RESUMEN
BACKGROUND: The aetiology of atypical haemolytic uraemic syndrome (aHUS) is, in contrast to classical, Shiga-like toxin induced HUS in children, largely unknown. Deficiency of human complement factor H and familial occurrence led to identification of the factor H gene (FH1) as the susceptibility gene, but the frequency and relevance of FH1 mutations are unknown. METHODS: We established a German registry for aHUS and analysed in all patients and 100 controls the complete FH1 gene by single strand confirmational polymorphism and DNA sequencing. In addition, complement C3 and factor H serum levels were assayed. Demographic data at onset of aHUS and follow up were compared for the mutation positive and negative groups. RESULTS: Of 111 patients with aHUS (68 female, 43 male, mean age 33 years) 14% had FH1 germline mutations, including two of eight patients with familial aHUS. For each of these eight patients, both parents were tested, and we were able to trace the mutation for five cases. In the other three cases (one with the mutation 3749 C/T, one with 3200 T/C, and one with 3566+1 G/A), we could not detect the mutation in either parent, although paternity was proven by genetic fingerprinting, suggesting that these subjects have new mutations. C3 was decreased in five mutation carriers but also in two non-carriers, and factor H was decreased in none of the carriers, but elevated in six carriers and 15 non-carriers. Clinical parameters including associated medications and diseases, and outcome of aHUS and of post-aHUS kidney transplantation were similar in the mutation positive and negative groups. CONCLUSION: FH1 germline mutations occur with considerable frequency in patients with aHUS. Hypocomplementaemia is not regularly associated with a germline mutation, and factor H serum levels can even be elevated. Screening for FH1 mutations contributes to the classification of aHUS.
Asunto(s)
Factor H de Complemento/genética , Síndrome Hemolítico-Urémico/genética , Adulto , Austria , Complemento C3/metabolismo , Factor H de Complemento/metabolismo , ADN/química , ADN/genética , Análisis Mutacional de ADN , Ensayo de Inmunoadsorción Enzimática , Femenino , Alemania , Síndrome Hemolítico-Urémico/sangre , Síndrome Hemolítico-Urémico/complicaciones , Humanos , Italia , Fallo Renal Crónico/complicaciones , Fallo Renal Crónico/terapia , Trasplante de Riñón , Masculino , Mutación , Polimorfismo Conformacional Retorcido-Simple , Sistema de Registros/estadística & datos numéricos , SuizaRESUMEN
Complement factor H (fH) is a member of a family of proteins involved in the regulation of complement activation (RCA). These proteins share a common structural motif, the Short Consensus Repeat (SCR), which is structurally conserved among related genes and between phylogenetically divergent species. fH is composed of 20 such SCRs and a variety of biological functions have been localised to specific SCR domains. The majority of individual SCRs identified are encoded by single exons, and processes such as gene conversion, duplication and exon shuffling have been implicated in the evolution and genomic radiation of SCR-encoding genes. We have analysed two GenBank sequence entries relating to two overlapping PAC clones sequenced at the Sanger Centre which contain the entire human fH gene and two adjacent fH-related (fHR) genes, fHR-1 and fHR-3. Here, we report the detailed analysis of the assembled 221 kb of contiguous, ungapped genomic sequence from human chromosome 1q32, in part employing the RUMMAGE-DP automated annotation tool. Genomic duplications involving fH and fHR exons were identified and Alu/L1 repeat dating established that the duplications occurred after the separation of rodent and primate lineages. The analysis indicates that retrotransposition as well as single and multiple exon duplication events are likely to have been involved in SCR radiation and RCA gene evolution, facilitated by conservation of splice-phasing and the single-exon, single-SCR nature of the encoded domains.
Asunto(s)
Factor H de Complemento/genética , ADN/química , Regiones no Traducidas 5' , Secuencia de Bases , Exones , Humanos , Datos de Secuencia Molecular , Señales de Clasificación de Proteína/genética , Secuencias Repetitivas de Ácidos Nucleicos , Lugares Marcados de Secuencia , Transcripción GenéticaRESUMEN
The RUNX1 gene on human chromosome 21q22.12 belongs to the 'runt domain' gene family of transcription factors (also known as AML/CBFA/PEBP2alpha). RUNX1 is a key regulator of hematopoiesis and a frequent target of leukemia associated chromosomal translocations. Here we present a detailed analysis of the RUNX1 locus based on its complete genomic sequence. RUNX1 spans 260 kb and its expression is regulated through two distinct promoter regions, that are 160 kb apart. A very large CpG island complex marks the proximal promoter (promoter-2), and an additional CpG island is located at the 3' end of the gene. Hitherto, 12 different alternatively spliced RUNX1 cDNAs have been identified. Genomic sequence analysis of intron/exon boundaries of these cDNAs has shown that all consist of properly spliced authentic coding regions. This indicates that the large repertoire of RUNX1 proteins, ranging in size between 20-52 kDa, are generated through usage of alternatively spliced exons some of which contain in frame stop codons. The gene's introns are largely depleted of repetitive sequences, especially of the LINE1 family. The RUNX1 locus marks the transition from a ~1 Mb of gene-poor region containing only pseudogenes, to a gene-rich region containing several functional genes. A search for RUNX1 sequences that may be involved in the high frequency of chromosomal translocations revealed that a 555 bp long segment originating in chromosome 11 FLI1 gene was transposed into RUNX1 intron 4.1. This intron harbors the t(8;21) and t(3;21) chromosomal breakpoints involved in acute myeloid leukemia. Interestingly, the FLI1 homologous sequence contains a breakpoint of the t(11;22) translocation associated with Ewing's tumors, and may have a similar function in RUNX1.
Asunto(s)
Empalme Alternativo , Cromosomas Humanos Par 21 , Proteínas de Unión al ADN/genética , Proteínas Proto-Oncogénicas , Factores de Transcripción/genética , Translocación Genética , Regiones no Traducidas 3' , Regiones no Traducidas 5' , Secuencia de Aminoácidos , Mapeo Contig , Subunidad alfa 2 del Factor de Unión al Sitio Principal , Islas de CpG , Exones , Orden Génico , Humanos , Secuencias Repetitivas Esparcidas , Intrones , Leucemia/genética , Datos de Secuencia Molecular , Proteína Proto-Oncogénica c-fli-1 , Seudogenes , Telómero/genética , Transactivadores/genéticaRESUMEN
Foreign particles and damaged host cells can activate the complement system leading to their destruction by the host defense system. Factor H (fH) plays a vital role in restricting complement activation on host cells through interactions with polyanions such as heparin, while allowing activation to proceed on foreign surfaces. Complement activation by damaged host cells is also down regulated by fH, which is localized to injured areas through interactions with C-reactive protein (CRP). A number of pathogens have developed mechanisms by which they can also bind fH and thus exploit its protective properties. One such organism is Group A Streptococcus (GAS) which mediates fH binding via its surface expressed M-protein. fH consists of 20 conserved short consensus repeat (SCR) units and mutagenesis studies indicate that the seventh repeat is responsible for interactions with heparin, CRP and M-protein. We recently performed molecular modelling of fH SCR 7 and identified a cluster of positively charged residues on one face of the domain. By alanine replacement mutagenesis, we demonstrated that these residues are involved in heparin, CRP and M protein binding, which indicates that there is a common site within fH SCR 7 responsible for multiple ligand recognition.
Asunto(s)
Antígenos Bacterianos , Proteínas de la Membrana Bacteriana Externa , Factor H de Complemento/química , Factor H de Complemento/metabolismo , Secuencia de Aminoácidos , Proteínas Bacterianas/inmunología , Sitios de Unión , Proteína C-Reactiva/inmunología , Proteínas Portadoras/inmunología , Activación de Complemento , Factor H de Complemento/genética , Heparina/química , Humanos , Técnicas In Vitro , Infecciones/etiología , Ligandos , Modelos Moleculares , Datos de Secuencia Molecular , Ácido N-Acetilneuramínico/inmunología , Estructura Terciaria de Proteína , Homología de Secuencia de Aminoácido , Streptococcus pyogenes/inmunologíaRESUMEN
Scleroderma is an enigmatic rheumatic disorder of uncertain etio-pathogenesis. Cancer has an approximately two-fold higher incidence in scleroderma patients than in the general population. There are preliminary data of acquired genetic damage in scleroderma but the significance of these observations are uncertain. To determine somatic mutation frequency at the glycophorin-A (GPA) locus in patients with limited and diffuse cutaneous scleroderma. The GPA assay measures the total somatic mutation frequency (Vf), composed of gene inactivating mutations (NO) and mutations arising from mitotic recombination (NN) in individuals heterozygous for the GPA MN blood group. Mutation frequency was determined using a validated GPA flow cytometric assay using fluorescent labeled monoclonal antibodies specific for the GPA blood groups M and N. This assay detects and enumerates progeny of red blood cell (rbc) precursor cells which have acquired genetic damage resulting in a loss of expression of one of the GPA alleles. It was found that patients with scleroderma (n = 23) had significantly elevated Vf as compared with young healthy controls (p < 0.001) and elderly controls (p = 0.03). Patients with diffuse scleroderma had higher mean Vf as compared with limited scleroderma (p = 0.055). In comparison with controls, patients with scleroderma exhibit a higher proportion of mitotic recombinant mutations than inactivating mutations (p < 0.002). There was no correlation between Vf and disease duration, age at onset or autoantibody status. We have documented evidence of acquired genetic damage at the GPA locus in scleroderma. Evidence of acquired genetic damage in this disorder may be importance in explaining both the etio-pathogenesis of scleroderma and the association of scleroderma with cancer.
Asunto(s)
Inestabilidad Genómica , Glicoforinas/genética , Esclerodermia Sistémica/genética , Adolescente , Anciano , Alelos , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , MutaciónRESUMEN
Magnesium has an established role in obstetrics and an evolving role in other clinical areas, in particular cardiology. Many of the effects involving magnesium are still a matter of controversy. Over the next decade, it is likely that improvements in the measurement of magnesium, a clearer understanding of the mechanisms of its actions and further results of clinical studies will help to elucidate its role, both in terms of treating deficiency and as a pharmacological agent.
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Deficiencia de Magnesio/metabolismo , Magnesio , Arritmias Cardíacas/prevención & control , Cationes , Eclampsia/tratamiento farmacológico , Femenino , Humanos , Intubación Intratraqueal , Magnesio/fisiología , Magnesio/uso terapéutico , Deficiencia de Magnesio/tratamiento farmacológico , Masculino , Infarto del Miocardio/prevención & control , Bloqueo Neuromuscular , Estado Nutricional , Preeclampsia/tratamiento farmacológico , EmbarazoRESUMEN
Factor H, a secretory glycoprotein comprising 20 short consensus repeat (SCR) or 'sushi' domains of about 60 amino acids each, is a regulator of the complement system. The complement-regulatory functions of factor H are targeted by its binding to polyanions such as heparin/sialic acid, involving SCRs 7 and 20. Recently, the SCR 7 heparin-binding site was shown to be co-localized with the Streptococcus Group A M protein binding site on factor H (T.K. Blackmore et al., Infect. Immun. 66, 1427 (1998)). Using sequence analysis of all heparin-binding domains of factor H and its closest homologues, molecular modeling of SCRs 6 and 7, and surface electrostatic potential studies, the residues implicated in heparin/sialic acid binding to SCR 7 have been localized to four regions of sequence space containing stretches of basic as well as histidine residues. The heparin-binding site is spatially compact and lies near the interface between SCRs 6 and 7, with residues in the interdomain linker playing a significant role.