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Biomedical subjects

Q Li

Publications and source records attributed to Q Li.

At least 703 records · Page 39Linked to original sources
PubMed1996

Oxidative stress activates metal-responsive transcription factor-1 binding activity. Occupancy in vivo of metal response elements in the metallothionein-I gene promoter.

Oxidative stress (tert-butylhydroquinone) rapidly induced metallothionein-I gene expression in mouse Hepa cells, and this effect was mediated predominantly through metal response promoter elements in transient transfection assays. In vivo genomic footprinting of the mouse metallothionein-I promoter after treatment of Hepa cells with hydrogen peroxide, tert-butylhydroquinone, or zinc suggested a rapid increase in occupancy of the metal response elements. More subtle changes also occurred in the constitutive genomic footprint at the composite major late transcription factor/antioxidant response element. This element may, in part, mediate induction by hydrogen peroxide. Electrophoretic mobility shift assays demonstrated a rapid (30 min) increase in the DNA binding activity of metal-responsive transcription factor-1 in Hepa cells treated with any of these inducers. In control cells, upstream stimulatory factor binding with the major late transcription factor site, and a nuclear protein complex distinct from AP-1, but specific for the antioxidant response element, were detected. The amounts of these complexes were not altered after these treatments. These studies indicate that metal-responsive transcription factor-1 plays a role in activating mouse metallothionein-I gene transcription in response to reactive oxygen species.

Animals↗
PubMed1996

Physical and biological characteristics of the antitumor drug actinomycin D analogues derivatized at N-methyl-L-valine residues.

The crystal structure of the DNA-actinomycin D (AMD) complex and a simple molecular modeling study indicated that AMD analogues derivatized at N-methyl-L-valine residues (fifth amino acid residue in the cyclic depsipeptide of AMD) could bind to DNA as strongly as the parent AMD. The analogues in which N-methyl-L-valine residues were replaced with L- and D-forms of N-methylvalines, N-methylthreonines, N-methylphenylalanies, N-methyltyrosines, and N-methyl-O-methyltyrosines have been totally synthesized. The characteristics of binding of the analogues to various DNAs including DNA-1 [d(TATATATGCATATATA)], DNA-2 [d(TATATACGCGTATATA)], DNA-3 [d(ATATATAGCTATATAT)], and DNA-4 [d(ATATATGGCCATATAT)] have been examined by using visible absorption spectrum methods. The association constants calculated from the absorption spectra indicate that the modifications of the N-methyl-L-valine residues in the AMD molecule do affect the DNA binding characteristics of the analogues. The L-aromatic analogues bind slightly better than the L-aliphatic analogues except for binding to DNA-1 (-TGCA-), whereas the D-aliphatic analogues bind consistently better than the D-aromatic analogues. In the L-form analogues, the L-Tyr analogue has the highest overall association constant, whereas the D-Val analogue has the highest association constant among the D-form analogues. In spite of substitution of bulky aromatic groups, the D-aromatic analogues bind to the DNA-1 quite well. However, D-aromatic analogues have significantly reduced their binding capacities to the other DNAs, indicating that the substitution of the D-aromatic residues creates a unique four-base sequence preference (-TGCA-). The RNA polymerase inhibitory activities of the AMD analogues in vivo have been examined using human cells (HeLa). All AMD analogues except for the L-Thr analogues severely inhibit RNA synthesis at relatively low drug concentrations. The D-Val, L-OMT, L-Phe, and D-Phe analogues inhibit RNA synthesis more strongly than the natural antibiotic (AMD itself).

Antibiotics, Antineoplastic↗
PubMed1996

The characterization of a human RHAMM cDNA: conservation of the hyaluronan-binding domains.

A full-length human RHAMM cDNA clone was isolated by a combination of screening a human breast cDNA expression library with the murine RHAMM 2 cDNA as well as 5' RACE and RT-PCR using messenger RNA from human breast cell line (MCF-10A). The full-length cDNA contained 725 aa that encoded an 84 kDa protein. Although the coding region of the human RHAMM cDNA resembles the murine RHAMM v4, it has additional unique N-terminal (489 bp) and C-terminal (33 bp) regions. Also, only 1 of 5 repeat sequences encoded in the murine cDNA are present in human cDNA. The overall homology between the overlapping region of human and mouse RHAMM v4 cDNA clone is 85%, but the HA binding motif (B[X7]B), shown to be critical for the signaling capability of this receptor, is 100% conserved.

Amino Acid Sequence↗
PubMed1996

Abrogation of c-kit/Steel factor-dependent tumorigenesis by kinase defective mutants of the c-kit receptor: c-kit kinase defective mutants as candidate tools for cancer gene therapy.

The growth and survival of many types of cancer cells are known to be supported by specific growth factor/cytokine systems. Among these, the activation of c-kit receptor and its ligand steel factor participates in several types of human carcinogenesis. W mutations of laboratory mouse strains are loss of functional mutations of the c-kit receptor. To examine the validity of these mutants in investigating c-kit-mediated carcinogenesis and in the treatment of c-kit-dependent tumors, we introduced various W mutations (W, Wv, and W42) into a transgenic mouse strain carrying human papillomavirus oncogenes, in which c-kit/Steel-mediated tumorigenesis occurs with a very high incidence. In all transgenic strains carrying a W mutation, the c-kit deficiency affected the tumorgenic process to various degrees. Tumor development was markedly suppressed in transgenic strains carrying kinase defective mutations (Wv and W42) in a heterozygous condition. In null-type (W) heterozygous transgenic mice, tumorigenesis was suppressed at a lower level. Moreover, minimal focal legions or, in some cases, no focal legions were found in the testes of W/Wv heterozygous transgenic mice, showing a close relationship between tumor cell growth and the degree of c-kit inactivation. These results indicated that c-kit activity is a pivotal determinant of testicular tumor development and that the kinase defective mutants of c-kit are valuable for treating c-kit-dependent cancer, as well as for clarifying the c-kit-mediated carcinogenesis.

Animals↗
PubMed1996

Imaging ROMK1 inwardly rectifying ATP-sensitive K+ channel protein using atomic force microscopy.

The inwardly rectifying K+ channel ROMK1 has been implicated as being significant in K+ secretion in the distal nephron. ROMK1 has been shown by immunocytochemistry to be expressed in relevant nephron segments. The development of the atomic force microscope has made possible the production of high resolution images of small particles, including a variety of biological macromolecules. Recently, a fusion protein of glutathione S-transferase (GST) and ROMK1 (ROMK1-GST) has been used to produce a polyclonal antibody for immunolocalization of ROMK1. We have used atomic force microscopy to examine ROMK1-GST and the native ROMK1 polypeptide cleaved from GST. Imaging was conducted with the proteins in physiological solutions attached to mica. ROMK1-GST appears in images as a particle composed of two units of similar size. Analyses of images indicate that the two units have volumes of approximately 118 nm3, which is close to the theoretical volume of a globular protein of approximately 65 kDa (the molecular mass of ROMK1-GST). Native GST exists as a dimer, and the images obtained here are consistent with the ROMK1-GST fusion protein's existence as a heterodimer. In experiments on ROMK1 in aqueous solution, single molecules appear to aggregate, but contact to the mica was maintained. Addition of ATP to the solution produced a change in height of the aggregates. This change (which was reversible) suggests that ATP induces a structural change in the ROMK1 protein. The data show that atomic force microscopy is a useful tool for examination of purified protein molecules under near-physiological conditions, and furthermore, that structural alterations in the proteins may be continuously investigated.

Adenosine Triphosphate↗
PubMed1996

Synthesis and structure-activity relationships of 2-pyridones: a novel series of potent DNA gyrase inhibitors as antibacterial agents.

Two novel series of 2-pyridones were synthesized by transposition of the nitrogen of 4-quinolones to the bridgehead position. This subtle interchange of the nitrogen atom with a carbon atom yielded two novel heterocyclic nuclei, pyrido[1,2-alpha]pyrimidine and quinolizine, which had not previously been evaluated as antibacterial agents and were found to be potent inhibitors of DNA gyrase. Quinolizines with a methyl group at the 9-position such as (S)-45a (ABT-719) demonstrate exceptional broad spectrum antibacterial activity. Most notably, they are active against resistant bacteria such as methicillin-resistant Staphylococcus aureus, vancomycin-resistant strains of enterococci, and ciprofloxacin-resistant organisms. In addition, 2-pyridones also possess favorable physiochemical and pharmacokinetic properties. These 2-pyridones were synthesized from the commercially available starting materials by 10-17 linear transformations. The structure of an adduct yielded by this sequence, (S)-45a (ABT-719), was determined by X-ray crystallographic analysis.

Animals↗
PubMed1996

Prostaglandin E2 stimulates a Ca2+-dependent K+ channel in human erythrocytes and alters cell volume and filterability.

To understand the mechanism by which human red blood cells (RBCs) contribute to hemostasis and thrombosis, we have examined the effects of metabolites released by activated platelets on intact RBCs. Prostaglandin E2 (PGE2), a signal molecule produced by activated platelets, was observed to lower the filterability of human erythrocytes by approximately 30% at 10(-10) M. PGE2 also caused a reduction in mean cell volume of approximately 10%. The shrinkage of red cells after PGE2 treatment was confirmed by documenting a decrease in osmotic fragility and an increase in cell density following exposure to the hormone. Careful analysis, however, revealed that only approximately 15% of the erythrocytes responded to stimulation with PGE2. Examination of the cause of cell shrinkage showed that induction of a PGE2-stimulated K+ efflux pathway leading to rapid loss of cellular K+ was responsible. The PGE2-stimulated K+ loss was also observed to be Ca2+-dependent, suggesting the possible involvement of the Gardos channel. Gardos channel participation was supported by the observation that two Gardos channel inhibitors, charybdotoxin and clotrimazole, independently blocked the PGE2-stimulated K+ efflux. Further evidence for Gardos channel activation came from experiments aimed at characterizing the efflux pathway followed by the obligatory counterion. Thus, K+ efflux was readily stimulated even when NO3- was substituted for Cl-, suggesting that neither KCl cotransport nor Na/K/2Cl cotransport plays a prominent role in the PGE2-induced cell shrinkage. Further, the anion transporter band 3 was implicated as the counterion efflux route, since DIDS inhibited the PGE2-stimulated cell volume change without blocking the change in membrane potential. Taken together, we propose that release of PGE2 by activated platelets constitutes part of a mechanism by which activated platelets may recruit adjacent erythrocytes to assist in clot formation.

Calcium↗
PubMed1996

In vivo search for butyrate responsive sequences using transgenic mice carrying A gamma gene promoter mutants.

We describe an in vivo approach, in transgenic mice, aimed to identify promoter elements responsible for the induction of gamma globin expression by butyrate. Transgenic lines carrying human A gamma gene promoter truncations at position -141, -201, -382, and -730 A gamma were treated with alpha amino butyric acid (alpha ABA), and effects on gamma globin expression were analyzed at the messenger RNA level. No induction of gamma gene expression was observed in animals carrying promoters truncated at positions -141, -201, or -382 A gamma, suggesting either that butyrate response elements (BRE) are not located in the proximal gamma gene promoter or, if they were, they require the cooperation of upstream sequences for gamma gene induction. Two animals from one line carrying the -730 A gamma truncation responded to alpha ABA treatment with significant increases in gamma gene expression, indicating that a BRE is located between position -382 and -730 region of the A gamma gene promoter. Because the maximum induction by alpha ABA is observed in transgenic mice carrying a A gamma gene promoter extending to nucleotide -1350, it is likely that another butyrate responsive element is located between -730 and -1350 of the A gamma gene promoter. These results indicate that the transgenic mouse model can be used for identification of DNA regions that contain cis elements involved in gamma globin gene inducibility.

Aminobutyrates↗
PubMed1996

Constitutively activated JNK is associated with HTLV-1 mediated tumorigenesis.

Human T cell leukemia virus type I (HTLV-1) is the etiologic agent of adult T-cell leukemia (ATL) and HTLV-1 associated myelopathy, also called tropical spastic paraparesis (HAM/TSP). Both clinical and in vitro evidence have demonstrated that the virus or its transactivator Tax, are transforming. However, transformation appears to require additional, as yet poorly characterized, genetic changes in infected cells. JNK is a recently characterized member of the MAP kinase family. Its signaling cascade is distinct from other members and has been demonstrated to play an important role in T-cell activation, at least partially through its downstream targets, c-jun and ATF-2. Here we demonstrate constitutive activation of the JNK cascade in human lymphocytes transformed in vitro by HTLV-1 and also in Tax transformed murine fibroblasts. Such activation is not induced by Tax expression alone, and occurs only when infected lymphocytes become IL-2 independent or immortalized. Constitutive JNK activation was also found in leukocytes isolated from ATL patients. The acquisition of constitutive JNK activation may represent an important later event in HTLV-1 tumorigenesis.

Animals↗