Identification and characterization of an Xq26-q27 duplication in a family with spina bifida and panhypopituitarism suggests the involvement of two distinct genes.

We investigated a family with a duplication, dup(X)q26-q27, that was present in two brothers, their mother, and their maternal grandmother. The brothers carrying the duplication displayed spina bifida and panhypopituitarism, whereas a third healthy brother inherited the normal X chromosome. Preferential inactivation of the X chromosome containing the duplication was evident in healthy carrier females. We determined the boundaries of the Xq26-q27 duplication. Via interphase FISH analysis we narrowed down each of the two breakpoint regions to approximately 300-kb intervals. The proximal breakpoint is located in Xq26.1 between DXS1114 and HPRT and is contained in YAC yWXD599, while the distal breakpoint is located in Xq27.3 between DXS369 and DXS1200 and contained in YAC yWXD758. The duplication comprises about 13 Mb. Evidence from the literature points to a predisposing gene for spina bifida in Xq27. We hypothesize that the spina bifida in the two brothers may be due to interruption of a critical gene in the Xq27 breakpoint region. Several candidate genes were mapped to the Xq27 critical region but none was shown to be disrupted by the duplication event. Recently, M. Lagerström-Fermér et al. (1997, Am. J. Hum. Genet. 60, 910-916) reported on a family with X-linked recessive panhypopituitarism associated with a duplication in Xq26; however, no details were reported on the extent of the duplication. Our study corroborates their hypothesis that X-linked recessive panhypopituitarism is likely to be caused by a gene encoding a dosage-sensitive protein involved in pituitary development. We place the putative gene between DXS1114 and DXS1200, corresponding to the interval defined by the duplication in the present family.

Genetic dissection of dome formation in a mammary cell line: identification of two genes with opposing action.

In this work, we extend the study of the genes controlling the formation of domes in the rat mammary cell line LA7 under the influence of DMSO. The role of the rat8 gene has already been demonstrated. We have now studied two additional genes. The first, called 133, is the rat ortholog of the human epithelial membrane protein 3 (EMP3), a member of the peripheral myelin protein 22 (PMP22)/EMP/lens-specific membrane protein 20 (MP20) gene family that encodes for tetratransmembrane proteins; it is expressed in the LA7 line in the absence of DMSO but not in its presence. The second gene is the beta subunit of the amiloride-sensitive Na(+) channel. Studies with antisense oligonucleotides show that the formation of domes is under the control of all three genes: the expression of rat8 is required for both their formation and their persistence; the expression of the Na(+) channel beta subunit is required for their formation; and the expression of gene 133 blocks the expression of the Na(+) channel genes, thus preventing formation of the domes. The formation of these structures is also accompanied by the expression of alpha(6)beta(1) integrin, followed by that of E-cadherin and cytokeratin 8. It appears, therefore, that dome formation requires the activity of the Na(+) channel and the rat8-encoded protein and is under the negative control of gene 133. DMSO induces dome formation by blocking this control.

Transcription map of Xq27: candidates for several X-linked diseases.

Human Xq27 contains candidate regions for several disorders, yet is predicted to be a gene-poor cytogenetic band. We have developed a transcription map for the entire cytogenetic band to facilitate the identification of the relatively small number of expected candidate genes. Two approaches were taken to identify genes: (1) a group of 64 unique STSs that were generated during the physical mapping of the region were used in RT-PCR with RNA from human adult and fetal brain and (2) ESTs that have been broadly mapped to this region of the chromosome were finely mapped using a high-resolution yeast artificial chromosome contig. This combined approach identified four distinct regions of transcriptional activity within the Xq27 band. Among them is a region at the centromeric boundary that contains candidate regions for several rare developmental disorders (X-linked recessive hypoparathyroidism, thoracoabdominal syndrome, albinism-deafness syndrome, and Borjeson-Forssman-Lehman syndrome). Two transcriptionally active regions were identified in the center of Xq27 and include candidate regions for X-linked mental retardation syndrome 6, X-linked progressive cone dystrophy, X-linked retinitis pigmentosa 24, and a prostate cancer susceptibility locus. The fourth region of transcriptional activity encompasses the FMR1 (FRAXA) and FMR2 (FRAXE) genes. The analysis thus suggests clustered transcription in Xq27 and provides candidates for several heritable disorders for which the causative genes have not yet been found.

Mapping of the MYCL2 processed gene to Xq22-23 and identification of an additional L MYC-related sequence in Xq27.2.

We report here the identification of a human genomic sequence from the q27.2 region of the X chromosome which shows a high homology to the L-MYC proto-oncogene. This sequence is not the MYCL2 homology, previously mapped to the long arm of the X chromosome at q22-qter by Morton et al., as we located the MYCL2-processed gene in Xq22-23, using a panel containing a combination of hybrid DNA carrying different portions of the human X chromosome. Based on computer analysis, the MYC-like sequence (MYCL3) is 98.2% identical to a portion of exon 3 of the MYCL1 gene and maps to the Xq27.2 region, between the DXS312 and DXS292 loci.

Identification and genomic organization of a gene coding for a new member of the cell adhesion molecule family mapping to Xq25.

The gene coding for a new member of the Immunoglobulin (Ig)-like domain-containing molecule superfamily has been identified and mapped to the human Xq25 chromosomal band. It contains 12 Ig-like domains in two clusters of 5 and 7 motifs, respectively, separated by a linker segment, followed by a transmembrane and a cytoplasmic region. The gene is conserved in mammals and is expressed in muscle, heart, brain, testis, and pancreas with transcripts of different length, suggesting that it is subjected to alternative processing. The transcript is assembled from 19 exons which are distributed along approx. 20kb; each Ig-like domain is contained in distinct exons which constitute the unit of repeated genomic duplications. Elucidation of the IGDC1 genomic structure will allow the investigation of the basis of its alternative transcription and of its possible involvement in diseases mapped to the Xq25 interval.

Identification of CXorf1, a novel intronless gene in Xq27.3, expressed in human hippocampus.

We have identified and characterized a novel human gene (Nomenclature Committee of the Genome Database GDB-assigned symbol CXorf1) that maps to the long arm of the X chromosome in Xq27 between loci DXS369 and DXS181, approximately 2.5 Mb centromeric to the FMR1 gene. The CXorf1 gene is conserved in primates, cow, and horse but not in mouse and rat. Northern blot analysis revealed two transcripts, present in the brain and in the G361 melanoma cell line. In situ hybridization experiments performed on sections of human hippocampus showed a clear, uneven localization of the CXorf1 mRNA in specific subfields of this brain area. In particular, CXorf1 was localized in the granular-cell layer of the dentate gyrus and in the CA2-CA3 subfields of Ammon's horn. CXorf1 is one of the first genes from this region to be characterized in detail and, on the basis of its chromosomal location and expression pattern, may have an important function in the brain.

Genomic organization of the human VP16 accessory protein, a housekeeping gene (HCFC1) mapping to Xq28.

The region between DXS52 and Factor VIII gene in the human Xq28 chromosomal band contains a G+C-rich isochore to which many genes have been mapped. We report here the isolation and characterization of a transcript mapping about 50 kb telomeric from the vasopressin type 2 receptor gene in a 180-kb YACs/cosmid contig containing the L1CAM gene at its centromeric end. The determined transcribed sequence from a human fetal brain library is identical to that of the recently identified accessory protein HCFC1 (host cell factor, also called C1) that activates herpes simplex virus VP16 (alpha TIF) transactivator protein for association with the octamer motif-binding protein Oct-1 (Cell 74: 115, 1993). The gene is expressed in a ubiquitous pattern and a larger transcript of approximately 10 kb is present in all the tissues tested, while an alternatively spliced RNA of approximately 8.0 kb is present in muscle and heart tissues. Genomic sequencing allowed us to determine that the sequenced transcript is assembled from 26 exons spread over a relatively small genomic region of approximately 24 kb. This alllowed us to determine that a previously reported cDNA clone arises from the splicing out of an internal portion of exon 8 which does not change the reading frame. All together these results raise the possibility that alternative mRNA processing could partly contribute to the diversity of the polypeptide HCFC1 family in a subset of tissues.

Isolation of a new gene in the Friedreich ataxia candidate region on human chromosome 9 by cDNA direct selection.

The Friedreich ataxia (FRDA) locus is localized on chromosome 9q13 in an interval less than 1 Mb between markers D9S202/FR1 and FR5. We cloned the FRDA candidate region in YACs, and we started a systematic search for transcripts in this region using the cDNA selection approach. Several overlapping cDNA clones mapping near the telomeric end of the FRDA minimum genetic region were isolated. Zoo blot analysis demonstrated that these cDNAs are well conserved among different species. A transcript of 4.8 kb was identified by hybridization to a Northern blot containing human brain poly(A)+ RNA. Partial sequence of these clones showed 100% homology with a previously described anonymous brain cDNA (EST01251). A search for mutations of this gene in FRDA patients and carriers is in progress. No mutations have been found to date, but we have identified a DNA polymorphism. This polymorphism was nonrecombinant with the disease in a previously described FRDA pedigree in which a recombination had occurred with more telomeric markers.

Linkage disequilibrium between FD1-D9S202 haplotypes and the Friedreich's ataxia locus in a central-southern Italian population.

We used two recently described genetic markers in the region of the Friedreich's ataxia locus to study 33 affected pedigrees from central-southern regions of Italy. These markers are predicted, by physical mapping, to be localised more closely to the Friedreich's ataxia locus than other previously described markers. No recombination was found between these markers and the disease locus. Strong linkage disequilibrium is present between the compound haplotype and the disease locus. Since this population was also previously studied by using three other more distal genetic markers, a total of five markers has been used to identify the extended haplotype. Homozygosity in consanguineous pedigrees was also studied. Extended haplotype analysis and homozygosity studies suggest the presence of few common disease causing mutations in our population.

Type 2 vasopressin receptor gene, the gene responsible nephrogenic diabetes insipidus, maps to Xq28 close to the LICAM gene.

Although long contigs have been assembled in Xq28, the interval between the anonymous probe St14 and the color vision locus is still incompletely defined. We report here that the recently cloned gene for type 2 vasopressin receptor (V2R) is physically linked to L1CAM using YACs and cosmids across about 180 kb of the region. Since it is known that L1CAM maps near the color pigment genes, this finding locates V2R in Xq28 in the area where nephrogenic diabetes insipidus (NDI) has been mapped by linkage analysis. The PFGE analysis of the clones positions V2R about 40 kb from the L1CAM gene in a region that appears to contain other unknown genes, since at least four putative CpG islands were identified by restriction analysis with rare cutter enzymes.

Isolation of a zinc finger motif (ZNF75) mapping on chromosome Xq26.

We report here the partial characterization of a new human zinc finger (ZNF75) gene of the Kruppel type mapping to the long arm of the X chromosome. A cosmid clone was isolated from a library specific to the Xq24-qter region by hybridization to a degenerate oligonucleotide representing the link between two contigous fingers of the C2H2 type. The sequence of the pertinent cosmid fragments demonstrated five consecutive zinc finger motifs, all pertaining to the Kruppel family. A reading frame starting at least 75 amino acids before the first zinc finger and ending 11 amino acids after the last one was identified; comparison with other ZF genes suggests that this genomic fragment represents the carboxy-terminal exon of the gene. Homology of approximately 55% in the zinc finger region was detected with many zinc finger genes including mouse Zfp-35 and human ZFN7 cDNA clones. Mapping using a panel of sematic cell hybrids and chromosomal in situ hybridization localized the gene to Xq26, in a region not previously known to contain zinc finger genes.