A deletion distinct from the classical homologous recombination of juvenile nephronophthisis type 1 (NPH1) allows exact molecular definition of deletion breakpoints

Betz, Regina C.; Rensing, Christopher; Schätzle, Silvia; Kuntzen, Thomas; Vetsi, Thalia; Imm, Anita; Hildebrandt, Friedhelm

doi:10.1002/1098-1004(200009)16:3<211::aid-humu4>3.0.co;2-y

Cited by 27 publications

(12 citation statements)

References 41 publications

(59 reference statements)

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“…2). None of these sequences are known to predispose to genomic rearrangements, although short stretches of identical sequence are often found at the sites of germline translocation and deletion breakpoints [20-22]. Genomic analysis of the parents and siblings of Case 2 indicated that the mutation was sporadic.…”

Section: Resultsmentioning

confidence: 99%

Multi-exon deletions of the FBN1gene in Marfan syndrome

et al. 2001

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BackgroundMutations in the fibrillin -1 gene (FBN1) cause Marfan syndrome (MFS), an autosomal dominant multi-system connective tissue disorder. The 200 different mutations reported in the 235 kb, 65 exon-containing gene include only one family with a genomic multi-exon deletion.MethodsWe used long-range RT-PCR for mutation detection and long-range genomic PCR and DNA sequencing for identification of deletion breakpoints, allele-specific transcript analyses to determine stability of the mutant RNA, and pulse-chase studies to quantitate fibrillin synthesis and extracellular matrix deposition in cultured fibroblasts. Southern blots of genomic DNA were probed with three overlapping fragments covering the FBN1 coding exonsResultsTwo novel multi-exon FBN1 deletions were discovered. Identical nucleotide pentamers were found at or near the intronic breakpoints. In a Case with classic MFS, an in-frame deletion of exons 42 and 43 removed the C-terminal 24 amino acids of the 5th LTBP (8-cysteine) domain and the adjacent 25th calcium-binding EGF-like (6-cysteine) domain. The mutant mRNA was stable, but fibrillin synthesis and matrix deposition were significantly reduced. A Case with severe childhood-onset MFS has a de novo deletion of exons 44–46 that removed three EGF-like domains. Fibrillin protein synthesis was normal, but matrix deposition was strikingly reduced. No genomic rearrangements were detected by Southern analysis of 18 unrelated MFS samples negative for FBN1 mutation screening.ConclusionsTwo novel deletion cases expand knowledge of mutational mechanisms and genotype/phenotype correlations of fibrillinopathies. Deletions or mutations affecting an LTBP domain may result in unstable mutant protein cleavage products that interfere with microfibril assembly.

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Section: Resultsmentioning

confidence: 99%

Multi-exon deletions of the FBN1gene in Marfan syndrome

et al. 2001

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“…The most notable structural feature is a Src homology 3 (SH3) 2 domain, a widespread protein-protein interaction module recognizing proline-rich motifs, which spans residues 157-207 of human nephrocystin. Another structural feature predicted from the nephrocystin amino acid sequence is a series of three 17-18-residue stretches, all within the N-terminal 105 amino acid residues, predicted to form short coiled-coil domains (7,8). The coiled-coil domain is an amphipathic ␣-helical heptad repeat recognized for its ability to mediate protein dimerization or oligomerization through hydrophobic interactions (9).…”

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confidence: 99%

“…In addition to human, cDNAs encoding full-length mouse nephrocystin have been described (7,8), as has a partial cDNA for canine nephrocystin (7). Comparison of the human and mouse sequences indicate that the N-terminal region of predicted coiled-coiled domains and the nearby SH3 domain are well conserved, with each showing 80% amino acid identity.…”

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confidence: 99%

“…However the C-terminal two-thirds of nephrocystin (human residues 228 -677 and mouse residues 241-691) 1 is even more highly conserved with 87% amino acid identity, indicating that this region of the protein also has an important function conserved through evolution. Protein sequence database searches reveal only one other protein with clear homology to the large conserved nephrocystin C-terminal region, a hypothetical Caenorhabditis elegans protein designated M28.7 that shows 23% overall identity and 42% similarity to the human sequence (8). Also conserved in M28.7 are the SH3 and putative coiled-coil domains further indicating it represents a distantly related nephrocystin ortholog.…”

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confidence: 99%

“…Expression studies indicate that the nephrocystin transcript is not uniquely expressed in the kidney but is widely detected in mouse embryos and in a variety of human and mouse adult tissues (7,8). Thus nephrocystin may function in many cell types, although the pathogenic consequence of loss-of-function is confined largely to the kidney.…”

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confidence: 99%

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Nephrocystin-conserved Domains Involved in Targeting to Epithelial Cell-Cell Junctions, Interaction with Filamins, and Establishing Cell Polarity

Donaldson¹,

Dise²,

Ritchie³

et al. 2002

Journal of Biological Chemistry

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Nephrocystin is the protein product of the gene mutated in juvenile nephronophthisis, an autosomal recessive cystic kidney disease afflicting children and young adults. Because the normal cellular function of nephrocystin is largely unknown, the molecular defects underlying disease pathogenesis remain obscure. Analysis of nephrocystin amino acid sequences from human and other species revealed three distinct conserved domains including Src homology 3 and coil-coil domains in the N-terminal region, as well as a large highly conserved C-terminal region bearing no obvious homology to other proteins and hence referred to as the "nephrocystin homology domain" (NHD). The objective of this study was to gain insight into nephrocystin function by defining functional properties of the conserved domains. We analyzed a series of nephrocystin deletion mutants expressed in Madin-Darby canine kidney and COS-7 cells. This analysis revealed previously unrecognized functional attributes of the NHD, including abilities to promote both self-association and epithelial cell-cell junctional targeting. We further observed that Madin-Darby canine kidney cell lines stably expressing a nephrocystin mutant with a deletion of the Src homology 3 domain have reduced ability to establish tight junctions as measured by transepithelial electrical resistance. Finally, from a two-hybrid screen and coimmunoprecipitation studies we identified members of the filamin family of actin-binding proteins as having the capacity to interact with the NHD. These findings support a functional role for nephrocystin as a docking protein involved in organizing a protein complex to regulate the actin cytoskeleton at sites of epithelial cell-cell adhesion and further suggest that these properties are important for establishing epithelial cell polarity.

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Translocation and gross deletion breakpoints in human inherited disease and cancer I: Nucleotide composition and recombination-associated motifs

et al. 2003

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Translocations and gross deletions are important causes of both cancer and inherited disease. Such gene rearrangements are nonrandomly distributed in the human genome as a consequence of selection for growth advantage and/or the inherent potential of some DNA sequences to be frequently involved in breakage and recombination. Using the Gross Rearrangement Breakpoint Database [GRaBD; www.uwcm.ac.uk/uwcm/mg/grabd/grabd.html] (containing 397 germ-line and somatic DNA breakpoint junction sequences derived from 219 different rearrangements underlying human inherited disease and cancer), we have analyzed the sequence context of translocation and deletion breakpoints in a search for general characteristics that might have rendered these sequences prone to rearrangement. The oligonucleotide composition of breakpoint junctions and a set of reference sequences, matched for length and genomic location, were compared with respect to their nucleotide composition. Deletion breakpoints were found to be AT-rich whereas by comparison, translocation breakpoints were GC-rich. Alternating purine-pyrimidine sequences were found to be significantly over-represented in the vicinity of deletion breakpoints while polypyrimidine tracts were over-represented at translocation breakpoints. A number of recombination-associated motifs were found to be over-represented at translocation breakpoints (including DNA polymerase pause sites/frameshift hotspots, immunoglobulin heavy chain class switch sites, heptamer/nonamer V(D)J recombination signal sequences, translin binding sites, and the chi element) but, with the exception of the translin-binding site and immunoglobulin heavy chain class switch sites, none of these motifs were over-represented at deletion breakpoints. Alu sequences were found to span both breakpoints in seven cases of gross deletion that may thus be inferred to have arisen by homologous recombination. Our results are therefore consistent with a role for homologous unequal recombination in deletion mutagenesis and a role for nonhomologous recombination in the generation of translocations.

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A deletion distinct from the classical homologous recombination of juvenile nephronophthisis type 1 (NPH1) allows exact molecular definition of deletion breakpoints

Cited by 27 publications

References 41 publications

Multi-exon deletions of the FBN1gene in Marfan syndrome

Multi-exon deletions of the FBN1gene in Marfan syndrome

Nephrocystin-conserved Domains Involved in Targeting to Epithelial Cell-Cell Junctions, Interaction with Filamins, and Establishing Cell Polarity

Translocation and gross deletion breakpoints in human inherited disease and cancer I: Nucleotide composition and recombination-associated motifs

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