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Y W Yang

Publications and source records attributed to Y W Yang.

At least 55 records · Page 3Linked to original sources
PubMed1991

Cloning and sequencing of bovine apolipoprotein E complementary DNA and molecular evolution of apolipoproteins E, C-I, and C-II.

Apolipoprotein (apo) E, a major protein component of plasma lipoproteins, is a physiological ligand for the low density lipoprotein (LDL) receptor as well as for a specific apoE receptor; it is therefore an important modulator of lipoprotein metabolism. In this study we cloned and sequenced bovine apoE complementary DNA. Comparison of nucleotide substitution rates shows that apoE is less conservative than apoA-I and evolves about 30% faster than an average mammalian protein. Although apoE is not a conservative protein, several regions have been well conserved among all eight mammalian sequences now available. These include a 33-amino-acid block immediately upsteam from the third intron/exon junction and the LDL receptor binding region. We have also compared published apoC-I and apoC-II sequences. Both proteins are less conservative than apoE. In particular, apoC-I shows no well-conserved region except for a small region in the common 33-amino-acid block, suggesting that the function of apoC-I does not have stringent structural requirements. On the other hand, in apoC-II the region encoded by exon 4, which consists of the last 29 amino acids of the polypeptide, has been rather well conserved, probably because this region is important for the activation of lipoprotein lipase and chylomicron and very low density lipoprotein metabolism.

Amino Acid Sequence↗
PubMed1991

The chick embryo fibroblast cation-independent mannose 6-phosphate receptor is functional and immunologically related to the mammalian insulin-like growth factor-II (IGF-II)/man 6-P receptor but does not bind IGF-II.

The insulin-like growth factor-II (IGF-II)/Mannose 6-P receptor (Man 6-P) is a multifunctional receptor that binds two unrelated ligands, IGF-II and lysosomal enzymes that contain Man 6-P recognition markers. Although this receptor has been extensively characterized in mammalian cells, binding of radiolabeled IGF-II to this receptor in avian cells and tissues has not been reported. In the present study, we demonstrate that chick embryo fibroblasts (CEFs) bind and internalize lysosomal enzymes in a Man 6-P-inhibitable fashion, and possess a protein immunologically related to the mammalian IGF-II/Man 6-P receptor that binds lysosomal enzymes with Man 6-P recognition markers but does not bind IGF-II. 1) When lysates of biosynthetically labeled CEFs were affinity-purified on beta-galactosidase-Sepharose, an approximately 250 kilodalton protein was observed in the Man 6-P eluate but not in the Glc 1-P or mannose eluates, that was precipitated by antisera to purified rat and bovine IGF-II/Man 6-P receptors, but not by nonimmune serum. 2) When CEFs were incubated with [35S]proteins enriched in lysosomal enzymes, Man 6-P inhibited binding (0 C) and uptake (34 C) in a dose-dependent fashion. Binding was unchanged in the absence of divalent cations. At low sugar concentrations, binding and uptake were inhibited selectively by Man 6-P and the conformationally similar sugar phosphate, Fru 1-P, a specificity similar to that of mammalian cation-independent Man 6-P receptors. 3) When affinity-purified lysates from biosynthetically labeled CEFs were incubated with antiserum to the rat IGF-II/Man 6-P receptor, a 245 kilodalton protein was immunoprecipitated from lysates that had been affinity purified on beta-galactosidase-Sepharose but not after purification on IGF-II-Sepharose. By contrast, a truncated IGF-II/Man 6-P receptor, presumably internalized from the fetal bovine serum used to feed the cells, was purified from lysates of unlabeled CEFs on IGF-II-Sepharose. Thus, CEFs possess a cation-independent Man 6-P receptor that is similar in size and immunological reactivity to the mammalian IGF-II/Man 6-P receptor, and binds and internalizes lysosomal enzymes but, unlike the mammalian receptor, does not bind IGF-II.

Animals↗
PubMed1991

Insulin rapidly inhibits insulin-like growth factor-binding protein-1 gene expression in H4-II-E rat hepatoma cells.

The insulin-like growth factor-binding proteins (IGFBPs) are thought to determine the distribution of IGF-I and IGF-II between the blood and tissue compartments and to modulate their biological activities. A dynamic metabolic role for one of the IGFBPs, IGFBP-1, is suggested by the fact that plasma IGFBP-1 was increased after fasting and diabetes and rapidly decreased by refeeding or insulin treatment, respectively. IGFBP-1 mRNA also is increased in the livers of diabetic rats and decreased by insulin treatment. To understand the molecular basis for this regulation, we have examined the effects of insulin on IGFBP-1 and IGFBP-1 mRNA in the H4-II-E cell line derived from the well differentiated H35 rat hepatoma. IGFBP-1, identified by ligand blotting and immunoblotting, is the major IGFBP in H4-II-E cells. Incubation of H4-II-E cells with insulin for 24 h decreased IGFBP-1 in the culture medium by approximately 50%. Inhibition was observed at physiological concentrations of insulin (ED50, less than 0.5 nM), but not at higher concentrations of IGF-II. These results, together with the fact that H4-II-E cells do not possess IGF-I receptors with which insulin might cross-react, suggest that insulin acts via the insulin receptor. Insulin inhibited IGFBP-1 in the medium by 80% in the absence of glucose, suggesting that the inhibition is a direct effect of insulin; glucose exerted a smaller independent effect in the absence of insulin. Insulin decreased IGFBP-1 mRNA in H4-II-E cells by 50% within 1 h and by 90% after 2-12 h of incubation. Nuclear run-on transcription assays indicated a corresponding decrease in the rate of IGFBP-1 gene transcription. Pretreatment of H4-II-E cells with dexamethasone stimulated IGFBP-1 transcription and increased steady state IGFBP-1 mRNA; stimulation was abolished by insulin treatment, indicating that inhibition by insulin was dominant over induction by dexamethasone. Thus, insulin, acting through the insulin receptor, rapidly decreases the abundance of IGFBP-1 mRNA in H4-II-E cells. Regulation occurs at least in part at the level of gene transcription. We propose that regulation of IGFBP-1 synthesis is an important component of the regulation of IGFBP-1 by insulin in vivo.

Animals↗
PubMed1990

The cDNA and protein sequences of mouse lactate dehydrogenase B. Molecular evolution of vertebrate lactate dehydrogenase genes A (muscle), B (heart) and C (testis).

Mouse lactate dehydrogenase-B cDNAs were isolated from cDNA libraries of macrophage (ICR strain) and thymus (F1 hybrid of C57BL/6 and CBA strains), and their nucleotide sequences determined. The lactate dehydrogenase-B cDNA insert of thymus clone mB188 consists of the protein-coding sequence (1002 nucleotides), the 5' (46 nucleotides) and 3' (190 nucleotides) non-coding regions, and poly(A) tail (19 nucleotides), while macrophage clone mB168 contains a partial lactate dehydrogenase cDNA insert from codon no. 55 to the poly(A) tail. Seven silent nucleotide substitutions at codon no. 142, 143, 186, 187, 241, 285 and 292, as well as a single nucleotide change in the 3' non-coding region, were found between these different strains of mice. The predicted sequence of 333 amino acids, excluding initiation methionine, was confirmed by sequencing and/or compositional analyses of a total of 103 (31%) amino acids from tryptic peptides of mouse lactate dehydrogenase-B protein. The nucleotide sequence of the mouse coding region for lactate dehydrogenase B shows 86% identity with that of the human isoenzyme, and only eight of the 139 nucleotide differences resulted in amino acid substitutions at residues 10, 13, 14, 17, 52, 132, 236 and 317. The rates of nucleotide substitutions at synonymous and nonsynonymous sites in the mammalian lactate dehydrogenase genes are calculated. The rates of synonymous substitutions for lactate dehydrogenase genes A (muscle) and B (heart) are considerably higher than the average rate computed from human and rodent genes. The rates of nonsynonymous substitutions for lactate dehydrogenase genes A (muscle) and B (heart), particularly the latter, are highly conservative. The rates of synonymous and nonsynonymous substitutions for the lactate dehydrogenase-C gene are about the same as the average rates for mammalian genes. A phylogenetic tree of vertebrate lactate dehydrogenase protein sequences is constructed. In agreement with the previous results, this analysis further indicates that lactate dehydrogenase-C gene branched off earlier than did lactate dehydrogenase-A and lactate dehydrogenase-B genes.

Amino Acid Sequence↗
PubMed1990

Molecular phylogeny of Rodentia, Lagomorpha, Primates, Artiodactyla, and Carnivora and molecular clocks.

Phylogenetic analysis of DNA sequences from primates, rodents, lagomorphs, artiodactyls, carnivores, and birds strongly suggests that the order Rodentia is an outgroup to the other four mammalian orders and that Artiodactyla and Carnivora belong to a superordinal clade. Further, there is strong evidence against the Glires concept, which unites Lagomorpha and Rodentia. The radiation among Lagomorpha, Primates, and Artiodactyla--Carnivora is very bush-like, but there is some evidence that Lagomorpha has branched off first. Thus, the branching sequence for these five orders of mammals seems to be Rodentia, Lagomorpha, Primates, Artiodactyla, and Carnivora. The branching date for Rodentia could be as early as 100 million years ago. The rate of nucleotide substitution in the rodent lineage is shown to be at least 1.5 times higher than those in the other four mammalian lineages.

Animals↗
PubMed1990

Tissue, developmental, and metabolic regulation of messenger ribonucleic acid encoding a rat insulin-like growth factor-binding protein.

Insulin-like growth factor-II (IGF-II) is the predominant insulin-like growth factor in fetal and neonatal rat serum and tissues. In serum, it occurs complexed to a 30-kDa nonglycosylated IGF-binding protein (IGFBP) that is immunologically related to the IGFBP in BRL-3A rat liver cells (rIGFBP-2). Levels of rIGFBP-2 and IGF-II decrease in rat serum after birth. Using a recently isolated cDNA clone for rIGFBP-2 as hybridization probe, we now compare the expression of rIGFBP-2 and IGF-II in fetal tissues and the effects of hypophysectomy and fasting on the abundance of these mRNAs in adult rat liver. rIGFBP-2 mRNA is expressed at high levels in term gestation liver and at lower levels in other tissues. The ratio of rIGFBP-2 to IGF-II mRNAs in stomach, kidney, and lung is similar to that seen in liver, whereas IGF-II mRNA is more abundant than rIGFBP-2 mRNA in muscle, intestine, heart, and skin. Both mRNAs are more abundant in fetal tissues than in the corresponding tissues from adult rats. Dexamethasone treatment of 4-day-old rats for 4 days caused a greater (90%) decrease in hepatic IGF-II mRNA than in rIGFBP-2 mRNA (50%), suggesting subtle differences in the developmental regulation of the two mRNAs. Even more striking differences were observed in the regulation of the two mRNAs in adult rats after hypophysectomy or fasting. Hepatic rIGFBP-2 mRNA was increased 10- to 20-fold compared to age-matched control rats, whereas IGF-II mRNA was not increased. A parallel increase in serum rIGFBP-2 was observed, suggesting that this regulation may result at least in part from the increased abundance of rIGFBP-2 mRNA. Thus, in addition to modulating the stimulation of growth and differentiation by IGF-II in fetal tissues, rIGFBP-2 may play a homeostatic role during catabolic states in the adult rat.

Animals↗
PubMed1990

Identification of rat cell lines that preferentially express insulin-like growth factor binding proteins rlGFBP-1, 2, or 3.

The bioavailability and action of the insulin-like growth factors (IGFs) are determined by specific IGF-binding proteins (IGFBP) to which they are complexed. Complementary DNA clones have been isolated that encode three related IGFBPs: human IGFBP-1 (hIGFBP-1), human IGFBP-3 (hIGFBP-3), and rat IGFBP-2 (rIGFBP-2). IGFBP-1 and IGFBP-3 are regulated differently in human plasma, suggesting that they have different functions. In order to study the molecular basis of the regulation of the different IGFBPs, we have identified a panel of rat cell lines that express a single predominant binding protein and developed an assay strategy to distinguish the different binding proteins. Proteins in conditioned medium were examined by ligand blotting, and by immunoprecipitation and immunoblotting using antibodies to rIGFBP-2 and hIGFBP-1; RNAs were hybridized to cDNA probes for rIGFBP-2 and hIGFBP-1. 1) C6 glial cells and B104 neuroblastoma cells express an approximately 40 kilodalton (kDa) glycosylated binding protein that most likely represents rIGFBP-3, the binding subunit of the 150 kDa IGF: binding protein complex in adult rat serum. The C6 and B104 binding proteins do not react with antibodies to rIGFBP-2, and RNAs from C6 and B104 cells do not hybridize to cDNA probes for rIGFBP-2 or hIGFBP-1. 2) BRL-3A, Clone 9, and TRL 12-15 cell lines derived from normal rat liver express rIGFBP-2, a 30 kDa nonglycosylated IGF-binding protein that is recognized by antibodies to rIGFBP-2 but not by antibodies to hIGFBP-1. RNAs from these cells hybridize to a rIGFBP-2 cDNA probe, but not to a hIGFBP-1 probe. 3) H35 rat hepatoma cells express a 30 kDa nonglycosylated IGFBP that is presumptively identified as rIGFBP-1. It does not react with antibodies to rIGFBP-2, but is recognized by polyclonal and monoclonal antibodies to hIGFBP-1. RNA from H35 cells hybridizes to a hIGFBP-1 cDNA probe, but not to a rIGFBP-2 probe. Expression of rIGFBP-1 by the H35 cell line has enabled us to establish and validate specific assays for this protein that allow us to study its regulation in intact rats. Identification of a panel of rat cell lines expressing specific IGFBPs should be useful in elucidating the molecular mechanisms of IGFBP regulation.

Animals↗
PubMed1989

Nucleotide sequence of cloned nad4 (urf4) gene from Chlamydomonas reinhardtii mitochondrial DNA.

Chlamydomonas reinhardtii mitochondrial (mt)DNA was digested with ClaI + HpaI and shotgun cloned into the M13mp19 vector cleaved with AccI + SmaI. One of the recombinant clones, with a 1.8-kb DNA insert, was completely sequenced using the dideoxy chain-termination method. Besides containing part of the cytochrome b (COB)-encoding gene (cob), this DNA fragment encodes subunit 4 of NADH dehydrogenase (NAD4). The deduced amino acid sequence and hydrophilicity plot indicate that NAD4 is highly hydrophobic. The nad4 gene shows a unique preference for certain codons which are also found in other C. reinhardtii mt proteins. Both the genes encoding NAD4 and COB are shown to be transcriptionally active by Northern hybridization. These closely linked genes suggest that RNA-processing events found in vertebrate mt are present in Chlamydomonas mt as well.

Amino Acid Sequence↗
PubMed1989

The 34 kilodalton insulin-like growth factor binding proteins in human cerebrospinal fluid and the A673 rhabdomyosarcoma cell line are human homologues of the rat BRL-3A binding protein.

Three members of a family of insulin-like growth factor binding proteins have been identified by nucleotide sequencing of cDNA clones: the binding subunit of the 150 kDa IGF-binding protein complex in human serum, the 30 kDa IGF binding protein in human amniotic fluid, and a 30 kDa binding protein (BP-3A) isolated from the rat BRL-3A cell line. The present study demonstrates by molecular hybridization and immunoreactivity that the human counterpart of rat BP-3A is a 34 kDa IGF binding protein that is present in human cerebrospinal fluid and is synthesized and secreted by the A673 human rhabdomyosarcoma cell line.

Amniotic Fluid↗
PubMed1989

Human interstitial retinoid-binding protein. Gene structure and primary structure.

Interstitial retinoid-binding protein (IRBP) is synthesized and secreted by rod photoreceptor cells into the interphotoreceptor matrix and is known to bind retinoids and fatty acids. We have used cDNA clones encoding human IRBP to isolate a 15-kilobase genomic fragment that encompasses the complete human IRBP gene. The IRBP gene spans more than 11 kilobases and is interrupted by three introns, all of which are positioned near the 3'-end of the coding sequence. The 3741-base pair coding region of IRBP appears to have been generated by quadruplication of an approximately 900 base pair long ancestral gene. The deduced amino acid sequence predicts a mature protein of 1,230 residues (calculated molecular weight 133,000). The protein sequence can be aligned into four homologous segments, each consisting of about 300 residues. Sequence similarity between segments is as high as 60% when conservative substitutions are taken into account. Two putative N-linked glycosylation sites are located in highly conserved domains in the center of the first and second segment of IRBP. A domain consisting of 41 residues at the COOH-terminal end of the third segment has 15 matching residues (38%) with an intradiscal loop of rhodopsin, a retinal-binding protein in rod photoreceptors.

Amino Acid Sequence↗
PubMed1989

Date of the monocot-dicot divergence estimated from chloroplast DNA sequence data.

The divergence between monocots and dicots represents a major event in higher plant evolution, yet the date of its occurrence remains unknown because of the scarcity of relevant fossils. We have estimated this date by reconstructing phylogenetic trees from chloroplast DNA sequences, using two independent approaches: the rate of synonymous nucleotide substitution was calibrated from the divergence of maize, wheat, and rice, whereas the rate of nonsynonymous substitution was calibrated from the divergence of angiosperms and bryophytes. Both methods lead to an estimate of the monocot-dicot divergence at 200 million years (Myr) ago (with an uncertainty of about 40 Myr). This estimate is also supported by analyses of the nuclear genes encoding large and small subunit ribosomal RNAs. These results imply that the angiosperm lineage emerged in Jurassic-Triassic time, which considerably predates its appearance in the fossil record (approximately 120 Myr ago). We estimate the divergence between cycads and angiosperms to be approximately 340 Myr, which can be taken as an upper bound for the age of angiosperms.

Animals↗
PubMed1989

Structure, specificity, and regulation of the insulin-like growth factor-binding proteins in adult rat serum.

Insulin-like growth factor-I (IGF-I), the principal IGF in adult rat serum, occurs complexed to specific binding proteins. After fractionation of serum on Sephadex G-200 at neutral pH, 62% of the immunoreactive IGF-I is recovered in the 150K region, 38% in the 40K region, and none is present as free 7.5K IGF-I. Adult rat serum also contains unoccupied binding sites for IGFs that also are predominantly (77%) located in the 150K region and have preferential binding affinity for IGF-II. IGF-binding protein components in the 150K and 40K regions were evaluated by affinity cross-linking to 125I-labeled IGFs and by ligand blotting (i.e. incubation of nitrocellulose blots of sodium dodecyl sulfate (SDS)-gels with [125I]IGFs). Affinity cross-linking of the 150K region revealed a major 43K binding protein complex and several minor covalent complexes of 97-210K that are formed during the cross-linking reaction. The 40K region of the gel filtration column contains a predominant 32K complex and smaller amounts of the 43K complex. Ligand blotting of the 150K region identifies a predominant cluster of binding components of about 40K and a smaller 29K protein. The apparent molecular masses of the 40K and 29K proteins are decreased by incubation with N-glycanase, indicating that they contain N-linked oligosaccharides. These glycoprotein components, designated gp40 and gp29, presumably combine with an acid-labile nonbinding subunit of about 100K to generate the 150K complex. The gp40 cluster represents glycosylation variants of a 34K protein; gp29 has been shown to correspond to an amino-terminal fragment of gp40. Ligand blotting of the 40K region indicates that it contains smaller amounts of gp40 and gp29, possibly representing free subunits not combined with the nonbinding subunit, as well as two proteins of apparent molecular mass 24K and 30K (p24 and p30) that are not glycosylated. Although p30 is similar in size to the binding protein from BRL-3A cells (BP-3A) that is present in fetal rat serum, immunoprecipitation and immunoblotting of whole and fractionated adult serum with an antiserum to BP-3A indicate that p30 in adult rat serum is an antigenically distinct protein. Serum levels of gp40 and gp29 are decreased by hypophysectomy and are restored by GH treatment; p24 and p30 show similar but smaller changes.

Animals↗
PubMed1989

The fetal rat binding protein for insulin-like growth factors is expressed in the choroid plexus and cerebrospinal fluid of adult rats.

The biological effects of the insulin-like growth factors, IGF-I and IGF-II, on their receptors are modulated by IGF-binding proteins. Recently, we isolated a cDNA clone for one member of the family of IGF-binding proteins, BP-3A, a 30 kilodalton (kDa) protein synthesized by the BRL-3A rat liver cell line. BP-3A is related to but distinct from two other cloned IGF-binding proteins, the human amniotic fluid binding protein and the glycosylated binding subunit of the 150 kDa IGF-binding protein complex in serum. It is expressed in multiple nonneural tissues and in serum in the fetal rat and decreases after birth, similar to the developmental pattern of IGF-II expression. IGF-I, IGF-II, and their receptors are expressed in brain. The present study examines the expression of BP-3A in the rat central nervous system. By Northern blot analysis, BP-3A mRNA is present at high levels in brain stem, cerebral cortex, and hypothalamus from 21-day gestation rats and, like IGF-II mRNA, persists in adult rat brain. The site of BP-3A mRNA synthesis was localized by in situ hybridization to coronal sections of adult rat brain using 35S-labeled oligonucleotides, 48 bases in length, complementary and anticomplementary to the coding region of BP-3A. Specific hybridization of the BP-3A probe was observed exclusively to the choroid plexus extending from the level of the medial preoptic nucleus to the arcuate nucleus of the hypothalamus, similar to the previously reported preferential localization of IGF-II mRNA to the choroid plexus. Synthesis of BP-3A mRNA by choroid plexus suggested that BP-3A might be secreted into the cerebrospinal fluid. A 30 kDa IGF-binding protein was demonstrated in rat cerebrospinal fluid that is recognized by antibodies to BP-3A and, like purified BP-3A, has equal affinity for IGF-I and IGF-II. By analogy with other transport proteins synthesized by the choroid plexus, BP-3A may facilitate the secretion of IGF-II to the cerebrospinal fluid and modulate its biological actions at distant sites within the brain.

Animals↗