence of four 3′ exons (3, 4, 5, and 6) in clone NH0563O05, in the interval between sWSS2794 and sWSS1765. This coincides exactly with the region containing the CS breakpoint (fig. 2). Bioinformatic analysis of genomic sequence from BAC RG250D13, upstream of the proposed CAGH44 coding region, confirms that the first ATG in the reported cDNA sequence is very likely to correspond to the start of the open reading frame. Therefore, exon 1 is indeed the first coding exon of this gene, although there may be additional exons upstream of this containing 5′ untranslated sequence. However, the ORF probably extends beyond the 3′ end of the currently reported cDNA sequence, since no in-frame stop codon has yet been reached. In addition, our PCR and vectorette analyses indicate that the last 43 bases (870–912) in the cDNA sequence reported by Margolis et al. (1997) do not come from the CAGH44 gene, but are likely to be an artifact of the original cloning procedure. This hypothesis is supported by the presence of an EcoRI site at 871–876 and by the observation that bases 878–912 are highly homologous (94% identity) to human ribosomal protein S16 mRNA.

Table 3
Genomic Organization of CAGH44[Note]

Given that CAGH44 maps to the same region as the CS breakpoint and encodes a brain-expressed polyglutamine repeat, we decided to analyze this gene in family KE. We did not detect any expansion of the regions encoding the polyglutamine stretches in members of the family. Furthermore, sequencing of the currently known CAGH44 coding region did not reveal any variant cosegregating with the disorder.


Our previous investigation of family KE provided the first clues to the chromosomal location of a gene involved in speech and language disorder. In the present work, we have used the extensive bioinformatic resources available to characterize the region of 7q31 that is likely to contain this gene. A significant proportion of this interval is represented by sequenced genomic clones; 87% of the framework STSs that map between D7S2459 and D7S643 are present within complete BAC or PAC sequence entries in Genbank. When these separate entries are assembled into large blocks of contiguous sequence and are integrated with other mapping data, the resulting sequence map is a powerful tool for investigating the region of interest, as illustrated here. For example, by searching for tandem dinucleotide repeats in specific intervals of the 7q31 sequence map, we were able to generate novel polymorphic markers for refined linkage analyses, allowing us to narrow the SPCH1 interval in family KE by several Mb.

In the absence of additional large families segregating verbal dyspraxia that might aid the fine mapping of SPCH1, the study of chromosomal rearrangements such as those described here provides an alternative means of narrowing the search for the gene. However, we note that observation of a breakpoint mapping to the critical interval in a patient affected with speech and language disorder does not in itself provide sufficient evidence of a causal role for the chromosomal abnormality. In addition, the breakpoint of a chromosomal rearrangement can sometimes map outside the transcription and promoter regions of the gene implicated in the etiology of the disease but still disrupt expression of this gene via a “position effect.” Some studies have demonstrated position effects acting as far as 900 kb from the gene responsible for the disorder (see Kleinjan and van Heyningen 1998). Therefore, drawing conclusions from the mapping of a chromosomal rearrangement can be difficult, unless they are substantiated by converging data from multiple cases or other kinds of analysis.

We have demonstrated that, although both the CS and BRD breakpoints map to 7q31, they are separated by >3.75 Mb of DNA. Thus, even allowing for the possibility of position effects, it seems unlikely that these breakpoints are influencing a single locus and that the patients have the same disease etiology. The developmental delay of BRD appears to be less selective than that of CS, involving behavioral problems and some general cognitive deficit, in addition to his speech and language difficulties. Warburton et al. (2000) previously mapped the BRD breakpoint to the CFTR–D7S643 interval and suggested that it may disrupt SPCH1. Our fine mapping of the BRD translocation, in combination with the refinement of the critical region from new linkage analyses of the KE family, indicates that this breakpoint in fact maps >1.45 Mb outside the current SPCH1 interval. The clone spanning the BRD breakpoint has been fully sequenced, but no transcripts have been found by electronic analyses of this sequence. Although it is possible that the BRD translocation could alter expression of the brain-specific KCND2 gene, whose promoter lies a few hundred kb distal to the breakpoint, our fine-mapping linkage results have excluded that locus as a candidate for SPCH1. It is worth noting that BRD has a right temporal lobe tumor, which, by virtue of its developmental origin, could interfere with the emergence of cognitive abilities, including those of speech and language.

In contrast, our analysis of the CS translocation places it within a single BAC (NH0563O05) inside the refined SPCH1 critical interval and suggests that it may disrupt the locus for CAGH44, a brain-expressed gene encoding a long polyglutamine tract. NH0563O05 is currently being sequenced and we are in the process of identifying the exact position of the breakpoint in order to confirm that the CS translocation does indeed interrupt the CAGH44 coding region. As stated above, further evidence will be required to properly establish a causal role of this breakpoint and the CAGH44 gene in the speech and language difficulties of CS.

The CAGH44 gene was first identified in a search for loci encoding polymorphic glutamine repeats whose expansion might cause neurological diseases (Margolis et al. 1997). Analysis of the polyglutamine-rich region showed that it is completely stable in normal individuals (Margolis et al. 1997), most likely because the CAG repeats that encode it are frequently interrupted by CAA codons, suggesting that they are unlikely to undergo the multistep gradual expansion typically associated with CAG-repeat disorders. Nevertheless, a recent study has shown that expansion of an impure CAG repeat (that of TATA-binding protein) can occur and can lead to a neurological disease (Koide et al. 1999). Therefore, on the basis of this and the results from the mapping of the CS translocation, CAGH44 appears to be a good candidate for SPCH1. Our mutation analysis of the currently known coding region for this gene did not detect expansion of the polyglutamine stretches or, indeed, any other variants cosegregating with the disorder in family KE. Since the most 3′ portion of the open reading frame was not isolated in the Margolis et al. (1997) study, we are at present identifying and fully characterizing the CAGH44 gene so that we may investigate it further in the KE family and in the patient CS.

We previously commented on the fact that the SPCH1 interval overlaps with a ~40-cM region identified in a genome screen for susceptibility to autism, a disorder which is often associated with speech and language abnormalities (Fisher et al. 1998; International Molecular Genetic Study of Autism Consortium 1998). Further support for a gene influencing autistic disorder on 7q has been found in additional linkage studies, but regions of linkage vary between data sets; whereas some show overlap with SPCH1, others implicate a region distal to this (Ashley-Koch et al. 1999; Barrett et al. 1999; International Molecular Genetic Study of Autism Consortium 1999; Philippe et al. 1999). We, and others, have identified chromosomal rearrangements (including translocations, inversions, and a duplication) involving 7q31 in patients with autism (Ashley-Koch et al. 1999; Vincent et al. 1999; Warburton et al. 2000; C. S. L. Lai, S. E. Fisher, P. A. Jacobs, E. R. Levy, C. G. Woods, and A. P. Monaco, unpublished data). However, breakpoints for these rearrangements map to a number of different sites, some within the SPCH1 interval and others distal to it, in a broad region of 7q31. Therefore, no simple relationship has yet emerged between the positions of chromosomal breakpoints in 7q and autism or language disorder. In addition, we note that the severe speech and language difficulties experienced by family KE are distinct from those normally associated with autistic disorder and that no members of this family have ever shown any autistic features. Thus, there is insufficient evidence from current clinical, cytogenetic, and linkage data to resolve the issue of whether there is a single 7q31 locus responsible for SPCH1 and autism susceptibility or two separate, adjacent loci contributing independently to these disorders.

In conclusion, our genomic characterization of the SPCH1 region has provided a framework for the investigations of family KE and the translocation patients presented in this report. These studies represent further steps towards the isolation of the first gene to be implicated in the development of speech and language.


We are very grateful to the KE family and to subjects CS and BRD and their families. We thank the Washington University Genome Sequencing Center for the generation of chromosome 7 sequence data. We thank Pam Warburton and Zoe Docherty for their help with the investigation of patient BRD. This study was funded by the Wellcome Trust. A.P.M. is a Wellcome Trust Principal Research Fellow.

Electronic-Database Information

URLs for data in this article are as follows:

Electronic PCR screening, http://www.ncbi.nlm.nih.gov/STS
GeneMap ’99, http://www.ncbi.nlm.nih.gov/genemap (for radiation-hybrid map data)
HGMP, http://www.hgmp.mrc.ac.uk (for PRIMER program)
NCBI BLAST, http://www.ncbi.nlm.nih.gov/BLAST/ (for homology searches of sequence data)
NHGRI chromosome 7–mapping data, http://genome.nhgri.nih.gov/chr7
UniGene, http://www.ncbi.nlm.nih.gov/UniGene/ (for clustering of ESTs)
WU-GSC chromosome 7 sequencing data and BLAST searches, http://genome.wustl.edu/gsc


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