Assignment by deletion mapping of the steroid sulfatase X-linked ichthyosis locus to Xp223

Summary A male child and his mother who are nullisomic and monosomic, respectively, for the distal portion of Xp because of an unbalanced X-Y translocation were tested for steroid sulfatase activity after clinical examination had yielded evidence for ichthyosis in the boy. Deficiency of steroid sulfatase was found in the male patient, while in his mother enzyme levels were in the heterozygous range. These results, based on cytogenetic evidence obtained with an elongation technique, indicate that the STS locus is at Xp223.     

Multipoint linkage analysis of steroid sulfatase (X-linked ichthyosis) and distal Xp markers

Six families with steroid sulfatase deficiency (STS; X-linked ichthyosis) have been studied with the Xg blood group (XG) and the DNA markers dic56 (DXS143), 782 (DXS85), pD2 (DXS43), and GMGX9. Carrier status of females was determined by assay of STS in hair roots. GMGX9 detects a frequent restriction fragment length polymorphism and also identifies a deletion in the majority of families with STS deficiency, including five of the six reported here. The linkage relationship of this marker to the others was studied in normal three-generation families yielding 32 phase-known meioses informative for two or more markers. No recombinants were observed between STS and GMGX9, giving a maximum lod score of 8.73 at zero recombination. Multipoint linkage analysis taking STS and GMGX9 as a single locus and incorporating two-point marker data and STS-XG data from published studies gave the map (Sequence: see text). This order was 2.4 times more likely than with (STS,GMGX9) and dic56 reversed and is supported by our findings in a male with steroid sulfatase deficiency due to a deletion of Xp22.3 which encompasses the XG locus. He is deleted for GMGX9 but shows normal hybridization to dic56 and 782.     

Xp21 contiguous gene syndromes: deletion quantitation with bivariate flow karyotyping allows mapping of patient breakpoints.

Bivariate flow karyotyping was used to estimate the deletion sizes for a series of patients with Xp21 contiguous gene syndromes. The deletion estimates were used to develop an approximate scale for the genomic map in Xp21. The bivariate flow karyotype results were compared with clinical and molecular genetic information on the extent of the patients' deletions, and these various types of data were consistent. The resulting map spans > 15 Mb, from the telomeric interval between DXS41 (99-6) and DXS68 (L1-4) to a position centromeric to the ornithine transcarbamylase locus. The deletion sizing was considered to be accurate to +/- 1 Mb. The map provides information on the relative localization of genes and markers within this region. For example, the map suggests that the adrenal hypoplasia congenita and glycerol kinase genes are physically close to each other, are within 1-2 Mb of the telomeric end of the Duchenne muscular dystrophy (DMD) gene, and are nearer to the DMD locus than to the more distal marker DXS28 (C7). Information of this type is useful in developing genomic strategies for positional cloning in Xp21. These investigations demonstrate that the DNA from patients with Xp21 contiguous gene syndromes can be valuable reagents, not only for ordering loci and markers but also for providing an approximate scale to the map of the Xp21 region surrounding DMD.     

Chromosome 7 long arm deletion breakpoints in preleukemia: mapping by pulsed field gel electrophoresis.

Chromosome 7 long arm deletions (7q-) are recurring chromosome abnormalities in leukemic bone marrow cells. In four patients we have previously localized the breakpoints in band 7q22 between the erythropoietin (EPO) and plasminogen activator inhibitor type 1 (PLANH1) genes that map 3 cM apart. The pro alpha 2(I) collagen (COL1A2, in band 7q22) and T cell receptor beta chain genes (TCRB, in band 7q35) have been found undeleted in one patient with an interstitial deletion. Pulsed field gel electrophoresis was used to map the breakpoints more accurately in two patients with a 7q- chromosome. The results suggested that lymphocytes and granulocytes give identical restriction patterns with several enzyme-probe combinations, and that a breakpoint possibly was within 195 kb of EPO in one patient but not in another. The gene order cen-COL1A2-EPO-breakpoint-tel was suggested but physical linkage between COL1A2 and EPO was not found. A new putative TCRB restriction fragment length polymorphism or inherited methylation site was detected.     

Physical mapping of translocation breakpoints in rye by means of synaptonemal complex analysis

A physical map including 40 translocation breakpoints has been constructed in rye by means of synaptonemal complex (SC) analysis of well-paired pachytene quadrivalents. The chromosome arms involved in such translocations were previously identified either from mitotic C-banding analysis or from the meiotic configurations observed in the progenies of crosses with a rye line having multiple chromosome rearrangements. The synaptonemal complexes formed by some translocation homozygotes were also analyzed, the relative pachytene SC length of their translocated chromosomes being compared to that observed in the corresponding translocation heterozygotes. In the translocations in which the position of the breakpoint could be well defined from mitotic C-banding analysis, a good correspondence between the relative position of the point showing partner exchange in the pachytene quadrivalents and the actual location of the breakpoint was established. It is concluded that the mapping of translocation breakpoints by SC analysis of pachytene quadrivalents provides a more accurate estimate of the position of the breakpoints than that obtained from mitotic C-banding analysis, due to the lack of evenly-distributed interstitial C-bands in most rye chromosomes. The distribution of the breakpoints along the chromosomes in relation to their spontaneous or induced origin is also discussed.     

Molecular characterization of deletion breakpoints in adults with 22q11 deletion syndrome

22q11 Deletion syndrome (22q11DS) is a common microdeletion syndrome with variable expression, including congenital and later onset conditions such as schizophrenia. Most studies indicate that expression does not appear to be related to length of the deletion but there is limited information on the endpoints of even the common deletion breakpoint regions in adults. We used a real-time quantitative PCR (qPCR) approach to fine map 22q11.2 deletions in 44 adults with 22q11DS, 22 with schizophrenia (SZ; 12 M, 10 F; mean age 35.7 SD 8.0 years) and 22 with no history of psychosis (NP; 8 M, 14 F; mean age 27.1 SD 8.6 years). QPCR data were consistent with clinical FISH results using the TUPLE1 or N25 probes. Two subjects (one SZ, one NP) negative for clinical FISH had atypical 22q11.2 deletions confirmed by FISH using the RP11-138C22 probe. Most (n = 34; 18 SZ, 16 NP) subjects shared a common 3 Mb hemizygous 22q11.2 deletion. However, eight subjects showed breakpoint variability: a more telomeric proximal breakpoint (n = 2), or more centromeric (n = 3) or more telomeric distal breakpoint (n = 3). One NP subject had a proximal nested 1.4 Mb deletion. COMT and TBX1 were deleted in all 44 subjects, and PRODH in 40 subjects (19 SZ, 21 NP). The results delineate proximal and distal breakpoint variants in 22q11DS. Neither deletion extent nor PRODH haploinsufficiency appeared to explain the clinical expression of schizophrenia in the present study. Further studies are needed to elucidate the molecular basis of schizophrenia and clinical heterogeneity in 22q11DS.     

An Xp22 microdeletion associated with ocular albinism and ichthyosis: approximation of breakpoints and estimation of deletion size by using cloned DNA probes and flow cytometry.

Ocular albinism of the Nettleship-Falls type (OA1) and X-linked ichthyosis (XI) due to steroid sulfatase (STS) deficiency are cosegregating in three cytogenetically normal half-brothers. The mother has patchy fundal hypopigmentation consistent with random X inactivation in an OA1 carrier. Additional phenotypic abnormalities that have been observed in other STS "deletion syndromes" are not present in this family. STS is entirely deleted on Southern blot in the affected males, but the loci MIC2X, DXS31, DXS143, DXS85, DXS43, DXS9, and DXS41 are not deleted. At least part of DXS278 is retained. Flow cytometric analysis of cultured lymphoblasts from one of the XI/OA1 males and his mother detected a deletion of about 3.5 million bp or about 2% of the X chromosome. Southern blot and RFLP analysis in the XI/OA1 family support the order tel-[STS-OA1-DXS278]-DXS9-DXS41-cen. An unrelated patient with the karyotype 46,X,t(X;Y) (p22;q11) retains the DXS143 locus on the derivative X chromosome but loses DXS278, suggesting that DXS278 is the more distal locus and is close to an XI/OA1 deletion boundary. If a contiguous gene deletion is responsible for the observed XI/OA1 phenotype, it localizes OA1 to the Xp22.3 region.     

Linkage studies in a family with X-linked recessive ichthyosis employing a cloned DNA sequence from the distal short arm of the X chromosome

Recently linkage has been described between the Duchenne muscular dystrophy (DMD) gene and a cloned DNA sequence, RC8, that detects restriction fragment length polymorphism and is derived from the distal short arm of the X chromosome. Positive lod scores between RC8 and Xg prompted us to examine the linkage relationship of RC8 to the steroid sulfatase-X-linked recessive ichthyosis (XRI) locus which is situated 15 cM proximal from Xg in the subtelomeric region of Xp. Unexpectedly, at least two crossovers were found among nine informative meioses of an informative family, suggesting that RC8 and XRI may be about 25 cM apart. This implies that the genetic distance between the Xg locus and the DMD locus may exceed 50 cM.     

Molecular mapping and cloning of the breakpoints of a chromosome 11p14.1-p13 deletion associated with the AGR syndrome

Chromosome 11p13 is frequently rearranged in individuals with the WAGR syndrome (Wilms tumor, aniridia, genitourinary anomalies, and mental retardation) or parts of this syndrome. To map the cytogenetic aberrations molecularly, we screened DNA from cell lines with known WAGR-related chromosome abnormalities for rearrangements with pulsed field gel (PFG) analysis using probes deleted from one chromosome 11 homolog of a WAGR patient. The first alteration was detected in a cell line from an individual with aniridia, genitourinary anomalies, mental retardation, and a deletion described as 11p14.1-p13. We have located one breakpoint close to probe HU11-164B and we have cloned both breakpoint sites as well as the junctional fragment. The breakpoints subdivide current intervals on the genetic map, and the probes for both sides will serve as important additional markers for a long-range restriction map of this region. Further characterization and sequencing of the breakpoints may yield insight into the mechanisms by which these deletions occur.     

Role of cholesterol sulfate in epidermal structure and function: Lessons from X-linked ichthyosis

X-linked ichthyosis is a relatively common syndromic form of ichthyosis most often due to deletions in the gene encoding the microsomal enzyme, steroid sulfatase, located on the short area of the X chromosome. Syndromic features are mild or unapparent unless contiguous genes are affected. In normal epidermis, cholesterol sulfate is generated by cholesterol sulfotransferase (SULT2B1b), but desulfated in the outer epidermis, together forming a ‘cholesterol sulfate cycle’ that potently regulates epidermal differentiation, barrier function and desquamation. In XLI, cholesterol sulfate levels my exceed 10% of total lipid mass (≈ 1% of total weight). Multiple cellular and biochemical processes contribute to the pathogenesis of the barrier abnormality and scaling phenotype in XLI. This article is part of a Special Issue entitled The Important Role of Lipids in the Epidermis and their Role in the Formation and Maintenance of the Cutaneous Barrier. Guest Editors: Kenneth R. Feingold and Peter Elias.     

Physical mapping of translocation breakpoints in a set of wheat-Aegilops umbellulata recombinant lines using in situ hybridization

Aegilops umbellulata Zhuk. carries genes at Glu-U1 loci that code for a pair of high-molecular-weight glutenin subunits not found in common wheat, Triticum aestivum. Wheat-Ae. umbellulata recombinant lines were produced with the aim of transferring genes coding for glutenin subunits from Ae. umbellulata into wheat with minimal flanking material. We used fluorescent genomic in situ hybridization to evaluate the extent of recombination and to map physically the translocation breakpoints on 11 wheat-Ae. umbellulata recombinant lines. In situ hybridization was able to identify alien material in wheat and showed breakpoints not only near the centromeres but also along chromosome arms. To characterize and identify chromosomes further, including deletions along the 1U chromosome, we used simultaneous multiple target in situ hybridization to localize a tandemly repeated DNA sequence (pSc119.2) and the 18S–25S and 5S rRNA genes. One line contained an Ae. umbellulata telocentric chromosome and another two had different terminal deletions, mostly with some wheat chromosome rearrangements. Although from six independent original crosses, the other eight lines included only two types of intercalary wheat-Ae. umbellulata recombination events. Five occurred at the 5S rRNA genes on the short arm of the Ae. umbellulata chromosome with a distal wheat-origin segment, and three breakpoints were proximal to the centromere in the long arm, so most of the long arm was of Ae. umbellulata origin. The results allow characterization of recombination events in the context of the karyotype. They also facilitate the design of crossing programmes to generate lines where smaller Ae. umbellulata chromosome segments are transferred to wheat with the potential to improve bread-making quality by incorporating novel glutenin subunits without undesirable linked genes.     

Xp22.3 interstitial deletion: A recognizable chromosomal abnormality encompassing VCX3A and STS genes in a patient with X-linked ichthyosis and mental retardation

X-linked ichthyosis is a genetic disorder affecting the skin and caused by a deficit in the steroid sulfatase enzyme (STS), often associated with a recurrent microdeletion at Xp22.31. Most of the STS deleted patients have X-linked ichthyosis as the only clinical feature and it is believed that patients with more complex disorders including mental retardation could be present as a result of contiguous gene deletion. In fact, VCX3A gene, a member of the VCX (variable charge, X chromosome) gene family, was previously proposed as the candidate gene for X-linked non-specific mental retardation in patients with X-linked ichthyosis.     

Physical and transcriptional mapping of the X-linked cleft palate and ankyloglossia (CPX) critical region

Cleft palate most commonly occurs as a sporadic multifactorial disorder with a clear but difficult to define genetic component. As a semi-dominant disorder, X-linked cleft palate (CPX) provides a useful model to investigate a congenital defect that is little influenced by non-genetic factors. By using an Icelandic kindred, CPX has been localised between DXS1196 and DXS1217 and mapped, in a 3-Mb yeast artificial chromosome contig, at Xq21.3. Markers generated from this physical map have now been used to construct a contig of P1 and bacterial artificial chromosome clones for genomic DNA sequencing. Genomic DNA sequence analysis has revealed two novel expressed genes and two pseudogenes in the order Cen-KLHL4-LAMRL5-CAPZA1P-CPXCR1-Tel. KLHL4 and CPXCR1 are widely expressed in fetal tissues, including the tongue, mandible and palate. DNA mutation screening of CPXCR1 has revealed several sequence variants present on all affected CPX chromosomes. However, these variants have also been detected at a lower frequency on unaffected chromosomes, indicating that they are polymorphisms that are unlikely to cause the CPX phenotype.     

Intrachromosomal rearrangements in avian genome evolution: evidence for regions prone to breakpoints

It is generally believed that the organization of avian genomes remains highly conserved in evolution as chromosome number is constant and comparative chromosome painting demonstrated there to be very few interchromosomal rearrangements. The recent sequencing of the zebra finch (Taeniopygia guttata) genome allowed an assessment of the number of intrachromosomal rearrangements between it and the chicken (Gallus gallus) genome, revealing a surprisingly high number of intrachromosomal rearrangements. With the publication of the turkey (Meleagris gallopavo) genome it has become possible to describe intrachromosomal rearrangements between these three important avian species, gain insight into the direction of evolutionary change and assess whether breakpoint regions are reused in birds. To this end, we aligned entire chromosomes between chicken, turkey and zebra finch, identifying syntenic blocks of at least 250 kb. Potential optimal pathways of rearrangements between each of the three genomes were determined, as was a potential Galliform ancestral organization. From this, our data suggest that around one-third of chromosomal breakpoint regions may recur during avian evolution, with 10% of breakpoints apparently recurring in different lineages. This agrees with our previous hypothesis that mechanisms of genome evolution are driven by hotspots of non-allelic homologous recombination.     

High frequency of mosaic CREBBP deletions in Rubinstein-Taybi syndrome patients and mapping of somatic and germ-line breakpoints

Rubinstein-Taybi syndrome (RSTS) is a rare malformation disorder caused by mutations in the closely related CREBBP and EP300 genes, accounting respectively for up to 60 and 3% of cases. About 10% of CREBBP mutations are whole gene deletions often extending into flanking regions. Using FISH and microsatellite analyses as a first step in the CREBBP mutation screening of 42 Italian RSTS patients, we identified six deletions, three of which were in a mosaic condition that has not been previously reported in RSTS. The use of region-specific BAC clones and small CREBBP probes allowed us to assess the extent of all of the deletions by mapping their endpoints to genomic intervals of 5-10 kb. Four of our five intragenic breakpoints cluster at the 5' end of CREBBP, where there is a peak of breakpoints underlying rearrangements in RSTS patients and tumors. The search for genomic motifs did not reveal any low-copy repeats (LCRs) or any greater density of repetitive sequences. In contrast, the percentage of interspersed repetitive elements (mainly Alu and LINEs in the CREBBP exon 2 region) is significantly higher than that in the entire gene or the average in the genome, thus suggesting that this characteristic may be involved in the region's vulnerability to breaking and nonhomologous pairing. The FISH analysis extended to the EP300 genomic region did not reveal any deletions. The clinical presentation was typical in all cases, but more severe in the three patients carrying constitutional deletions, raising a question about the possible underdiagnosis of a few cases of mild RSTS.     

Memory,mood and associated neuroanatomy in individuals with steroid sulphatase deficiency (X-linked ichthyosis)

Steroid sulphatase (STS) cleaves sulphate groups from steroid hormones, and steroid (sulphate) levels correlate with mood and age-related cognitive decline. In animals, STS inhibition or deletion of the associated gene, enhances memory/neuroprotection and alters hippocampal neurochemistry. Little is known about the consequences of constitutive STS deficiency on memory-related processes in humans. We investigated self-reported memory performance (Multifactorial Memory Questionnaire), word-picture recall and recent mood (Kessler Psychological Distress Scale, K10) in adult males with STS deficiency diagnosed with the dermatological condition X-linked ichthyosis (XLI; n = 41) and in adult female carriers of XLI-associated genetic variants (n = 79); we compared results to those obtained from matched control subjects [diagnosed with ichthyosis vulgaris (IV, n = 98) or recruited from the general population (n = 250)]. Using the UK Biobank, we compared mood/memory-related neuroanatomy in carriers of genetic deletions encompassing STS (n = 28) and non-carriers (n = 34,522). We found poorer word-picture recall and lower perceived memory abilities in males with XLI and female carriers compared with control groups. XLI-associated variant carriers and individuals with IV reported more adverse mood symptoms, reduced memory contentment and greater use of memory aids, compared with general population controls. Mood and memory findings appeared largely independent. Neuroanatomical analysis only indicated a nominally-significantly larger molecular layer in the right hippocampal body of deletion carriers relative to non-carriers. In humans, constitutive STS deficiency appears associated with mood-independent impairments in memory but not with large effects on underlying brain structure; the mediating psychobiological mechanisms might be explored further in individuals with XLI and in new mammalian models lacking STS developmentally.