A novel beta-thalassemia mutation [IVS-II-5 (G-->C)] in a Chinese family from Guangxi Province, P.R. China.
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Biomedical subjects
Publications and source records attributed to S Liang.
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A thermosensitive mutant was selected on the basis of its resistance after a heat shock to the lethal effects of lomofungin, a drug that inhibits RNA synthesis. We demonstrate that the single mutation conferring thermosensitivity and lomofungin resistance is heteroallelic to sec23-1. This new allele of sec23 is designated sec23-11. The yeast SEC23 gene has previously been reported to function in protein transport from the endoplasmic reticulum (ER) to the Golgi apparatus. After a 1-min shift to elevated temperature, a sec23-1 mutant becomes defective in vesicle formation from the ER. We show here that upon a shift to high temperature, both sec23 alleles confer defects in ER-to-Golgi transport, and both mutants stop accumulating nascent RNA after 20 to 30 min. Transcription and early maturation of ribosomal RNA, the major RNA species, are normal in a sec23-11 mutant, but the matured ribosomal RNA is then degraded. This ribosomal RNA instability may reflect improper assembly of ribosomal subunits, due to failure to localize essential factors such as ribosomal proteins to the nucleus. In addition to the sec23 mutants, other ER-to-Golgi secretion mutants exhibit a strong RNA biosynthetic defect, and we conclude that continued post-ER protein transport, not just a functional SEC23 gene product (Sec23p), is required for ribosomal RNA stability. We also identified an extragenic multicopy suppressor of the sec23-11 mutant, the essential STS1 gene ("Sec Twenty-three Suppressor 1"). Besides restoring partial temperature resistance to the sec23-11 mutant, overexpression of STS1 also overcomes both the RNA synthesis and the protein transport blocks in this mutant. In contrast, multicopy STS1 exerts only small effects on the sec23-1 mutant. The predicted STS1 gene product (Sts1p) is a low-abundance protein of 36,500 daltons, with no signal peptide and no hydrophobic stretch of sufficient length to span a membrane. Like Sec23p, Sts1p is found in a cytosolic compartment. These features suggest that Sts1p may play a catalytic or regulatory role in conjunction with the more abundant Sec23p.
Some of the acidic reagents or medicines (i.e. HCl, GSH & GSSG, Vit. C, Vit. B6) and basic reagents (Tris) were used to regulate the pH of water soluble protein and urea soluble protein solution of human lenses. At a certain pH, the lens protein solution appears opalescence, the pH of opacity solution which is measured by pH meter is around 5.6-6.3. The opalescence of water soluble and urea soluble lens protein solution of human fetal, adult and cataractous lenses were also determined by gradient acidic and basic solution. The result show that the opacity is more obvious in soluble protein solution of cataractous lenses and clear lenses than in that of fetal lenses, and the lens urea soluble protein has a marked opalescence in comparison with the water soluble protein. So we assume that cataractogenesis might be associated with the change of pH within lens.
The total proteins, water--soluble proteins and the urea--soluble proteins of the lens from fetal, adult and senile cataract were determined by SDS--PAGE. It was found that there was a 43KD polypeptide in the capsular--epithelium of all lens stated above. The band on SDS-PAGE corresponding to 43KD polypeptide was wide in the water--soluble proteins of the cortex and nucleus of the lenses obtained from over 14-year-old individuals. It became blurred in the urea--soluble proteins of cortex and nucleus of lens with aging, whereas it became almost disappeared in the senile cataract lens.
UV-spectrophotometry and fluorometry were used to study Malonaldehyde (MDA) and Superoxide Dismutase (SOD) in normal, cataractous human lenses and red blood cells of the patients with cataract. MDA content of senile and complicated cataractous lenses was significantly higher than that of normal human lenses, while that of complicated cataract was significantly higher than that of senile cataract. SOD activity of senile and complicated cataractous lenses was significantly lower than that of normal human lenses, while there was no marked difference between senile and complicated cataractous lenses. Significant correlation between cataractous lenses and red blood cells was not found in MDA content and SOD activity. There was a negative correlation between SOD and MDA in normal human lenses, but no correlation between SOD and MDA in cataractous lenses. The study shows that lipid peroxidation may be one of the possible mechanisms of cataractogenesis in human, and emphasizes the role of SOD in prevention of photoperoxidative damages to the tissues.
Retinal S-antigen and interphotoreceptor retinoid-binding protein (IRBP) were purified by DEAE-Sephacel ionexchange chromatography and Con A Sepharose affinity chromatography. Amino acid analysis of the S-antigen and IRBP was carried out and compared with that presented by others. The possible reason for difference with regard to some amino acids and related problems were analyzed and discussed.
Human fetal gamma 2-crystallin solution was irradiated by ultra-violet (UV) (lambda 365 nm) at varying time, and analysed by light scattering, SDS-PAGE with DACM, which could specifically combine to free sulphydryl, as well as tryptophan fluorescence and its quenching with acrylamide. The results showed that light scattering of gamma 2-crystallin solution increased after 24 hour irradiation; SDS-PAGE reveled formation of dimer which did not have free sulphydryl and low molecular weight peptide of gamma 2-crystallin. The tryptophan fluorescence of gamma 2-crystallin decreased, and acrylamide fluorescence quenching effect did not show significant change. It suggested that human fetal gamma 2-crystallin photooxidized by UV could aggreate by disulfide and degrated to low molecular weight peptide, and its tryptophan were also oxidized in some degree.
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A two-site ELISA and a bioassay were used to detect NGF-like activity in human brain tissue. Both assays detected mouse NGF and recombinant human NGF with approximately equal sensitivity, whereas the antibodies showed little cross-reactivity with the recombinant human proteins NT-3 and brain-derived neurotrophic factor. NGF-like activity was detected in fresh human cortical samples obtained from epileptic patients, with the highest activity observed in the right hemisphere of men. NGF-like activity was subsequently measured in autopsy samples of frontal and occipital cortex from patients with Alzheimer's disease (AD) and from individuals with no history or pathological evidence of AD. Based on both the ELISA and the bioassay measurements, NGF-like activity was significantly elevated in both brain regions in AD. These results demonstrate the feasibility of detecting NGF-like activity in both fresh and postmortem human brain tissue and further suggest that AD is characterized by increased, rather than decreased, levels of cortical beta-NGF. The AD-related increase in NGF may be a consequence of degenerative changes in the basal forebrain cholinergic system.
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Chick sympathetic (E-9) or telencephalic (E-7) neurons were cultured at low density on poly-DL-ornithine (PORN), poly-L-lysine (POLS), laminin or laminin-covered PORN or POLS and monitored with time-lapse videomicroscopy. Neurons migrated on laminin, or laminin-covered PORN or POLS, but not on PORN or POLS alone. Neuronal migration did not involve interactions with other cells indicating that neurons are capable of independent migration when exposed to a laminin substrate.
RNA12-1 is a dominant temperature-sensitive (Ts) yeast mutant which has previously been reported to exhibit a defect in RNA accumulation at 37 degrees C. We further characterized this mutant through analyses of rRNA transcription rates and maturation. The results show that pre-rRNA is normally synthesized but that subsequent maturation is severely affected by a temperature upshift: the nascent rRNAs are under-methylated and little mature rRNA can be observed at 37 degrees C. Likewise, the accumulation of some mRNAs for ribosomal proteins is also prevented at 37 degrees C. The RNA12-1 mutation is recessive at 32 degrees C, which made it possible to clone the wild-type rna12+ gene by complementation of the Ts phenotype with plasmids from a multicopy yeast genomic library. The predicted gene product is a protein of 96,630 Da with no significant sequence similarity to any known proteins. Gene disruption is not lethal at either the permissive or the restrictive temperature. The gene is located on chromosome XIII, downstream of the ADH2 gene and 10 cM from the ADE4 gene. Furthermore, the mutant allele RNA12-1 was cloned and sequenced. A point mutation found in this allele leads to dominant thermosensitivity at 37 degrees C when the mutant gene is introduced into a wild-type strain. Taken together, these data suggest that the rna12+ gene product plays a dispensable role in early maturation of pre-rRNA but that its mutant gene product can interfere with the normal function of other proteins required for pre-rRNA maturation.
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Three primers were designed, one specific for alpha 1-globin DNA, a second for alpha 2-globin DNA, and a third that is common for both alpha 1- and alpha 2-globin DNA. These three primers can be applied for selective amplification of the two globin DNA fragments, which is useful for identification of nondeletional types of Hb H disease. Fifty-nine DNA samples of Hb H patients from Guangxi were studied by selective amplification, and 27 cases (45.8%) were confirmed as nondeletional types. Of these, 22 (81.5%) had the Hb Constant Spring (CS) mutation and one had the Hb Quong Sze (QS) mutation; both were identified by hybridization with synthesized oligonucleotide probes. Nondeletional Hb H disease in Guangxi seems to be more severe than the deletional types. The average hemoglobin level of the nondeletional Hb H/CS (--/alpha alpha) is 6.8 g/dl, which is lower than that of the deletional types (7.9 g/dl), while the levels of Hb H and Hb Bart's were much higher in the patients with Hb H/CS than in those with the deletional types.
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The Pd(0)-catalyzed intramolecular olefination of vinyl and aryl triflates has been studied with a view to gaining insight into the question of kinetically preferred reductive elimination when at least two options are available. In either series, a non-activated alkene participant was found to be converted most readily into the non-conjugated cyclization product. This trend is seen despite the more forcing conditions necessary to engage the less reactive aryl triflates in ring closure. On the other hand, when the pendant chain is terminated by an alpha-methyl acrylate unit, the conjugated diene is kinetically preferred. The two reactions appear to be closely balanced energetically since product distributions are not greatly disparate. Nevertheless, their complementarity could be utilized to advantage in the synthesis of polycyclic molecules possessing multiple sterogenic centers.
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Interphotoreceptor retinoid-binding protein, IRBP, not only functioning as a shuttle to carry the retinoid between photoreceptor cells and pigment epithelium, but also inducing experimental autoimmune uveoretinitis (EAU), was purified by ConA Sepharose affinity chromatography from the fractions containing IRBP obtained in the course of ion-exchange chromatography by which the bovine retinal S-antigen was purified. This much simplified method allows more rapid purification of the two kinds of protein. EAU was successfully induced with injecting 50 micrograms of IRBP emulsified with Freund's complete adjuvant into the right hind footpads of Lewis rats. It was characterized by panophthalmia or endophthalmia with the severest damage in the posterior retina. Lymphocytes, mononuclear and polymorphonuclear cells were found to be the inflammatory cells of infiltration, with lymphocytes being predominant.