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PubMedDate not supplied

A novel splicing mutation in exon 4 (456G>A) of the GH1 gene in a patient with congenital isolated growth hormone deficiency.

Isolated Growth Hormone Deficiency (IGHD) due to GH1 gene defects has a variable inheritance pattern: autosomal recessive, autosomal dominant, and X-linked. the autosomal dominantly inherited form, IGHD II, is mainly caused by heterozygous mutations of splicing around the exon 3/IVs3 boundary region of the GH1 gene resulting in exon 3 skipping of transcripts. We have previously reported findings on GH1 gene mutations in 28 russian patients with severe congenital IGHD (-3.22+/-1.2 height sDs at the age of 1yr); five heterozygous dominant negative splice site mutations in intron 2, intron 3, and exon 4 of the GH1 gene were identified in 32.1% of the cohort. In the present report we describe a novel 456G>A heterozygous mutation of splicing of the last base of the 3'-acceptor splice site of exon 4 within the GH1 in a 4.2-year old, extremely short (-5.32 height sDs) girl with congenital IGHD. the mutation involves a highly conserved GGGgtg sequence of the exon 4/IVs4 boundary region of the GH1 gene. the predicted effect of the 456 G>A mutation is perturbed splicing with possible skipping of exon 4 of the GH1 gene. the novel heterozygous 456 G>A mutation in exon 4 expands the spectrum of dominant negative splicing defects within the GH1 gene, responsible for congenital IGHD.

Child, Preschool↗

PubMed2005

A duplication in the canine beta-galactosidase gene GLB1 causes exon skipping and GM1-gangliosidosis in Alaskan huskies.

GM(1)-gangliosidosis is a lysosomal storage disease that is inherited as an autosomal recessive disorder, predominantly caused by structural defects in the beta-galactosidase gene (GLB1). The molecular cause of GM(1)-gangliosidosis in Alaskan huskies was investigated and a novel 19-bp duplication in exon 15 of the GLB1 gene was identified. The duplication comprised positions +1688-+1706 of the GLB1 cDNA. It partially disrupted a potential exon splicing enhancer (ESE), leading to exon skipping in a fraction of the transcripts. Thus, the mutation caused the expression of two different mRNAs from the mutant allele. One transcript contained the complete exon 15 with the 19-bp duplication, while the other transcript lacked exon 15. In the transcript containing exon 15 with the 19-bp duplication a premature termination codon (PTC) appeared, but due to its localization in the last exon of canine GLB1, nonsense-mediated RNA decay (NMD) did not occur. As a consequence of these molecular events two different truncated GLB1 proteins are predicted to be expressed from the mutant GLB1 allele. In heterozygous carrier animals the wild-type allele produces sufficient amounts of the active enzyme to prevent clinical signs of disease. In affected homozygous dogs no functional GLB1 is synthesized and G(M1)-gangliosidosis occurs.

Alleles↗

PubMed1998

A partial cDNA sequence of the ovine insulin receptor gene: evidence for alternative splicing of an exon 11 region and for tissue-specific regulation of receptor isoform expression in sheep muscle, adipose tissue and liver.

Insulin is as integral and important to the management of metabolism in ruminants as it is in non-ruminants. The suggestion of a lowered ruminant sensitivity and/or responsivity to insulin may relate more to the insulin receptor than to the hormone itself. We screened an ovine cDNA library using degenerate primers and polymerase chain reaction (PCR) to detect and sequence a cDNA portion corresponding to exons 10, 11 and 12 of the human insulin receptor gene in which a 36 base pair (bp) segment (exon 11) is alternatively spliced to produce two distinct receptor isoforms differing in functional characteristics including binding affinity for insulin. The ovine cDNA segment (nucleotides 671 to 770) displayed 84, 84, and 78% nucleotide homology to equivalent segments from the human, rhesus monkey and rat respectively. Reverse transcription PCR (RT-PCR) of selected tissues (liver, m. longissimus dorsi, m. rectus capitis and omental, perirenal and subcutaneous fats) taken at slaughter from three male, pure Dutch Texel lambs (experiment 1) and five male Texel-Greyface crossbred lambs (experiment 2) revealed two mRNA products in each tissue (including spleen; experiment 2 only) corresponding to cDNAs of molecular sizes 161 and 197 bp -- a difference of 36 bp. Sequence alignment showed the 36 bp segment to be homologous to the alternatively spliced exon 11 region of the human insulin receptor gene and to be highly conserved with that from other species. The abundance of the exon 11(+) isoform in the purebred Texel genotype was significantly higher in liver than in perirenal fat and rectus capitis and longissimus dorsi skeletal muscles (P<0.05) and higher also than in subcutaneous and omental fats (P<0.01). There was, however, no difference in the abundance of the exon 11(+) isoform between the individual muscle and fat depots in this sheep genotype. The abundance of the exon 11(+) isoform in the crossbred Texel genotype was significantly higher in liver (P<0. 05) than in the muscles (rectus capitis, P<0.05; longissimus dorsi, P<0.001), all three fats (P<0.001) and spleen (P<0.001). In the crossbred genotype, the abundance of the exon 11(+) isoform was higher in skeletal muscle than in all three fat depots (P<0.001), in which the isoform abundance was similar. Altered ratios of expression of the two products of this alternative splicing event could determine tissue sensitivity and/or responsivity to insulin and provide a mechanism for the management of nutrient partitioning and nutrient utilisation between tissues which is fundamental to the growth of tissues and manipulation of carcass characteristics in meat-producing animals.

Adipose Tissue↗

PubMed2001

The role of exon 5 in fibroblast collagenase (MMP-1) substrate specificity and inhibitor selectivity.

Interstitial collagen is degraded by members of the matrix metalloproteinase (MMP) family, including MMP-1. Previous work has shown that the region of MMP-1 coded for by exon 5 is implicated both in substrate specificity and inhibitor selectivity. We have constructed a chimeric enzyme, the exon 5 chimera, consisting primarily of MMP-1, with the region coded for by exon 5 replaced with the equivalent region of MMP-3, a noncollagenolytic MMP. Unlike MMP-3, the exon 5 chimera is capable of cleaving type I collagen, but the activity is only 2.2% of trypsin-activated MMP-1. 'Superactivation' of the chimera has no discernible effect, suggesting that the salt bridge formed in 'superactive' MMP-1 is not present. The kinetics for exon 5 chimera cleavage of two synthetic substrates display an MMP-3 phenotype, however, cleavage of gelatin is slightly impaired as compared to the parent enzymes. The K(iapp) values for the exon 5 chimera complexed with synthetic inhibitors and N-terminal TIMP-2 also show a more MMP-3-like behaviour. However, the k(on) values for N-terminal TIMP-1 and N-terminal TIMP-2 are more comparable to those for MMP-1. These data show that the region of MMP-1 coded for by exon 5 is involved in both substrate specificity and inhibitor selectivity and the structural basis for our findings is discussed.

Amino Acid Sequence↗

PubMed2001

A nonsense mutation in MLH1 causes exon skipping in three unrelated HNPCC families.

Germline mutations in the DNA mismatch repair genes MSH2 and MLH1 are responsible for the majority of hereditary nonpolyposis colorectal cancer (HNPCC) families. A common mutation mechanism is to disrupt MLH1 and MSH2 mRNA splicing. The disruption creates aberrant mRNAs lacking specific coding exons (exon skipping). Here, we report a novel skipping of MLH1 exon 12 caused by an AAG to TAG nonsense mutation at codon 461 in three HNPCC families of North American origins. The nonsense codon was found in a conserved haplotype in the three unrelated families and seems to represent a founder mutation. The skipping created an aberrant MLH1 mRNA transcript lacking exon 12. The effect of the codon 461 nonsense mutation on exon 12 skipping is evident even though it was placed in a minigene construct containing entirely different coding sequences. Notably, the effect of the nonsense mutation on exon skipping is incomplete. Accordingly, a second aberrant MLH1 transcript encompassing the nonsense codon is also produced. Whereas the latter transcript is unstable, presumably because of nonsense-mediated mRNA decay, neither of the aberrant transcripts seems to affect the stability of wild-type MLH1 mRNA. This study demonstrates that the germ-line nonsense mutation at codon 461 of MLH1 disrupts normal MLH1 mRNA processing, and that exon skipping underlies pathogenesis in these HNPCC families.

Adaptor Proteins, Signal Transducing↗

PubMedDate not supplied

[Spliced oct-1 gene mRNA isoforms with untranslated exons and a deletion in the region coding for the POU-specific domain].

Transcription factor Oct-1 is involved in expression regulation of housekeeping genes, in lymphocyte differentiation, and in the immune response. Tissue-specific oct-1 mRNA isoforms are known to be expressed in lymphoid cells. Four new mouse isoforms were identified. Of these, two were tissue-specific (oct-1R alpha and oct-1R beta) and contained exon 1L. The oct-1R alpha was shown to contain an additional fragment, which corresponds to an exon located in the 3'-region of mouse otf-1. No homolog was found in human OTF-1. The oct-1R alpha isoform proved to lack an exon coding for a fragment of the POU domain. This deletion results in a loss of the first helix of the domain, and the mutant protein is devoid of affinity for octamer ATGCAAAT. Two other mRNA isoforms, oct-1d and oct-1e, were shown to contain untranslated regions between exons 1U and 2. The regions correspond to exons 1i and 2i located between exons 1U and 1L in the 5'-region of the mouse oct-1 gene. Human OTF-1 was not found to contain exon 1i. On evidence of these and published data, it was assumed that a set of oct-1 isoforms is present in the cell, reflecting the complexity of expression regulation of oct-1 and the multiplicity of its functions.

5' Untranslated Regions↗

PubMed2003

Active transport of the survival motor neuron protein and the role of exon-7 in cytoplasmic localization.

Spinal muscular atrophy (SMA) is a neurodegenerative disease caused by deletion and/or mutation of the survival motor neuron protein Gene (SMN1) that results in the expression of a truncated protein lacking the C terminal exon-7. Whereas SMN has been shown to be an important component of diverse ribonucleoprotein (RNP) complexes, its function in neurons is unknown. We hypothesize that the active transport of SMN may be important for neurite outgrowth and that disruption of exon-7 could impair its normal intracellular trafficking. SMN was localized in granules that were associated with cytoskeletal filament systems and distributed throughout neurites and growth cones. Live cell imaging of enhanced green fluorescent protein (EGFP)-SMN granules revealed rapid, bidirectional and cytoskeletal-dependent movements. Exon-7 was necessary for localization of SMN into the cytoplasm but was not sufficient for granule formation and transport. A cytoplasmic targeting signal within exon-7 was identified that could completely redistribute the nuclear protein D-box binding factor 1 into the cytoplasm. Neurons transfected with SMN lacking exon-7 had significantly shorter neurites, a defect that could be rescued by redirecting the exon-7 deletion mutant into neurites by a targeting sequence from growth-associated protein-43. These findings provide the first demonstration of cytoskeletal-based active transport of SMN in neuronal processes and the function of exon-7 in cytoplasmic localization. Such observations provide motivation to investigate possible transport defects or inefficiency of SMN associated RNPs in motor neuron axons in SMA.

Actin Cytoskeleton↗

PubMed1992

Expression of variable exon A-, B-, and C-specific CD45 determinants on peripheral and thymic T cell populations.

A mAb (I/24) has been generated that is specific for a determinant on mouse CD45 molecules. Reactivity of this mAb with a panel of CD45 transfected cell lines demonstrated that the determinant recognized is dependent upon expression of one or more CD45 variable exons and that exon C is sufficient for its expression. The exon C-specific epitope detected by I/24 is expressed at high density on essentially all B lymphocytes and at an intermediate density on the vast majority of CD8+ splenic T cells. Two distinct subpopulations of CD4+ splenic T cells were detected, a minor subpopulation that expresses this exon determinant at high density and a major subpopulation that expresses it at a much lower density. This first identification of a CD45RC-specific reagent allowed a comparison of the expression of exon A-, exon B-, and exon C-specific determinants on peripheral and thymic lymphoid populations. When splenic lymphocytes were analyzed for expression of CD45RA (reactive with mAb 14.8), CD45RB (reactive with mAb 23G2 or mAb 16.A), and CD45RC (reactive with mAb I/24) determinants, it was found that each of these CD45 determinants had a distinct pattern of expression on CD4+ and CD8+ T cells and B cells. CD45RB and RC epitopes were also detected at high density on a small proportion (0.7 to 4.1%) of thymocytes. Both CD45RB and RC epitopes were found predominantly on CD4-CD8- and CD4-CD8+ thymocytes but were also found on small numbers of CD4+CD8+ and CD4+CD8- cells. The population of thymocytes that expressed CD45RB and CD45RC determinants displayed a novel TCR CD3 phenotype characterized by a level of expression that was intermediate between that seen in the larger CD3 bright and CD3 dull populations of thymocytes.

Animals↗

PubMed2004

[Exon polymorphism of human RHD gene].

OBJECTIVE: To study exon polymorphism of human RHD gene and investigate the genetic mechanism of RhD-negative individuals. METHODS: PCR using sequence-specific primers (PCR-SSP) was performed on 40 RhD-positive, 120 RhD-negative and 2 weak D blood samples. RESULTS: All 10 exons could be detected in the 40 RhD-positive and 2 weak D samples. Out of the 120 RhD-negative samples, 28 (23.33%) carried 10 exons, 19 (15.83%) lost most of the 10 exons (with mainly intermediate deletion), and 73 (60.83%) had deletion of all the 10 exons; 19 samples of Del phenotype identified from the 120 RhD-negative samples had all the 10 exons. CONCLUSION: Polymorphism of the exon structure of RHD gene is present in RhD-negative individuals, characterized chiefly by gross deletion, partial deletion and non-deletion.

Asian People↗

PubMed2006

[Direct sequencing and analysis of exons of Wilson' disease gene with the most mutations: a study in Hunan Han patients].

OBJECTIVE: To detect hot point mutations of ATP7B gene in Hunan Han patients with Wilson' disease (WD). METHODS: The genomic DNA of 22 WD patients was extracted and exons 5, 8, 12, 13 were amplified by PCR. Screening for the mutations was done by direct sequencing and analysed by BLAST. RESULTS: Fifteen of the 22 patients were found with mutations. Ten heterozygous Arg778Leu (2273G --> T) mutations were found in exon 8, all of them were accompanied with 2250C --> G polymorphism (Leu770Leu). Seven patients were found with 2855G --> A (Arg952Lys) polymorphism (4 heterozygous and 3 homozygous), 3 of them had Arg778Leu mutation in exon 8 and one with heterozygous mutation Gly943Asp (2828G --> A) in exon 12 simultaneously. Only one patient was found with heterozygous Pro992Leu (2975C --> T) mutation in exon 13. No mutations were found in exon 5. CONCLUSION: Arg778Leu is the hot point mutation of ATP7B gene in Hunan Han patients with Wilson' disease while exon 5 is not.

Adenosine Triphosphatases↗

PubMed1991

Sequence of DNA flanking the exons of the HEXA gene, and identification of mutations in Tay-Sachs disease.

The rapid identification of mutations causing Tay-Sachs disease requires the capacity to readily screen the regions of the HEXA gene most likely to be affected by mutation. We have sequenced the portions of the introns flanking each of the 14 HEXA exons in order to specify oligonucleotide primers for the PCR-dependent amplification of each exon and splice-junction sequence. The amplified products were analyzed, by electrophoresis in nondenaturing polyacrylamide gels, for the presence of either heteroduplexes, derived from the annealing of normal and mutant DNA strands, or single-strand conformational polymorphisms (SSCP), derived from the renaturation of single-stranded DNA. Five novel mutations from Tay-Sachs disease patients were detected: a 5-bp deletion of TCTCC in IVS-9; a 2-bp deletion of TG in exon 5; G78 to A, giving a stop codon in exon 1; G533 to T in exon 5, producing the third amino acid substitution detected at this site; and G to C at position 1 of IVS-2, expected to produce abnormal splicing. In addition, two mutations, (G1496 to A in exon 13 and a 4-bp insertion in exon 11) that have previously been reported were identified.

Alleles↗

PubMed1991

Human alpha 1 (XIII) collagen gene. Multiple forms of the gene transcripts are generated through complex alternative splicing of several short exons.

The structure of the gene for the human alpha 1(XIII) collagen chain (COL13A1) was determined from genomic clones spanning 140,000 base pairs (bp), including about 3,000 bp of the 5'-end-flanking region and 5,000 bp of the 3'-end-flanking region. The gene was shown to contain 39 exons. There were eight exons of 27 bp, five of 36 bp, four of 54 bp, three of 45 bp, and two of 42 bp. The rest of the exons coding for translated sequences had sizes varying between 24 and 153 bp. The genomic clones did not contain exons 3 and 4 whose sizes could, however, be estimated from cDNA clones. S1 nuclease mapping and primer extension analyzes indicated five closely spaced initiation sites of transcription. Sequencing of the 5'-end-flanking region did not reveal a typical TATA box but a four times repeated TATTTAT sequence that may serve as true TATA boxes. Two CCAAT boxes were found starting at positions-13 and -194, and furthermore, the promoter region contains two GC boxes. Previous studies on alpha 1 (XIII) collagen cDNA and genomic clones showed that the primary transcript undergoes complex alternative splicing generating at least four different forms of mRNAs. The present work demonstrated that sequences of seven exons are alternatively used. These exons contain sequences coding for pure collagenous regions, pure noncollagenous regions, and an exon coding for a junction of a collagenous and noncollagenous domain.

Amino Acid Sequence↗

PubMed1991

Effect of deletion of glycoprotein IIb exon 28 on the expression of the platelet glycoprotein IIb/IIIa complex.

We have isolated from an HEL cell cDNA library an alternatively spliced transcript for the platelet membrane glycoprotein IIb (GPIIb) that resulted from the deletion of the 34 amino acids of exon 28 of the GPIIb gene. Confirming an earlier report, we also detected this transcript in platelet mRNA. To determine the consequences of exon 28 deletion on the expression of the GPIIb/IIIa heterodimer, we expressed cDNA for GPIIb-28 in COS-1 cells, either individually or simultaneously with a cDNA for GPIIIa. When recombinant GPIIb-28 was expressed alone, it did not acquire resistance to the enzyme endo-beta-N-acetylglucosaminidase H, was not cleaved into heavy and light chains, and was not transported to the cell surface. However, when recombinant GPIIb-28 was coexpressed with recombinant GPIIIa, GPIIb/IIIa heterodimers were assembled. Nevertheless, these heterodimers failed to complete posttranslational processing and were degraded intracellularly. Exon 28 contains one site for Asn-linked glycosylation. To determine if loss of this glycosylation site was responsible for the effects of exon 28 deletion, we removed the site from the exon 28 of intact GPIIb by oligonucleotide-mediated mutagenesis. However, absence of the carbohydrate appended to exon 28 did not prevent normal GPIIb/IIIa heterodimer expression. Our studies indicate that absence of the amino acids encoded by GPIIb exon 28 sufficiently perturbs the quaternary configuration of the GPIIb/IIIa heterodimer to impair its subsequent intracellular transport and processing. They also indicate that this alternatively spliced form of GPIIb mRNA, although present in megakaryocytes, is unlikely to make a significant contribution to the GPIIb/IIIa complexes expressed on platelets.

Base Sequence↗

PubMed1990

Deletion of a splice donor site ablates expression of the following exon and produces an unphosphorylated RB protein unable to bind SV40 T antigen.

Studies of mutated retinoblastoma (RB) proteins in human tumor cells potentially reveal regions of the normal RB gene product that are required for its cancer suppression function. We here characterize a mutated RB protein of Mr 104,000 (p104) from a primary small-cell lung carcinoma. Unlike normal RB protein (pp110RB), p104 was unphosphorylated and unable to bind T antigen of SV40 both in vivo and in vitro. On the other hand, nuclear localization and DNA binding activity were preserved in the mutated protein. p104 was immunoprecipitable with four separate polyclonal antibodies recognizing different epitopes of the RB polypeptide, suggesting the presence of most exons in their correct reading frame. Following reverse transcription and in vitro amplification, RB mRNA from this tumor was shown to lack nucleotides encoded by exon 16. Analysis of genomic DNA from this tumor showed that exon 16 and its flanking splice donor and acceptor sequences were present and entirely normal; however, a 43-base pair (bp) region containing the splice donor site of intron 15 was deleted instead. Exon 15 was joined directly to exon 17 during mRNA processing via a cryptic splice donor site; exon 16 was presumably skipped because the preceding mutated intron was of insufficient length (less than 80 bp) for normal RB mRNA processing. These results demonstrate that loss of a single small exon disrupts several important biochemical properties of RB protein. In addition, sequence features of the 43-bp depletion suggest involvement of a novel deletional mechanism.

Antigens, Polyomavirus Transforming↗

PubMed1988

Identification of a mutation that causes exon skipping during collagen pre-mRNA splicing in an Ehlers-Danlos syndrome variant.

Recent biochemical studies have shown that the fibroblasts from a patient with Ehlers-Danlos Syndrome Type VIIB produce nearly equal amounts of normal and shortened pro-alpha 2(I) collagen chains (Wirtz, M.K., Glanville, R. W., Steinmann, B., Rao, V. H., and Hollister, D. (1987) J. Biol. Chem. 262, 16376-16385). Compositional and sequencing studies of the abnormal pro-alpha 2(I) chain identified an interstitial deletion of 18 residues corresponding to the N-telopeptide of the collagen molecule. Since this region is encoded by a 54-base pair exon, number 6, the protein defect could have been caused by gene deletion, abnormal pre-mRNA splicing, or both. Here, in order to elucidate the molecular nature of this mutation we have analyzed the sequences of pro-alpha 2(I) collagen cDNA and genomic clones obtained from RNA and DNA of the patient's fibroblasts. Using oligomer-specific cloning we identified a cDNA that contains a 54-base pair deletion corresponding precisely to the sequence of exon 6. Identification of the normal gene was based on the finding of an identical sequence polymorphism in a normal cDNA and in the genomic clone derived from one of the two collagen alleles. The other gene, instead, displayed a base substitution (T to C) in the obligatory GT dinucleotide of the 5' splice-site sequence of intron 6. Analysis of nearly 100 base pairs immediately 5' to exons 5, 6, and 7, and 3' to exons 5 and 7 did not reveal any additional change. Therefore, the data strongly suggest that the observed GT-to-GC transition at the splice donor site of intron 6 generates an abnormally spliced mRNA in which the sequence of exon 5 is joined to the sequence of exon 7. Since skipping of exon 6 does not interfere with the coding frame of the mRNA, the resulting shortened polypeptide, albeit utilized in the assembly of a procollagen trimer, ultimately causes the Ehlers-Danlos Syndrome Type VII phenotype.

Alleles↗

PubMed1989

Inhibition of calpain by a synthetic oligopeptide corresponding to an exon of the human calpastatin gene.

Calpastatin is a widely distributed endogenous inhibitor protein specifically acting on calpain (Ca2+-dependent cysteine endopeptidase). The inhibitor consists of four inhibitory domains (Domains 1-4) with mutually homologous sequences. NH2-terminal Domain L is non-homologous, and all domains have 120-140 residues each. A human calpastatin genomic DNA clone was isolated using a previously obtained human calpastatin cDNA probe. Sequence analysis has revealed that the clone contains Domain 1 and segments of neighboring domains (Domains L and 2). Each of three highly conserved, restricted regions within Domain 1 was located on separate exons, 1A, 1B, and 1C. Exon 2A, corresponding to the first exon of Domain 2, is homologous to Exon 1A and follows Exon 1D of Domain 1. A 27-residue peptide encoded by Exon 1B, including a 12-residue middle conserved sequence, was chemically synthesized and tested for protease inhibitory activities. The synthetic peptide showed strong inhibition against calpain I (low Ca2+-requiring form), and calpain II (high Ca2+-requiring form), but no inhibition against papain or trypsin. These results indicated that Exon 1B forms a self-sufficient functional subdomain of the calpastatin inhibitory domain.

Amino Acid Sequence↗

PubMed1986

Molecular mapping of murine I region recombinants. II. Crossing over in the E beta gene is restricted to a 4.5 kb stretch of DNA that excludes the beta 1 exon.

The crossing over in five murine I-region recombinants (Is/Ik) was studied by restriction fragment length polymorphism analysis after Southern blot hybridization by using I-region-specific probes. These recombinants included three recently developed strains, B10.ASR1, B10.ASR11, and B10.ASR12; and two strains B10.S(9R) and B10.HTT derived earlier. Although these recombinants were reciprocal in haplotype orientation to the three recombinants we reported recently, these too crossed over within the same 7 kb stretch of DNA in the E beta gene. This 7 kb stretch of DNA included the 3' half of the first intron, the beta 1 exon, the second intron, and the beta 2 exon. A comparison of the cDNA sequences of the two parental E beta alleles revealed that although the beta 2 exons were identical, there were several nucleotide differences between the two beta 1 exons. This allowed us to determine the parental origin of the beta 1 exon in the recombinants at the level of transcription by using S1 nuclease mapping. Thus we were able to show that in each case the 3' portion of the first intron and the beta 1 exon were upstream from the site of crossover. All eight recombinants involving the k and s haplotypes now can be mapped within a 4.5 kb stretch of DNA, which includes only the beta 1-beta 2 intron and the beta 2 exon of the E beta gene. These findings imply that the I-E molecules expressed in these recombinants will probably have conserved sequences and therefore will exhibit identical I-E-restricted immune responses, despite the fact that crossing over could have occurred at different sites within the beta 1-beta 2 intron.

Alleles↗

PubMed1995

Inherited human complement C5 deficiency. Nonsense mutations in exons 1 (Gln1 to Stop) and 36 (Arg1458 to Stop) and compound heterozygosity in three African-American families.

Hereditary C5 deficiency has been reported in several families of different ethnic backgrounds and from different geographic regions, but the molecular genetic defect causing C5 deficiency has not been delineated in any of them. To examine the molecular basis of C5 deficiency in the African-American population, the exons and intron/exon boundaries of the C5 structural genes from three C5-deficient (C5D) African-American families were sequenced, revealing two nonsense mutations. The nonsense mutations are located in exon 1 (C84AG to TAG) in two of the C5D families (Rhode Island and North Carolina) and in exon 36 (C4521GA to TGA) in the third C5D family (New York). The exon 1 and 36 mutations are contained in codons that encode the first amino acid of the C5 beta-chain (Gln1 to Stop) and residue 1458 in the alpha-chain (Arg1458 to Stop), respectively. Allele-specific PCR and sequence analyses demonstrated that the exon 1 mutation is present in only one of the C5 null genes in both the Rhode Island and North Carolina families, and the exon 36 mutation is contained in only one C5 null gene in the New York family. Neither of the nonsense mutations was found in the European or Caucasian-American C5D individuals examined. Collectively, these data indicate that: 1) C5 deficiency is caused by several different molecular genetic defects, 2) C5 deficiency in the African-American population can be explained in part by two distinct nonsense mutations in exons 1 and 36, and 3) compound heterozygosity exists in all of the reported African-American C5D families.

Alleles↗