PCR, Southern blotting and oocyte expression to assess the efficacy of subtractive hybridization.
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Animals↗
Biomedical subjects
S Somlo
Publications and source records attributed to S Somlo.
Evidence for a third genetic locus for autosomal dominant polycystic kidney disease.
Autosomal dominant polycystic kidney disease (ADPKD) is a genetically heterogeneous disease with loci on chromosomes 16p and 4q. It has a moderately high spontaneous mutation rate, although the relative frequency of such mutations at each gene locus is unknown. In studying genetic heterogeneity in the French-Canadian population, we identified a family in which a classical clinical presentation of ADPKD resulted from a mutation at a locus genetically distinct from either of the previously described loci for this disease. This suggests the existence of a third genetic locus for ADPKD.
Aged↗
Linkage disequilibrium in the region of the autosomal dominant polycystic kidney disease gene (PKD1).
The gene for autosomal dominant polycystic kidney disease (PKD1) is located on chromosome 16p, between the flanking markers D16S84 and D16S125 (26.6prox). This region is 750 kb long and has been cloned. We have looked at the association of 10 polymorphic markers from the region, with the disease and with each other. This was done in a set of Scottish families that had previously shown association with D16S94, a marker proximal to the PKD1 region. We report significant association between two CA repeat markers and the disease but have not found evidence for a single founder haplotype in these families, indicating the presence of several mutations in this population. Our results favor a location of the PKD1 gene in the proximal part of the candidate region.
Alleles↗
A transducin-like gene maps to the autosomal dominant polycystic kidney disease gene region.
A novel human gene (sazD) that maps to the autosomal dominant polycystic kidney disease region shares sequence similarity with members of the beta-transducin superfamily. The cDNA sazD-c predicts an approximately 58-kDa protein (sazD) with seven internal repeats, similar to the WD-40 motif of the transducin family. The size of this protein family has been expanding rapidly; however, neither the structure nor the function of this repeated motif is known. Preliminary data do not suggest that sazD is mutated in patients with polycystic kidney disease.
Adult↗
Autosomal dominant polycystic kidney disease: localization of the second gene to chromosome 4q13-q23.
At least two loci are known to exist for autosomal dominant polycystic kidney disease (ADPKD). One was localized to 16p, but the second less common locus has remained unlinked. Over 100 microsatellite markers, distributed across all chromosomes, have been typed on informative family members from the large Sicilian kindred in which the genetic heterogeneity was first discovered. Both the affected and the unaffected status of every family member used in the study were confirmed by renal ultrasonography. This search has resulted in the successful localization of a second ADPKD gene to chromosome 4q. It was found to be flanked by the markers D4S231 and D4S414, defining a segment that spans about 9 cM. The new locus has been designated PKD4. This second localization will allow researchers to target another ADPKD gene for isolation in an effort to understand the pathogenesis of this common disorder. Furthermore, when flanking markers for the second ADPKD gene are used in conjunction with flanking markers for PKD1, the accuracy of the diagnosis of the subtype of ADPKD present in any particular family will be enhanced. This will improve the accuracy of linkage-based presymptomatic diagnoses by reducing the error due to genetic heterogeneity.
Chromosome Mapping↗
Inherited diseases of the kidney.
It has long been known that a number of diseases affecting the kidney are the result of genetic defects passed on through the generations. Whereas some of these defects are rare, others, eg, the cystic diseases, are among the most common. Our understanding of the underlying pathobiology in these disorders based on physiologic and cell biologic studies is variable--we suspect that the V2 vasopressin receptor is defective in nephrogenic diabetes insipidus; we know that the glomerular basement membrane in Alport syndrome is abnormal; we suspect that a tumor suppressor gene is defective in Wilms tumor; and we lack a unifying hypothesis regarding cystic degeneration of the kidney. The advent and rapid progress of molecular biology have permitted an entirely new approach to understanding these diseases, allowing the expected identification of mutations in the V2 receptor, the unexpected finding that a novel collagen gene is responsible for many Alport syndrome cases, and the somewhat less-unexpected finding that only one of several genes responsible for renal cancers has been identified. Further, we are beginning to unravel the complex pathways responsible for cystic changes in the kidney. This review integrates these molecular biologic discoveries with the known pathobiology of disease to achieve a more complete understanding of the whole process.
Animals↗
A kindred exhibiting cosegregation of an overlap connective tissue disorder and the chromosome 16 linked form of autosomal dominant polycystic kidney disease.
Autosomal dominant polycystic kidney disease (ADPKD) is a disorder of adult onset manifested by bilaterally enlarged cystic kidneys frequently associated with progressive renal failure. The mutated gene (PKD1) responsible for 85 to 95% of cases has been localized to a small segment on the distal tip of the short arm of chromosome 16. A clinical spectrum of heritable connective tissue disorders that remain unclassifiable under the present nosology but that contain elements of the Marfan's syndrome have previously been described. The genetic localization and molecular basis of such overlap connective tissue disorders (OCTD) have not been elucidated. In this report, a kindred in which ADPKD and OCTD appear to cosegregate is described. The connective tissue phenotype in this family includes aortic root dilation, aortic and vertebral artery aneurysms with dissection, and aortic valve incompetence, as well as pectus abnormalities, pes planus, joint laxity, arachnodactyly, scoliosis, dolichostenomelia, and high arched palate. ADPKD was manifest primarily as bilateral renal cysts with or without renal failure. The DNA of all living family members was studied with markers recognizing polymorphic loci flanking the PKD1 region (3'HVR and O90a), as well as markers from the loci of chromosomes 15 and 5, associated with fibrillin genes FBN1 and FBN2, respectively. In this kindred of 20 family members traced through five generations, cosegregation of ADPKD and the OCTD phenotype was observed in 12 of 12 meioses and 3 of 3 phase known. Both markers for PKD1 were tightly linked to both ADPKD and OCTD, whereas there was no evidence for linkage with either fibrillin locus. In this family, the ADPKD and OCTD mutations are genetically linked. The presence of OCTD with ADPKD identifies a group of patients at significantly greater risk for sudden death from aortic root and other vascular aneurysmal dissection and rupture.
Adolescent↗
Positional cloning approach to the dominant polycystic kidney disease gene, PKD1.
Positional cloning is a powerful strategy for identifying the site of disease-producing mutations when the underlying biochemical defect is unknown. The approach also offers new methods for the presymptomatic diagnosis of genetic disease. Using these methods we have localized the PKD1 gene, mutated in the majority of PKD1 families, to a small (500 kb) segment of chromosome 16, band p13.3. Virtually all of this interval has been cloned in cosmids and lambda bacteriophage. Over 20 sets of non-overlapping cDNA clones have been isolated from the region. Sequence and mutational analyses are currently underway. In addition, a set of polymorphic clones has been identified for presymptomatic diagnosis. Included in this set are several highly variable [CA]n microsatellite repeats. These highly informative markers can be rapidly assayed from a small amount of genomic DNA using the polymerase chain reaction. Despite these advances, presymptomatic diagnosis cannot be established with certainty in many families. However, identification of the PKD1 gene itself will eventually allow diagnosis by direct detection of mutations.
Base Sequence↗
Human-mouse homologies in the region of the polycystic kidney disease gene (PKD1).
Autosomal dominant polycystic kidney disease (PKD1) is linked to the alpha-globin locus near the telomere of chromosome 16p. We established the existence of a conserved linkage group in mouse by mapping conserved sequences and cDNAs from the region surrounding the PKD1 gene in the mouse genome. Results obtained with the BXD recombinant strain system and somatic cell hybrids show the homologous region to be located on mouse chromosome 17 near the globin pseudogene Hba-ps4, an unprocessed alpha-like globin gene. The markers we mapped are widely distributed over the region known to contain the PKD1 gene, and it is therefore likely that the mouse homologue of PKD1 is also located on mouse chromosome 17.
Animals↗
The gene for autosomal dominant polycystic kidney disease lies in a 750-kb CpG-rich region.
PKD1, the locus most commonly affected by mutations that produce autosomal dominant polycystic kidney disease (ADPKD), has previously been localized to chromosome 16p13.3. Since no cytogenetic abnormalities have been found in association with ADPKD, flanking genetic markers have been required to define an interval--the PKD1 region--that contains the PKD1 gene. In this report we demonstrate, through the construction of a long-range restriction map that links the flanking genetic markers GGG1 (D16S84) and 26.6PROX (D16S125), that the PKD1 gene lies within an extremely CpG-rich 750-kb segment of chromosome 16p13.3. Approximately 90% of this region has been cloned in three extensive cosmid/bacteriophage contigs. The cloned DNA is a valuable resource for identifying new closer flanking genetic markers and for isolating candidate genes from the region.
Chromosomes, Human, Pair 16↗
Fine genetic localization of the gene for autosomal dominant polycystic kidney disease (PKD1) with respect to physically mapped markers.
PKD1, the gene for the chromosome 16-linked form of autosomal dominant polycystic kidney disease, has previously been genetically mapped to an interval bounded by the polymorphic loci Fr3-42/EKMDA2 distally and O327hb/O90a proximally. More recently, 26.6PROX was identified as the closest proximal flanking locus. We set out to refine the localization of PKD1 by identifying a series of single recombinant events between the flanking markers Fr3-42/EKMDA2 and O327hb/O90a and analyzing them with a new set of polymorphic loci that have been physically mapped within the PKD1 interval. We identified 11 such crossovers in eight families; 6 of these fell into the interval between GGG1 and 26.6PROX, a distance of less than 750 kb. Three of these crossovers placed PKD1 proximal to GGG1 and two crossovers placed PKD1 distal to 26.6PROX. Both of the latter also placed PKD1 telomeric to a locus 92.6SH1.0, which lies 200-250 kb distal to 26.6PROX. The sixth recombinant, however, placed the disease mutation proximal to the locus 92.6SH1.0. Several possible explanations for these observations are discussed. An intensive study to locate deletions, insertions, and other chromosomal rearrangements associated with PKD1 mutations failed to detect any such abnormalities. Thus we have defined, in genetic and physical terms, the segment of 16p13.3 where PKD1 resides and conclude that a gene-by-gene analysis of the region will be necessary to identify the mutation(s).
Chromosome Mapping↗
The molecular genetics of autosomal-dominant polycystic kidney disease of the PKD1 type.
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Chromosome Mapping↗
A positional cloning approach to inherited renal disease.
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Amino Acid Sequence↗
CpG island in the region of an autosomal dominant polycystic kidney disease locus defines the 5' end of a gene encoding a putative proton channel.
In an attempt to isolate candidate genes for autosomal dominant polycystic kidney disease, a number of CpG-rich islands have been identified from a region defined genetically as the site of disease mutations. Genomic fragments adjacent to one of these islands were used to isolate cDNAs from both HeLa cells and cultured cystic epithelium that encode a 155-amino acid peptide having four putative transmembrane domains. The corresponding transcript was found in all tissues tested but was most abundant in brain and kidney. Potential control response elements were identified in the genomic region 5' to the initiation codon. The deduced amino acid sequence has 93% similarity to the 16-kDa proteolipid component that is believed to be part of the proton channel of the vacuolar H(+)-ATPase. Possible roles for a mutated proton channel in the pathogenesis of cystic disease were considered. However, sequencing of cDNAs corresponding to both alleles of an affected individual revealed no differences in the deduced amino acid sequence. Moreover, transcript size and abundance were not altered in cystic kidney.
Amino Acid Sequence↗
Cosmid walking and chromosome jumping in the region of PKD1 reveal a locus duplication and three CpG islands.
The locus responsible for the most common form of autosomal dominant polycystic kidney disease (PKD1) is located on chromosome 16p13.3. Genetic mapping studies indicate that PKD1 is flanked on the proximal side by the DNA marker 26.6 (D16S125). Here we show that 26.6 has undergone a locus duplication and that the two loci are less than 150kb apart. One of the two loci contains a polymorphic TaqI site that has been used in genetic studies and represents the proximal boundary for the PKD1 locus. We demonstrate that the polymorphic locus is the more proximal of the two 26.6-hybridizing loci. Therefore, four cosmids isolated from the distal 26.6-hybridizing locus contain candidate sequences for the PKD1 gene. These cosmids were found to contain two CpG islands that are likely markers for transcribed regions. A third CpG island was detected and cloned by directional chromosome jumping.
Chromosome Walking↗
The diagnosis and prognosis of autosomal dominant polycystic kidney disease.
BACKGROUND: Autosomal dominant polycystic kidney disease is usually caused by a mutant gene at the PKD1 locus on the short arm of chromosome 16, but in about 4 percent of families with the disorder it is caused by unknown mutations elsewhere in the genome. The natural course of the disease in both genetic forms is not well characterized. METHODS: We studied 17 families with autosomal dominant polycystic kidney disease to compare presymptomatic diagnosis by ultrasonography with diagnosis by genetic-linkage studies and to relate clinical variation of the disease to whether the PKD1 mutation was implicated. RESULTS: In 10 families the disorder was found to cosegregate with polymorphic DNA markers flanking the PKD1 locus, in 2 families it did not, and in 5 families linkage could not be determined. In the 10 families with the PKD1 mutation, 46 percent of the members less than 30 years old who had a 50 percent risk of inheriting a mutation had renal cysts, as compared with 11 percent of the members of the two families without linkage (P less than 0.001). In the PKD1 families, all 67 diagnoses made by ultrasonography were confirmed by determination of the genotype as inferred from linkage. Forty of 48 members (83 percent) less than 30 years old who inherited the PKD1 mutation had renal cysts. All 27 members 30 years old or older who inherited the mutation had renal cysts, suggesting that the probability of a false negative diagnosis did not exceed 0.13 in this age group (P less than 0.05). The mean (+/- SE) age at the onset of end-stage renal disease among members of the PKD1 families was 56.7 +/- 1.9 years, as compared with 69.4 +/- 1.7 years among members with cysts in the families without linkage (P = 0.0025). Hypertension and renal impairment were less frequent and occurred later in the families without the PKD1 mutation. CONCLUSIONS: At present, in most persons with a 50 percent risk of autosomal dominant polycystic kidney disease, imaging techniques are the only mode of reaching a diagnosis before symptoms appear. In such persons a negative ultrasonographic study during early adult life indicates that the likelihood of inheriting a PKD1 mutation is small. In the few who inherit a non-PKD1 mutation for polycystic kidney disease, renal failure is likely to occur relatively late in life.
Adult↗
Nerve growth cones isolated from fetal rat brain: subcellular fractionation and characterization.
The biochemical and functional characterization of the nerve growth cone is of major interest for studies on mechanisms involved in nervous system development. We describe the isolation from fetal brain of membrane-bound fragments of nerve growth cones by density gradient fractionation. These so-called growth cone particles are highly uniform and identifiable on the basis of their organelle complement. Furthermore, they co-purify in mixing experiments with fragments of radiolabeled and light microscopically identified nerve growth cones from primary cultures. The possibility of isolating growth cone fragments in quantity renders feasible the analysis of molecular mechanisms involved in growth cone function.
Animals↗