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Biomedical subjects

X Lu

Publications and source records attributed to X Lu.

At least 451 records · Page 25Linked to original sources
PubMed1994

The Caenorhabditis elegans locus lin-15, a negative regulator of a tyrosine kinase signaling pathway, encodes two different proteins.

The Caenorhabditis elegans locus lin-15 negatively regulates an intercellular signaling process that induces formation of the hermaphrodite vulva. The lin-15 locus controls two separate genetic activities. Mutants that lack both activities have multiple, ectopic pseudo-vulvae resulting from the overproduction of vulval cells, whereas mutants defective in only one lin-15 activity appear wild-type. lin-15 acts non-cell-autonomously to prevent the activation of a receptor tyrosine kinase/ras signaling pathway. We report here the molecular characterization of the lin-15 locus. The two lin-15 activities are encoded by contiguous genomic regions and by two distinct, non-overlapping transcripts that may be processed from a single mRNA precursor by trans-splicing. Based on the DNA sequence, the 719- and 1,440-amino acid lin-15 proteins are not similar to each other or to known proteins. lin-15 multivulva mutants, which are defective in both lin-15 activities, contain deletions and insertions that affect the lin-15 genomic region.

Amino Acid Sequence↗
PubMed1994

Inclusion of energy straggling in a numerical method for electron dose calculation.

Energy straggling along electron trajectories has been incorporated into a numerical algorithm for electron beam dose calculations. Landau's theory is used to predict, at any point in the absorber, the broadening of the primary electron energy spectrum due to energy loss straggling. Numerical calculations have been performed for electron beams with energies of 10-30 MeV incident upon water in order to determine the variation of dose with depth and variation of energy spectra with pathlength. These calculations are compared with the results of Monte Carlo simulations performed with the EGS4 code. The inclusion of energy loss straggling in the numerical calculations leads to predictions of energy spectra and dose deposition that are in good agreement with modified Monte Carlo simulations in which bremsstrahlung is ignored and the energy given to knock-on electrons is deposited at the site of their creation. Less satisfactory agreement was achieved when these calculations were compared to full Monte Carlo simulations that included the bremsstrahlung events and transported the knock-on electrons. It is concluded that bremsstrahlung energy loss must also be included into this algorithm, if an acceptable dose computation accuracy is to be achieved for clinical applications.

Computer Simulation↗
PubMed1994

Fine mapping and functional characterization of two immuno-dominant regions from the preS2 sequence of hepatitis B virus.

A set of monoclonal antibodies (mAbs) directed against the preS2 region of hepatitis B virus (HBV) surface antigen (HBsAg) was generated by immunization of mice with native HBsAg isolated from the blood of HBV carriers. According to (1) mutual competition binding of mAb to natural HBsAg, (2) recognition of full-length preS2 displayed on hepatitis B core particles, (3) recognition of synthetic partial preS2 peptides, and (4) Western blotting using a fusion protein library of truncated preS2 fragments of different legths, mAbs were assigned to two groups which coincided with groups I and III described by Mimms et al. [Virology 1990; 176:604-619]. All mAbs recognized linear epitopes and were glycosylation independent. Six out of eight fine-mapped mAbs recognized common epitopes located in the amino-terminal part of the preS sequence between amino acids 131 and 144 (group I), and inhibited binding of HBsAg to polymerized human serum albumin. Only two mAbs recognized a carboxy-terminal HBV-genotype-specific epitope covering amino acid residues 162 to 168 (group III). These mAbs bound to the highly variable proteolysis-sensitive hinge of preS2. Although four out of six mAbs targeted to immunodominant region I require the full-length sequence 131-L[Q/L]DPRVRGLY[F/L]PAG-144, two mAbs recognize the shorter and slightly carboxy-terminal-shifted sequences 133-DPRVRGLY[F/L]-141 or 135-PVRGLY[F/L]PAG-144. Together with previously identified preS2 epitopes 133-DPRVRGL-139, 137-RGLYFPA-143, and 132-QDPR-135, these data indicate diversity of the immune response against epitopes within the same immunodominant region. This diversity may be generated by a labile secondary structure. Sequence analysis suggests the transition from an alpha-helix to a loop structure at this site.

Amino Acid Sequence↗
PubMed1994

Identification of two N-terminal non-alpha-helical domain motifs important in the assembly of glial fibrillary acidic protein.

The non-alpha-helical N-terminal domain of intermediate filament proteins plays a key role in filament assembly. Previous studies have identified a nonapeptide motif, SSYRRIFGG, in the non-alpha-helical N-terminal domain of vimentin that is required for assembly. This motif is also found in desmin, peripherin and the type IV intermediate filament proteins. GFAP is the only type III intermediate filament protein in which this motif is not readily identified. This study has identified two motifs in the non-alpha-helical N-terminal domain of mouse GFAP that play important roles in GFAP assembly. One motif is located at the very N terminus and has the consensus sequence, MERRRITS-ARRSY. It has some characteristics in common with the vimentin nonapeptide motif, SSYRRIFGG, including its location in the non-alpha-helical N-terminal domain and a concentration of arginine residues. Unlike the vimentin motif in which even conserved sequence changes affect filament assembly, the GFAP consensus sequence, MERRRITS-ARRSY, can be replaced by a completely unrelated sequence; namely, the heptapeptide, MVRANKR, derived from the lambda cII protein. When fused to GFAP sequences with sequential deletions of the N-terminal domain, the lambda cII heptapeptide was used to help identify a second motif, termed the RP-box, which is located just upstream of the GFAP alpha-helical rod domain. This RP-box affected the efficiency of filament assembly as well as protein-protein interactions in the filament, as shown by sedimentation assays and electron microscopy. These results are supported by previous data, which showed that the dramatic reorganization of GFAP within cells was due to phosphorylation-dephosphorylation of a site located in this RP-box. The results in this study suggest the RP-box motif to be a key modulator in the mechanism of GFAP assembly, and support a role for this motif in both the nucleation and elongation phases of filament assembly. The RP-box motif in GFAP has the consensus sequence, RLSL-RM-PP. Sequences similar to the GFAP RP-box motif are also to be found in vimentin, desmin and peripherin. Like GFAP, these include phosphorylation and proteolysis sites and are adjacent to the start of the central alpha-helical rod domain, suggesting that this motif of general importance to type III intermediate filament protein assembly.

Amino Acid Sequence↗
PubMed1994

First-trimester anesthesia exposure and the risk of central nervous system defects: a population-based case-control study.

OBJECTIVES: Although up to 2% of women undergo surgery during pregnancy, teratogenic effects of general anesthesia have not been adequately studied. Recently, an association between first-trimester operations and central nervous system defects has been described. This issue was explored in a population-based case-control study. METHODS: Case patients included live-born and stillborn infants with central nervous system defects born to residents of metropolitan Atlanta, Ga, between 1968 and 1980. Control patients included normal babies frequency matched to case patients by race, birth hospital, and period of birth. Conditional logistic regression analysis was used to adjust for potential confounding factors. RESULTS: Of 694 mothers of infants with central nervous system defects, 12 reported first-trimester anesthesia exposure; 34 of 2984 control mothers reported such exposure (odds ratio [OR] = 1.7, 95% confidence interval [CI] = 0.8, 3.3). A striking association was observed between reported anesthesia exposure and hydrocephalus with another major defect (OR = 9.6, 95% CI = 3.8, 24.6). The strongest association was that of anesthesia exposure with hydrocephalus and eye defects (OR = 39.6, 95% CI = 7.5, 209.2). CONCLUSIONS: An increased risk of hydrocephalus with other defects was found among offspring of mothers with reported first-trimester anesthesia. Further studies are needed to explore the possible teratogenic effects of general anesthesia.

Abnormalities, Drug-Induced↗
PubMed1994

[A genetic study of adult polycystic kidney disease].

The 3'HVR has been considered as the most useful probe in making the linkage analysis of adult polycystic kidney disease (APKD). We examined three APKD families using this probe. Their polymorphism information was then computed for linkage study using LINKAGE package and computer programme HOMOG for homogeneity test. One family was defined as non PKD 1, because the disease locus was failed to show linkage to the 3'HVR marker. Of the other two families, one can still not be identified as non PKD1, but had distinguishable meiotic recombinations, another was in favour of linkage to 3'HVR and therefore judged to be PKD1. The presence of genetic heterogeneity could significantly reduce the value of linked probe analysis as a tool for gene diagnosis of APKD.

Adolescent↗
PubMed1994

[Role of cardiac renin-angiotensin system in swimming induced physiological myocardial hypertrophy].

To determine the contribution of cardiac renin-angiotensin system (RAS) to the physiological myocardial hypertrophy induced by swimming training and the relationship between locally produced and circulating RAS, both ventricular and plasma angiotensin (Ang) I and II contents, ventricular angiotensin converting enzyme (ACE) and plasma renin activity (PRA) were detected by means of radioimmunoassay and biochemical method. It was shown that after 5 weeks of swimming, the ventricular wet weight to body weight ratio (V/Bwt) and Ang II in both left and right ventricles and ACE activity increased markedly as compared with the controls (P < 0.05). Furthermore, significantly positive correlation was found between the ventricular Ang II and V/Bwt (r = 0.7721, P < 0.001), while the plasma Ang I and II and PRA remained at the control level. No correlation was found between plasma Ang II and V/Bwt. These above findings suggest that cardiac RAS may play an important role in physiological myocardial hypertrophy and to a large extent is in dependent on circulating RAS.

Angiotensin II↗
PubMed1993

1H-NMR studies and secondary structure of the RGD-containing snake toxin, albolabrin.

Albolabrin is a naturally occurring peptide from snake venom containing the sequence Arg-Gly-Asp (RGD). It inhibits platelet aggregation by blocking the binding of fibrinogen to the glycoprotein Gp IIb-IIIa, on the surface of activated platelets. Albolabrin consists of 73 residues with six intramolecular disulphide bonds. The 1H-NMR spectrum of albolabrin has been assigned using homonuclear two-dimensional techniques and its secondary structure determined. Like kistrin and echistatin, two related peptides from snake venom, albolabrin appears to have little regular secondary structure in solution. Several bends and two short distorted beta sheets are observed. The RGD sequence, important for binding to the receptor, lies in a mobile loop joining two strands of one of these beta sheets. This loop undergoes a pH-dependent conformational change.

Amino Acid Sequence↗
PubMed1993

Solid-phase von Willebrand factor contains a conformationally active RGD motif that mediates endothelial cell adhesion through the alpha v beta 3 receptor.

The interaction of von Willebrand factor (vWF) with the alpha v beta 3 integrin of human umbilical vein endothelial cells is dependent on the RGD sequence present at residues 1744-1746 of the mature vWF subunit. We compared vWF and its two dimeric fragments, SpIII (residues 1-1365) and SpII (residues 1366-2050), as adhesion substrates. Solid-phase vWF and SpII supported endothelial cell adhesion, whereas SpIII, which contains the glycoprotein (GP) Ib binding domain, did not. Soluble SpII inhibited adhesion to immobilized ligands, whereas soluble vWF did not, suggesting that exposure of the cell attachment domain involves a conformational modification of vWF. Dendroaspin and albolabrin, two RGD-containing peptides of the disintegrin family, were potent inhibitors of cell adhesion to vWF (IC50 approximately 15 nmol/L). Complete inhibition of endothelial cell adhesion to vWF was obtained in the presence of F(ab')2 of monoclonal antibody 9 to vWF, which blocks vWF binding to platelet GPIIb/IIIa. In contrast, monoclonal antibody 713 to vWF, which blocks its binding to platelet GPIb, did not inhibit cell adhesion to vWF. These results indicate that endothelial cell adhesion to vWF is mediated by an RGD-dependent interaction with alpha v beta 3, but does not seem to involve a GPIb-like receptor, and show the importance of the conformation of the RGD sequence.

Amino Acid Sequence↗
PubMed1993

Differential induction of transcriptionally active p53 following UV or ionizing radiation: defects in chromosome instability syndromes?

Accumulation of p53 protein was seen in the nuclei of mammalian cells following DNA damage caused by ultraviolet radiation (UV), X-ray, or a restriction enzyme. Promoters containing p53-binding sites show a dramatic transcriptional response to DNA damage. The p53 response to X-ray is rapid, reaching a peak at 2 hr after radiation, but is very transitory and reduced in magnitude compared with that seen in response to UV. We find no substantive defect in the p53 response of cells from ataxia telangiectasia or xeroderma pigmentosum complementation group A patients. In contrast, 2 out of 11 primary cultures from Bloom's patients showed a complete absence of p53 accumulation following UV irradiation or SV40 infection and a grossly delayed and aberrant response following X-ray.

Apoptosis↗
PubMed1993

Synthetic RGD peptides derived from the adhesive domains of snake-venom proteins: evaluation as inhibitors of platelet aggregation.

Synthetic peptides based on the RGD domains of the potent platelet aggregation inhibitors kistrin and dendroaspin were generated. The 13-amino-acid peptides were synthesized as dicysteinyl linear and disulphide cyclic forms. In platelet-aggregation studies, the cyclic peptides showed 3-fold better inhibition than their linear equivalents and approx. 100-fold greater potency than synthetic linear RGDS peptides derived from fibronectin. An amino acid substitution, Asp10-->Ala, in the kistrin-based peptide gave a 4-fold decrease in potency in the linear peptide, but produced a 2-fold elevation in the inhibitory activity of the cyclic form, generating a peptide of potency comparable with that of the parent protein.

Adenosine Diphosphate↗
PubMed1993

Mitochondrial RNA abundance in differentiating human colonic epithelial tumor cells estimated through use of a mitochondrial genome map.

We describe a procedure for quantification of mitochondrial (mt) RNA present in total RNA extracts of HT-29 human colonic adenocarcinoma cells grown under conditions for rapid growth (25 mM glucose) or differentiation (25 mM trehalose or 5 mM butyrate). Purified mt DNA was fragmented into specific coding regions using restriction endonuclease sites predicted from HeLa cell mt DNA sequence and probed with either 32P-labelled mt DNA or cDNA made from total RNA of HT-29 or HeLa cells. The amounts of probe that hybridized to various gene-encoded mt DNA fragments or RNA were quantified by laser densitometry. Use of 13 restriction endonucleases revealed that most if not all the mt DNA of HT-29 and K562 leukemic cells was comparable in size to that of HeLa cells. Relative levels of mt RNA from rapidly growing HT-29 and HeLa cells were lower than those measured for differentiated HT-29 cells induced by either trehalose or butyrate. In rapidly growing HT-29 cells and HeLa cells, the highest levels of specific mt RNAs were those encoded by mt DNA sequences immediately flanking the nested promoters and the heavy-strand replication origin (OH). Expression patterns of specific mt RNAs from HT-29 cells treated with butyrate and with trehalose were similar, but not identical. In either case, the mt RNAs that increased the most were those coded by mt DNA sequences located downstream from the light-strand replication origin (OL), suggesting a novel pattern of expression not seen before.

Butyrates↗
PubMed1993

Priming immunization determines T helper cytokine mRNA expression patterns in lungs of mice challenged with respiratory syncytial virus.

Defining the mechanism for the vaccine-enhanced illness associated with respiratory syncytial virus (RSV) is critical for advancing RSV vaccine development. Previous studies in which infants were vaccinated with formalin-inactivated alum-precipitated whole virus did not protect from RSV infection, and those infected had a high incidence of severe illness. In contrast, previous clinical trials evaluating live attenuated RSV showed no associated vaccine-enhanced illness. We have used a mouse model to explore the immunopathogenesis of RSV infection. In this study cytokine mRNA expression was examined using 32P-labeled oligonucleotide probes in Northern blot analyses of polyA RNA extracted from lungs of mice primed with various vaccine preparations then challenged nasally with live RSV. We have shown that upon challenge, priming of mice with inactivated virus or subunit F glycoprotein induced a pattern of cytokine mRNA expression suggesting a dominant Th2-like lymphocyte response (relative increase in IL-4 mRNA expression). In contrast, challenge of mice primed with live RSV by parenteral or mucosal routes induced a Th1-like pattern of cytokine mRNA expression (relative decrease in IL-4 mRNA expression compared to IFN-gamma mRNA expression). Thus, the formulation and route of delivery of vaccine products can influence the pattern of cytokine expression in lung upon RSV challenge.

Animals↗
PubMed1993

The ethanolamine requirement of keratinocytes for growth is not due to defective synthesis of ethanolamine phosphoacylglycerols by the decarboxylation pathway.

We have investigated whether the growth requirement of keratinocytes for ethanolamine is due to defective synthesis of ethanolamine phosphoacylglycerols (EPG) via decarboxylation of serine phosphoacylglycerols. Proliferating keratinocytes readily incorporated [3H]ethanolamine into phosphatidylethanolamine (PE) and [3H]serine into phosphatidylserine (PS) and PE. Non-proliferating keratinocytes in ethanolamine-free medium incorporated [3H]glycerol into phosphatidylcholine (PC), PS and PE in decreasing order of label incorporated. The order of decreasing incorporation of glycerol after addition of ethanolamine to the medium was PC > PE > PS. Incubation of non-proliferating keratinocytes with [3H]serine resulted in incorporation of label into PS and PE. The extent of incorporation of [3H]serine into PS in non-proliferating keratinocytes was not less than that in proliferating cells. Addition of ethanolamine to the medium of non-proliferating keratinocytes did not change the quantity of label incorporated into PS, but resulted in a decrease of label incorporated into PE. When cells were prelabelled overnight with [3H]serine and subsequently incubated in medium containing ethanolamine, the loss of label from PS was inhibited relative to that of control cells incubated in medium without ethanolamine. The activity of PS decarboxylase activity in keratinocyte mitochondria was inhibited by phosphoethanolamine and PE, but not by ethanolamine or CDP-ethanolamine. Both proliferating and non-proliferating keratinocytes incorporated [3H]serine into ether-linked ethanolamine phospholipids. Taken together, the above results suggest that (1) both proliferating and non-proliferating keratinocytes are able to synthesize PE and ether-linked ethanolamine phospholipids from serine, and therefore the ethanolamine-requirement of the cells is not due to a defective decarboxylase pathway; (2) any inability of the decarboxylase pathway to meet cellular EPG requirement is not due to decreased synthesis of serine phospholipids; (3) synthesis of PE via decarboxylation, the major route in nonproliferating keratinocytes, appears to decrease when ethanolamine is made available and the CDP-ethanolamine pathway is functioning; (4) phosphoethanolamine and increased PE produced from the CDP-ethanolamine pathway may inhibit PS decarboxylase activity in the cells and provide a means of coordinating the synthesis of PE by the two pathways to prevent excess production.

Cell Division↗
PubMed1993

Crystal structure of human immunodeficiency virus type 1 reverse transcriptase complexed with double-stranded DNA at 3.0 A resolution shows bent DNA.

The crystal structure of a ternary complex of human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) heterodimer (p66/p51), a 19-base/18-base double-stranded DNA template-primer, and a monoclonal antibody Fab fragment has been determined at 3.0 A resolution. The four individual subdomains of RT that make up the polymerase domains of p66 and p51 are named fingers, palm, thumb, and connection [Kohlstaedt, L. A., Wang, J., Friedman, J. M., Rice, P. A. & Steitz, T. A. (1992) Science 256, 1783-1790]. The overall folding of the subdomains is similar in p66 and p51 but the spatial arrangements of the subdomains are dramatically different. The template-primer has A-form and B-form regions separated by a significant bend (40-45 degrees). The most numerous nucleic acid interactions with protein occur primarily along the sugar-phosphate backbone of the DNA and involve amino acid residues of the palm, thumb, and fingers of p66. Highly conserved regions are located in the p66 palm near the polymerase active site. These structural elements, together with two alpha-helices of the thumb of p66, act as a clamp to position the template-primer relative to the polymerase active site. The 3'-hydroxyl of the primer terminus is close to the catalytically essential Asp-110, Asp-185, and Asp-186 residues at the active site and is in a position for nucleophilic attack on the alpha-phosphate of an incoming nucleoside triphosphate. The structure of the HIV-1 RT/DNA/Fab complex should aid our understanding of general mechanisms of nucleic acid polymerization. AIDS therapies may be enhanced by a fuller understanding of drug inhibition and resistance emerging from these studies.

Base Sequence↗
PubMed1993

Transformation of monoamine oxidase-B primary amine substrates into time-dependent inhibitors. Tertiary amine homologues of primary amine substrates.

A family of N-methylated and N,N-dimethylated alkyl and arylalkylamines was prepared and more than half of the analogues were shown to be time-dependent pseudo-first-order inhibitors of monoamine oxidase-B. Some of the time-dependent inactivators were reversible and others were irreversible with respect to prolonged dialysis following inactivation. Partition ratios ranged from zero to 11,000. These results are rationalized in terms of a combination of an inductive effect and a stereoelectronic effect as a result of hindered rotation of an active site covalent adduct. A molecular mechanics calculation indicates that there is at least 10 kcal/mol of torsional energy to be overcome in order for the enzyme adduct to be released. These findings show that tertiary amine homologues of primary amine substrates of monoamine oxidase are time-dependent inhibitors, and this should be useful in the design of new inactivators of this enzyme.

Alkylation↗