A transcription map of the minimally deleted region from 13q14 in B-cell chronic lymphocytic leukemia as defined by large scale sequencing of the 650 kb critical region.

Extensive analysis of tumors has demonstrated homozygous and heterozygous deletions in chromosome region 13q14.3 in B-cell chronic lymphocytic leukemia (B-CLL), suggesting the site of a tumor suppressor gene. Since previous searches for this gene have not yielded any viable candidates, we now present the sequence of the BACs which span the minimally deleted approximately 650 kb region between markers D13S319 and D13S25. This sequence has allowed us to create the definitive transcription map for the region which reveals 93 ESTs and 12 Unigene clusters in this region. Using gene prediction programs, a further 19 potential genes are also identified. The genes show an asymmetrical distribution throughout the region with most of them clustering at the extreme ends. This sequencing effort provides for the definitive structure of the B-CLL deletion region and the identification of the vast majority of the potential candidate genes. Of all the genes identified, only three have homologies to known genes: two L1 repeat genes and rabbit epididymal protein 52. This 13q14.3 sequence provides the final substrate from which to characterize the B-CLL tumor suppressor gene.

Lack of association of the (AAAT)6 allele of the GXAlu tetranucleotide repeat in intron 27b of the NF1 gene with autism.

A novel allele of the GXAlu tetranucleotide repeat in intron 27b of the neurofibromatosis 1 (NF1) gene has recently been reported to be present in 4.7% of autistic patients but not in controls. We have found the novel GXAlu allele absent in 204 patients from the South Carolina Autism Project and 200 controls. The autism population studied includes a significant number of patients with hypotonia, stereotyped behaviors, or postural, gait, and motor abnormalities similar to those seen in the patients previously reported to possess the novel GXAlu allele. This suggests that the novel (AAAT)6 GXAlu allele is not associated with autism.

Fine mapping of the PTGFR gene to 1p31 region and mutation analysis in human breast cancer.

The 1p31 chromosomal region shows loss of heterozygosity (LOH) in up to 50% of human breast cancer, indicating the presence of a tumor suppressor gene at this location. Many efforts have been made to identify candidate genes responsible for breast cancer on the short arm of chromosome 1. It was shown that prostaglandins have been implicated in the tumorigenesis pathway, perhaps via interactions with their cell surface receptors. The prostaglandin F2 receptor gene (PTGFR) was tentatively mapped to 1p31 adjacent to the region undergoing LOH in human breast cancer. We undertook a mutation study in 34 sporadic human breast tumors using a variant of SSCP, incorporation PCR SSCP (IPS). Several nucleotide variants were detected in different tumors. Here we report the nature of these nucleotide changes and the possible involvement of the PTGFR gene in the etiology of human cancer.

The HOPA gene dodecamer duplication is not a significant etiological factor in autism.

A recent study has suggested that a dodecamer duplication in the HOPA gene in Xq13 may occur in a significant portion of male patients with autism. We have determined the incidence of this duplication in 202 patients from the South Carolina Autism Study. The incidence of the duplication was not significantly different between patients and controls. Three of the female patients inherited the duplication from nonautistic fathers. In addition, there was no systematic skewing of X inactivation in the female patients with the duplication, or in nonautistic mothers and sisters with the duplication. These findings suggest that the dodecamer duplication in the HOPA gene does not play a significant role in the etiology of autism.

DCAMKL1, a brain-specific transmembrane protein on 13q12.3 that is similar to doublecortin (DCX).

Mutations in the human doublecortin (DCX), a brain-specific putative signaling protein, cause X-linked lissencephaly and subcortical band heterotopia. A predicted 740-amino-acid protein from human brain has two distinct regions, an N-terminal 345-amino-acid region 78% similar to the DCX protein and a C-terminal 427-amino-acid region that contains two transmembrane domains and is 98% homologous to a rat Ca2+/calmodulin-dependent protein kinase. We have designated this protein DCAMKL1. It maps to chromosome 13q12.3-q13, within a 540-kb YAC clone containing markers D13S805 and D13S1164. Northern analysis detected three major transcript isoforms of the DCAMKL1 gene expressed differentially and predominantly in human fetal and adult brain and during mouse embryogenesis (11-17 dpc). These results and its homology with the DCX and Ca2+/calmodulin dependent kinase proteins suggest a likely role for DCAMKL1 transmembrane protein in developing and adult brain, possibly in a pathway of cortical development.

Molecular cloning and characterization of TRPC5 (HTRP5), the human homologue of a mouse brain receptor-activated capacitative Ca2+ entry channel.

A novel human gene, TRPC5, was cloned from the region of Xq23 that contains loci for nonsyndromic mental retardation (MRX47 and MRX35) and two genes, DCX and HPAK3, implicated in two X-linked disorders (LISX and MRX30). Within a single YAC, we have determined the order cen-HPAK3(5'-3')-DCX(3'-5')-DXS7012E-TRPC5(3'-5' )-ter. TRPC5 encodes a 974-residue novel human protein (111.5 kDa predicted mass) and displays 99% homology with mouse TRP5, (MGD-approved symbol Trrp5) a novel member of a family of receptor-activated Ca2+ channels. It contains eight transmembrane domains, including a putative pore region. A transcript larger than 9.5 kb is observed only in fetal and adult human brain, with a relatively higher level in the adult human cerebellum. We devised an efficient method, Incorporation PCR SSCP (IPS), for detection of gene alterations. Five single-nucleotide variations in the TRPC5 gene were identified in males with mental retardation. However, these were found to be polymorphic variants. Exclusive expression of the TRPC5 gene in developing and adult brain suggests a possible role during development and provides a candidate gene for instances of mental retardation and other developmental defects.

Human doublecortin (DCX) and the homologous gene in mouse encode a putative Ca2+-dependent signaling protein which is mutated in human X-linked neuronal migration defects.

Subcortical band heterotopia (SBH) and classical lissencephaly (LIS) result from deficient neuronal migration which causes mental retardation and epilepsy. A single LIS/SBH locus on Xq22.3-q24 was mapped by linkage analysis and physical mapping of the breakpoint in an X;2 translocation. A recently identified gene, doublecortin ( DCX ), is expressed in fetal brain and mutated in LIS/SBH patients. We have identified four novel missense mutations in the gene, one familial mutation with LIS in a male and SBH in the carrier females, one de novo mutation in an SBH female, and two mutations in sporadic SBH female patients. The DCX gene is found to be expressed exclusively at a very high level in the adult frontal lobe. We have also cloned the X-linked mouse doublecortin (Dcx) gene. It encodes isoforms of a highly hydrophilic 40 kDa protein, homologous to its human counterpart and containing several potential phosphorylation sites. Both human and mouse DCX proteins are homologous to a CNS protein containing a Ca2+/calmodulin kinase domain, suggesting that the DCX protein may belong to a novel class of intracellular proteins involved in neuronal migration through Ca2+-dependent signaling.

Integrated STS/YAC physical, genetic, and transcript map of human Xq21.3 to q23/q24 (DXS1203-DXS1059).

A map has been assembled that extends from the XY homology region in Xq21.3 to proximal Xq24, approximately 20 Mb, formatted with 200 STSs that include 25 dinucleotide repeat polymorphic markers and more than 80 expressed sequences including 30 genes. New genes HTRP5, CAPN6, STPK, 14-3-3PKR, and CALM1 and previously known genes including BTK, DDP, GLA, PLP, COL4A5, COL4A6, PAK3, and DCX are localized; candidate loci for other disorders for which genes have not yet been identified, including DFN-2, POF, megalocornea, and syndromic and nonsyndromic mental retardation, are also mapped in the region. The telomeric end of the contig overlaps a yeast artificial chromosome (YAC) contig from Xq24-q26 and with other previously published contigs provides complete sequence-tagged site (STS)/YAC-based coverage of the long arm of the X chromosome. The order of published landmark loci in genetic and radiation hybrid maps is in general agreement. Combined with high-density STS landmarks, the multiple YAC clone coverage and integrated genetic, radiation hybrid, and transcript map provide resources to further disease gene searches and sequencing.

Evidence that a dodecamer duplication in the gene HOPA in Xq13 is not associated with mental retardation.

A recent study suggested that a dodecamer duplication in exon 42 of the HOPA gene in Xq13 may be a significant factor in the etiology of X-linked mental retardation. In an effort to investigate this possibility, we determined the incidence of the dodecamer duplication in cohorts of non-fragile X males with mental retardation from three countries, cohorts of fragile X males from two countries, 43 probands from families with X-linked mental retardation and control cohorts from three countries. The duplication was found in 3.6-4.0% of male patients from two non-fragile X groups (Italy and South Carolina), in 1.2% from another non-fragile X group (South Africa), but in no male patients from families with X-linked mental retardation (South Carolina). The dodecamer duplication was also found in several white males with fragile X syndrome from France (5%) and South Africa (22.2%). Additionally, the duplication was found in 1.5% of South Carolinian newborn males, 2.5% South Carolinian male college students, 5% Italian male controls and 4.5% of the white South African controls. None of the black South African non-fragile X individuals with mental retardation, the fragile X or the control samples tested carried the duplication, suggesting that the duplication is rare in the black South African population. The incidence of the duplication was not significantly different between any of the groups in the study. Therefore, results of our studies in four different populations do not corroborate the findings of the previous study, and indicate that the HOPA dodecamer duplication does not convey an increased susceptibility to mental retardation.