C3 metabolism in a patient with deficiency of the second component of complement (C2) and discoid lupus erythematosus.
(9/259)
A patient with a hereditary deficiency of the second component of complement and discoid lupus erythematosus with features of systemic lupus erythematosus was studied. The propositus had a 9-year history of rash and arthralgia. Transient renal disease had completely resolved; there was a history of seizures. Examination of his serum disclosed antinuclear antibodies but no total haemolytic complement activity. C2 was absent. Serum concentrations of C1s, C3, C5 and C9 were elevated; other complement components were present in normal concentration, including C3 pro-activator. The patient's C3 pro-activator was electrophoretically converted by inulin and four of five lipopolysaccharides, but was poorly converted by aggregated human IgG. Two separate turnover studies with radiolabelled C3 showed fractional catabolic rates of 3-03 and 2-48% of the remaining plasma pool/hr (range of three normals: 1-62-2-18%/hr); and estimated C3 synthetic rates of 2-74 and 2-31 mg/kg/hr (range of three normals: 0-89-1-40 mg/kg/hr). Serum complement profiles of the patient's family demonstrated that the C2 deficiency was inherited as an autosomal codominant. One sibling, homozygous for C2 deficiency, and three other siblings, both parents and one daughter, all heterozygous for C2 deficiency, are in good health. Immunofluorescent studies of the patient's diseased skin exhibited substantial deposits of IgG, IgM, C1q, and C4 but not of later acting complement components, properdin, or C3 proactivator. These studies do not support the notion that inflammation in C3-deficient individuals with lupus erythematosus is mediated by the alternative complement pathway. (+info)
Transcriptional termination and coupled polyadenylation in vitro.
(10/259)
Using a coupled, in vitro transcription and polyadenylation system we have investigated the molecular mechanism of transcriptional termination by RNA polymerase II (PolII). We showed previously that specific G-rich sequences pause transcription and then activate polyadenylation. We show that physiological pause sites activate polyadenylation in our in vitro system. We also investigate the mechanism of PolII transcriptional termination, and show that these transcripts are either directly released from the transcription complex or are 3' end processed while still attached to the complex. We also show that 3' product (generated by cleavage/polyadenylation) remains associated with the transcription complex, but is rapidly degraded on it. (+info)
Phylogenetic analysis of retroposon family as exemplified on human chromosome 13: further evidence for recent proliferation.
(11/259)
The retroposon SINE-R.C2 was first identified as a human-specific insertion in the complement C2 gene. In our previous study, SINE-R type retroposons, derived from the endogenous retrovirus HERV-K family, have been found to be hominoid specific. In this report on human chromosome 13, we identified eighteen new SINE-R retroposons resembling those we have previously reported on the sex chromosomes and on chromosomes 7 and 17. Phylogenetic analysis using the neighbor-joining method revealed that four SINE-R retroposons (13-16, 21, 23, 25) on chromosome 13 were closely related to the human-specific retroposon SINE-R.C2, with a high degree of sequence homology (95-97%). Such elements differ from the HERV-K10. LTR sequence from which they are derived in being deleted for the promoter region. Therefore while the evidence adds to the case that some classes of SINE-R element have continued to proliferate in hominid and hominoid evolution and may, as in the case of Fukuyama type muscular dystrophy, be a cause of insertional mutagenesis, they are less likely than the HERV-K10 LTR to have a positive effect on host gene activity. (+info)
Two clusters of acidic amino acids near the NH2 terminus of complement component C4 alpha'-chain are important for C2 binding.
(12/259)
Previous work has indicated a role for the NH2-terminal segment of the C3 alpha'-chain in the binding interactions of C3b with a number of its protein ligands. In particular, we have identified two clusters of acidic residues, namely, E736 and E737 and to a lesser extent D730 and E731, as being important in the binding of C3b to factor B and complement receptor 1 and the binding of iC3b to complement receptor 3. Whereas human C3 and C4 have an overall sequence identity of 29%, over a segment near the NH2 termini of their respective alpha'-chains the sequence identity is 56% (70% chemical similarity). Given the functional similarity between the C4b-C2 and C3b-B interactions in the respective formation of the classical and alternative pathway C3 convertases, as well as the sequence conservation of two acidic clusters, we hypothesized that residues 744EED and 749DEDD within the NH2-terminal segment of the C4 alpha'-chain would mediate in part the binding of C2 to C4b. We tested this hypothesis using three independent approaches. Site-directed mutagenesis experiments revealed that replacing subsets of the charged residues by their isosteric amides within either acidic cluster resulted in molecules having reduced C2 binding activity. Moreover, a synthetic peptide (C4 residues 740-756) encompassing the two acidic clusters was a specific inhibitor of the binding of C2 to red cell-associated C4b. Finally, Ab raised against the above peptide was able to block the interaction between red cell-associated C4b and fluid phase C2. Taken together, these results strongly suggest that the NH2-terminal acidic residue-rich segment of C4 alpha'-chain contributes importantly to the interaction of C4b with C2. (+info)
Hypochlorite-induced alterations to canine serum complement.
(13/259)
Changes in the concentration of the components of complement produced by NaOC1 both in vitro and in vivo are recorded. C1, C4 and C7 are particularly sensitive to this oxidizing agent, although all components decrease at high concentrations of NaOC1. Following oxidation, complement componenets return rapidly to normal. Data are presented to indicate that part of this repair mechanism is due to the action of reducing agents such as ascorbic acid and part is due to the synthesis of the individual components. The unique sensitivity of complement components to oxidation make this treatment of potential value in suppressing the inflammatory response. (+info)
Hypoxia increases the sensitivity of the L-type Ca(2+) current to beta-adrenergic receptor stimulation via a C2 region-containing protein kinase C isoform.
(14/259)
The effects of hypoxia on the L-type Ca(2+) current (I:(Ca-L)) in the absence and presence of the ss-adrenergic receptor agonist isoproterenol (Iso) were examined. Exposing guinea pig ventricular myocytes to hypoxia alone resulted in a reversible inhibition of basal I:(Ca-L). When cells were exposed to Iso in the presence of hypoxia, the K:(0.5) for activation of I:(Ca-L) by Iso was significantly decreased from 5.3+/-0.7 to 1.6+/-0.1 nmol/L. The membrane-impermeant thiol-specific oxidizing compound 5, 5'-dithio-bis(2-nitrobenzoic acid) (DTNB) attenuated the inhibition of basal I:(Ca-L) by hypoxia 81.3+/-9.4% but had no effect on the increase in sensitivity of I:(Ca-L) to Iso. In addition, DTT mimicked the effects of hypoxia on basal I:(Ca-L) and the increase in sensitivity to Iso. Neither the inhibitors of guanylate cyclase LY-83583 or methylene blue nor the NO synthase inhibitor N:(G)-monomethyl-L-arginine monoacetate had any effect on the basal inhibition of I:(Ca-L) or the decrease in K:(0.5) for activation of I:(Ca-L) by Iso during hypoxia. However, the protein kinase C (PKC) inhibitors bisindolylmaleimide I and Go 7874 significantly attenuated the increase in sensitivity of I:(Ca-L) to Iso. More specifically, the response was attenuated when cells were dialyzed with a peptide inhibitor of the C2 region-containing classical PKC isoforms. The same effect was not observed with the PKCepsilon peptide inhibitor. These results suggest that hypoxia regulates I:(Ca-L) through the following 2 distinct mechanisms: direct inhibition of basal I:(Ca-L) and an indirect effect on the sensitivity of the channel to ss-adrenergic receptor stimulation that is mediated through a classical PKC isoform. (+info)
Lack of evidence of a specific role for C4A gene deficiency in determining disease susceptibility among C4-deficient patients with systemic lupus erythematosus (SLE).
(15/259)
The aim of the present study was to investigate the prevalence of C4 and C2 deficiencies and to characterize genomic alterations in C4 genes in a large cohort of 125 unselected patients with SLE. We determined the protein concentration and functional activity of C2 and C4, as well as the C4 phenotype. C4 genotyping included Taq 1 restricted fragment lengh polymorphism (RFLP) analysis and polymerase chain reaction using sequence-specific primers (SSP-PCR). Type I C2 deficiency was diagnosed by PCR. Overall, 79.2% of the patients exhibited abnormalities of the C4 genes including deletion, non-expression, gene conversion and duplication. Among C4-deficient patients (n = 66, 52.8% prevalence), 41.0% of the patients exhibited a C4A deficiency and 59.0% a C4B deficiency. Half of the C4 deficiencies were due to a gene deletion. There was a strong association between C4A and C4B gene deletion and the presence of the DRB1*03 allele. Among the silent C4A genes, only two cases were related to a 2-bp insertion in exon 29 of the C4A gene. A gene conversion was demonstrated in eight patients (6.4%). One patient had a homozygous C4A deficiency. Three (2.4%) patients presented with a heterozygous type I C2 deficiency and none with homozygous deficiency. Our results argue against a specific role for C4A gene deficiency in determining disease susceptibility among patients with SLE that are C4-deficient. (+info)
Human immunodeficiency virus type 1 induces expression of complement factors in human astrocytes.
(16/259)
Since the brain is separated from the blood immune system by a tight barrier, the brain-resident complement system may represent a central player in the immune defense of this compartment against human immunodeficiency virus (HIV). Chronic complement activation, however, may participate in HIV-associated neurodegeneration. Since the level of complement factors in the cerebrospinal fluid is known to be elevated in AIDS-associated neurological disorders, we evaluated the effect of HIV type 1 (HIV-1) on the complement synthesis of brain astrocytes. Incubation of different astrocytic cell lines and primary astrocytes with HIV-1 induced a marked upregulation of the expression of the complement factors C2 and C3. The synthesis of other secreted or membrane-bound complement proteins was not found to be altered. The enhancement of C3 production was measured both on the mRNA level and as secreted protein in the culture supernatants. HIV-1 laboratory strains as well as primary isolates were capable of inducing C3 production with varied effectiveness. The usage of viral coreceptors by HIV-1 was proved to be a prerequisite for the upregulation of C3 synthesis, which was modulated by the simultaneous addition of cytokines. The C3 protein which is secreted after incubation of the cells with HIV was shown to be biologically active as it can participate in the complement cascade. (+info)