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M Hsu

Publications and source records attributed to M Hsu.

At least 19 recordsLinked to original sources
PubMed2003

Antigenic variations in the CD4 induced sites of the CCR5-tropic, pathogenic SHIVsf162p3 gp120 variants.

In vivo passage of non-pathogenic, CCR5-tropic simian/human immunodeficiency virus (SHIV) - SHIVsf162 resulted in a pathogenic isolate, SHIVsf162p3. In an attempt to characterize envelope (Env)-mediated properties that may contribute to its pathogenicity, major (P3 major) and minor (P3 minor) Env gp120 variants were cloned from the plasma of a SHIVsf162p3-infected animal, and expressed in the context of luciferase reporter viruses. Entry mediated by these envelopes and susceptibility to neutralization by CD4 induced-site (CD4i) antibodies (MAbs) was analyzed in comparison to parental SF162. Sequence analysis revealed that the P3 major and minor variant Envs contained 14 and 17 amino acid changes, respectively, compared with SF162. The rank order of entry mediated by the three envelopes was P3 major > SF162 > P3 minor, whereas the reverse order was observed for susceptibility to neutralization by CD4i MAbs. Since CD4i epitopes overlap the coreceptor (CoR) binding site, these findings suggest that the amino acid changes accumulated upon in vivo passage of SHIVsf162 result in Env gp120 structural rearrangements that modulate the exposure and/or conformation of the CoR binding site. This, in turn, led to increased entry and infectivity of the P3 major variant and may be responsible, in part, for the enhanced pathogenicity of SHIVsf162p3.

Amino Acid Sequence↗
PubMed2002

Pre-treatment with R-lipoic acid alleviates the effects of GSH depletion in PC12 cells: implications for Parkinson's disease therapy.

Oxidative stress is believed to play a key role in the degeneration of dopaminergic neurons in the substantia nigra (SN) of Parkinson's disease (PD) patients. An important biochemical feature of PD is a significant early depletion in levels of the thiol antioxidant compound glutathione (GSH) which may lead to the generation of reactive oxygen species (ROS), mitochondrial dysfunction, and ultimately to subsequent neuronal cell death. In earlier work from our laboratory, we demonstrated that depletion of GSH in dopaminergic PC12 cells affects mitochondrial integrity and specifically impairs the activity of mitochondrial complex I. Here we report that pre-treatment of PC12 cells with R-lipoic acid acts to prevent depletion of GSH content and preserves the mitochondrial complex I activity which normally is impaired as a consequence of GSH loss.

Animals↗
PubMed2001

HIV-1 nucleocapsid protein and the secondary structure of the binary complex formed between tRNA(Lys.3) and viral RNA template play different roles during initiation of (-) strand DNA reverse transcription.

In human immunodeficiency virus type 1 (HIV-1), the tRNA(Lys.3) primer and viral RNA template can form a specific complex that is characterized by extensive inter- and intramolecular interactions. Initiation of reverse transcription from this complex has been shown to be distinguished from subsequent elongation by early pausing events, such as at the +1 and +3 nucleotide positions. One major concern regarding the biological relevance of these results is that most kinetic studies of HIV-1 reverse transcription have been performed using tRNA(Lys.3)-viral (v) RNA complexes that were formed by heat annealing. In contrast, tRNA(Lys.3) in viruses is placed onto the primer binding site by nucleocapsid (NC) sequences of the Gag protein. In this study, we have further characterized the initiation features of reverse transcription in the presence of HIV-1 NC protein. In contrast to results obtained with a heat-annealed tRNA(Lys.3).vRNA complex, we found that polymerization reactions catalyzed by HIV-1 reverse transcriptase did not commonly pause at the +1 nucleotide position when a NC-annealed RNA complex was used, and that this was true regardless whether NC was actually still present during reverse transcription. This activity of NC required both zinc finger motifs, as demonstrated by experiments that employed zinc finger-mutated forms of NC protein (H23C NC and ddNC), supporting the involvement of the zinc fingers in the RNA chaperone activity of NC. However, NC was not able to help reverse transcriptase to escape the +3 pausing event. Mutagenesis of a stem structure within the tRNA(Lys.3). vRNA complex led to disappearance of the +3 pausing event as well as to significantly reduced rates of reverse transcription. Thus, this stem structure is essential for optimal reverse transcription, despite its role in promotion of the +3 pausing event.

Amino Acid Motifs↗
PubMed2001

The hunt for a cure for Parkinson's disease.

Several exciting new scientific advances have been made in the past decade toward both understanding the causes of and finding a cure for Parkinson's disease. Heartened by an acceleration in research findings in the past several years, the government has recently called for an infusion of funds from both the National Institutes of Health and private foundations into this burgeoning area of biomedical research. Most currently available conventional treatments for the disease only temporarily delay symptom presentation while doing nothing to halt disease progression. However, the rapidly accelerating pace of research in this field has left researchers hopeful that Parkinson's will be the first major age-related neurodegenerative disease for which we have a viable cure. In this article, advances in various areas of Parkinson's disease research are reviewed.

Animals↗
PubMed2001

The tetrahymena ribozyme cleaves a 5'-methylene phosphonate monoester approximately 10(2)-fold faster than a normal phosphate diester: implications for enzyme catalysis of phosphoryl transfer reactions.

Single-atom substrate modifications have revealed an intricate network of transition state interactions in the Tetrahymena ribozyme reaction. So far, these studies have targeted virtually every oxygen atom near the reaction center, except one, the 5'-bridging oxygen atom of the scissile phosphate. To address whether interactions with this atom play any role in catalysis, we used a new type of DNA substrate in which the 5'-oxygen is replaced with a methylene (-CH2-) unit. Under (kcat/Km)S conditions, the methylene phosphonate monoester substrate dCCCUCUT(mp)TA4 (where mp indicates the position of the phosphonate linkage) unexpectedly reacts approximately 10(3)-fold faster than the analogous control substrates lacking the -CH2- modification. Experiments with DNA-RNA chimeric substrates reveal that the -CH2- modification enhances docking of the substrates into the catalytic core of the ribozyme by approximately 10-fold and stimulates the chemical cleavage by approximately 10(2)-fold. The docking effect apparently arises from the ability of the -CH2- unit to suppress inherently deleterious effects caused by the thymidine residue that immediately follows the cleavage site. To analyze the -O- to -CH2- modification in the absence of this thymidine residue, we prepared oligonucleotide substrates containing methyl phosphate or ethyl phosphonate at the reaction center, thereby eliminating the 3'-terminal TA4 nucleotidyl group. In this context, the -O- to -CH2-modification has no effect on docking but retains the approximately 10(2)-fold effect on the chemical step. To investigate further the stimulatory influence on the chemical step, we measured the "intrinsic" effect of the -O- to -CH2- modification in nonenzymatic reactions with nucleophiles. We found that in solution, the -CH2- modification stimulates chemical reactivity of the phosphorus center by <5-fold, substantially lower in magnitude than the stimulatory effect in the catalytic core of the ribozyme. The greater stimulatory effect of the -CH2- modification in the active site compared to in solution may arise from fortuitous changes in molecular geometry that allow the ribozyme to accommodate the phosphonate transition state better than the natural phosphodiester transition state. As the -CH2- unit lacks lone pair electrons, its effectiveness in the ribozyme reaction suggests that the 5'-oxygen of the scissile phosphate plays no role in catalysis via hydrogen bonding or metal ion coordination. Finally, we show by analysis of physical organic data that such interactions in general provide little catalytic advantage to RNA and protein phosphoryl transferases because the 5'-oxygen undergoes only a small buildup of negative charge during the reaction. In addition to its mechanistic significance for the Tetrahymena ribozyme reaction and phosphoryl transfer reactions in general, this work suggests that phosphonate monoesters may provide a novel molecular tool for determining whether the chemical step limits the rate of an enzymatic reaction. As methylene phosphonate monoesters react modestly faster than phosphate diesters in model reactions, a similarly modest stimulatory effect on an enzymatic reaction upon -CH2- substitution would suggest rate-limiting chemistry.

Animals↗
PubMed2001

The ontogeny of scarless healing II: EGF and PDGF-B gene expression in fetal rat skin and fibroblasts as a function of gestational age.

Twenty years ago, surgeons noted the ability of early-gestation fetal skin to heal in a scarless manner. Since that time, numerous investigators have attempted to elucidate the mechanisms behind this phenomenon. As a result of this effort, it is now well established that many animals undergo a transition late in development from scarless cutaneous healing to a scar-forming, adultlike phenotype. The authors have been interested in the role played by cytokines known to be involved in the adult wound-healing process and how they relate to scarless repair. They therefore asked the following question: Are genes for epidermal growth factor (EGF) and platelet-derived growth factor-B (PDGF-B) expressed differentially as a function of gestational age in fetal rat skin and dermal fibroblasts? To answer this question, skin from fetal Sprague-Dawley rats (N = 56) at time points that represented both the scarless and scar-forming periods of rat gestation was harvested. In addition, fibroblasts derived from fetal rat skin were cultured in vitro at similar times. These cells were expanded in culture and, when confluent, total ribonucleic acid from both fibroblasts and whole skin was extracted and subjected to Northern blot analysis with probes for EGF and PDGF-B. Results demonstrated that neither EGF nor PDGF-B gene expression changed markedly as a function of gestational age in fetal fibroblasts alone. In whole skin, however, both EGF and PDGF-B demonstrated a marked decrease in gene expression with increasing gestational age. Furthermore, the most striking decrease in gene expression for both cytokines came between 16 and 18 days of gestation-the transition point between scarless and scar-forming repair in the fetal rat. These data suggest that EGF and PDGF may play a role in the mechanism of scarless cutaneous repair. Moreover, it appears that fetal fibroblasts are not the cell type responsible for this differential gene expression. These results raise questions about the unique cytokine milieu likely to be present during the time of scarless healing and the cells that ultimately guide the mechanisms leading to skin regeneration.

Animals↗
PubMed2001

Discoidin domain receptors and their ligand, collagen, are temporally regulated in fetal rat fibroblasts in vitro.

The biochemical regulation of collagen deposition during adult cutaneous wound repair is poorly understood. Likewise, how collagen is perceived and modulated in fetal scarless healing remains unknown. Recently, discoidin domain receptors-1 and 2 (DDR1 and DDR2) with tyrosine kinase activity have been identified as novel receptors for collagen. In light of these findings, it was speculated that the production of collagen receptors DDR1 and DDR2 by fetal fibroblasts may be temporally regulated to correlate with the ontogeny of embryonic scar formation. More specifically, because DDRs directly bind collagen and transmit the signals intracellularly, it was hypothesized that they may play an important role in fetal scarless healing by ultimately regulating and modulating collagen production and organization. As part of a fundamental assessment to elucidate the role of DDRs in scarless fetal wound repair, the endogenous expression of DDR1, DDR2, collagen I, and total collagen, as a function of fetal Sprague-Dawley rat skin fibroblasts of different gestational ages, representing scar-free ( E16.5) periods was determined. Using explanted dermal fibroblasts of gestational days E13.5, E16.5, E18.5, and E21.5 (term gestation = 21.5 days) fetuses (n = 92), [3H]proline incorporation assay and Northern and Western blotting analysis were performed to compare the expressions of these molecules with scar-free and scar-forming stages of embryonic development. These results revealed a pattern of increasing collagen production with increasing gestational ages, whereas DDR1 expression decreased with increasing gestational age. This observation suggests that elevated levels of DDR1 may play an important role in scarless tissue regeneration by early gestation fetal fibroblasts. In contrast, DDR2 was expressed by fetal rat fibroblasts at a similar level throughout gestation. These data demonstrate for the first time the temporal expression of collagen and DDR tyrosine kinases in fetal rat fibroblasts as a function of gestational ages. Overall, these data suggest that differential temporal expression of the above-mentioned molecules during fetal skin development may play an important role in the ontogeny of scar formation. Future studies will involve the characterization of the biomolecular functions of these receptor kinases during fetal wound repair.

Animals↗
PubMed2001

Ontogeny of expression of transforming growth factor-beta 1 (TGF-beta 1), TGF-beta 3, and TGF-beta receptors I and II in fetal rat fibroblasts and skin.

Fetal cutaneous wounds that occur in early gestation heal without scar formation. Although much work has been done to characterize the role of transforming growth factor-beta (TGF-beta) isoforms in the adult wound repair process, their function in fetal scarless wound repair is not well understood. The authors hypothesized that the pattern of expression for TGF-beta isoforms and their receptors may influence the phenotypic transition from scarless to scar-forming repair observed during fetal gestation. Using time-dated fetal Sprague-Dawley rat fibroblasts and unwounded skin at gestational ages 14, 16, 18, and 21 days postcoitum of the scarless (< or =16 days) and scar-forming (>16 days) periods of gestation (term = 21.5 days), the authors analyzed the endogenous messenger RNA (mRNA) levels of TGF-beta 1 and TGF-beta 3 and their signaling receptors TGF-beta-RI and TGF-beta-RII. Northern blot analyses in both fibroblasts and unwounded skin revealed that levels of TGF-beta 1 were not differentially expressed, whereas more TGF-beta 3 mRNA transcript was found in early than in late gestation. Fibroblast expression of TGF-beta-RI showed no substantial differences, whereas expression of TGF-beta-RII increased during gestation. In contrast, expression of both TGF-beta-RI and TGF-beta-RII in unwounded skin showed decreasing levels as a function of gestational age. The differential levels of TGF-beta 1 and TGF-beta 3 suggest that the ratio of these cytokines may provide a predominantly antiscarring or profibrotic signal upon wounding during the scar-free or scar-forming periods of gestation, respectively. Furthermore, lower amounts of the ligand-binding TGF-beta-RII seen in early gestation fibroblasts suggest a decreased ability to perceive ligand during the period of scarless repair.

Animals↗
PubMed2001

Differential expression of transforming growth factor-beta receptors I and II and activation of Smad 3 in keloid fibroblasts.

Keloids represent a dysregulated response to cutaneous wounding that results in an excessive deposition of extracellular matrix, especially collagen. However, the molecular mechanisms regulating this pathologic collagen deposition still remain to be elucidated. A previous study by this group demonstrated that transforming growth factor (TGF)-beta1 and -beta2 ligands were expressed at greater levels in keloid fibroblasts when compared with normal human dermal fibroblasts (NHDFs), suggesting that TGF-beta may play a fibrosis-promoting role in keloid pathogenesis.To explore the biomolecular mechanisms of TGF-beta in keloid formation, the authors first compared the expression levels of the type I and type II TGF-beta receptors in keloid fibroblasts and NHDFs. Next, they investigated the phosphorylation of Smad 3, an intracellular TGF-beta signaling molecule, in keloid fibroblasts and NHDFs. Finally, they examined the regulation of TGF-beta receptor II by TGF-beta1, TGF-beta2, and TGF-beta3 ligands. Our findings demonstrated an increased expression of TGF-beta receptors (types I and II) and increased phosphorylation of Smad 3 in keloid fibroblasts relative to NHDFs. These data support a possible role of TGF-beta and its receptors as fibrosis-inducing growth factors in keloids. In addition, all three isoforms of recombinant human TGF-beta proteins could further stimulate the expression of TGF-beta receptor II in both keloids and NHDFs. Taken together, these results substantiate the hypothesis that the elevated levels of TGF-beta ligands and receptors present in keloids may support increased signaling and a potential role for TGF-beta in keloid pathogenesis.

Activin Receptors, Type I↗
PubMed2001

Place learning in virtual space. III: Investigation of spatial navigation training procedures and their application to fMRI and clinical neuropsychology.

This paper describes the utilization of a desktop virtual environment task, the Computer-Generated (C-G) Arena, in the study of human spatial navigation. First, four experiments examined the efficacy of various training procedures in the C-G Arena. In Experiment 1, participants efficiently located a hidden target after only observing the virtual environment from a fixed position (placement learning). In Experiment 2, participants efficiently located a hidden target after only observing an experimenter search the virtual environment (observational learning). In Experiment 3, participants failed to display a latent learning effect in the virtual environment. In Experiment 4, all training procedures effectively taught participants the layout of the virtual environment, but the observational learning procedure most effectively taught participants the location of a hidden target within the environment. Finally, two experiments demonstrated the application of C-G Arena procedures to neuroimaging (Experiment 5) and neuropsychological (Experiment 6) investigations of human spatial navigation.

Adolescent↗
PubMed2000

The effect of mutations in the HIV-1 nucleocapsid protein on strand transfer in cell-free reverse transcription reactions.

Interactions between the nucleocapsid protein (NC) and reverse transcriptase of HIV-1 have been shown to promote the initiation of reverse transcription. We assayed the effect of NC on later events, using a strand transfer system with donor and acceptor HIV RNA templates and found that the presence of NC resulted in increased synthesis of full-length strand-transferred (FLST) DNA. This effect also occurred with mutated forms of NC that lacked both zinc fingers, or that contained a point mutation (histidine-->cysteine) at amino acid 23. In contrast, NC-derived proteins containing only the proximal or distal zinc fingers, or lacking the N- and C-termini, were all unable to catalyze the synthesis of FLST DNA. Band-shift assays using both the mutated and wild-type forms of these proteins revealed that all the NC proteins promoted strand association between (-) strong-stop DNA [(-)ssDNA] and acceptor RNA. The zinc finger motifs were dispensable for full-length processive reverse transcription, and the N- and C-termini were required; however, all NC domains were dispensable for association of (-)ssDNA and acceptor RNA. This suggests that annealing is a less stringent reaction than DNA polymerization.

Cell-Free System↗
PubMed2000

The use of newborn rats and an adenoviral gene delivery vector as a model system for wound-healing research.

An attractive experimental method to elucidate the role of growth factors and cytokines in cutaneous wound healing would be to overexpress or "knock out" a molecule using a gene delivery vector and observe the impact on the wound repair process. As a first step toward developing an adenoviral gene delivery procedure to study wound repair, the authors injected beta-galactosidase (beta-gal) adenoviruses either subcutaneously or intradermally into the dorsal skin of 10-day-old postnatal Sprague-Dawley rats. Histological analysis and beta-gal staining were used to determine the expression and localization of the transferred gene. Beta-gal expression was observed as early as day 1 and up to day 7 postintradermal injection and day 9 postsubcutaneous injection, with no obvious inflammatory reaction detected at the injection sites. Furthermore, as expected, greater beta-gal expression was observed in the dermis of intradermally injected rats compared with the dermis of subcutaneously injected rats. Next, the authors sought to determine whether cutaneous wounds would heal before dissipation of the transferred gene. They created incisional and excisional wounds on the backs of similar-age rats. They found that incisional wounds closed by day 5 postwounding, whereas excisional wounds closed by day 14 postwounding. Their study demonstrated that an adenoviral vector delivered a gene efficiently into newborn rat skin and maintained the gene expression for at least as long as it would take for an incisional wound to heal. The combined use of newborn rat wound models and an adenoviral vector may provide a useful in vivo system to define the biological roles of growth factors and cytokines involved in the wound repair process. These discoveries may lead to the development of gene therapy approaches for abnormal wound healing.

Adenoviridae↗
PubMed2000

Cellular signaling by tyrosine phosphorylation in keloid and normal human dermal fibroblasts.

Keloids represent a dysregulated response to cutaneous wounding that results in disfiguring scars. Unique to humans, keloids are characterized by an accumulation of extracellular matrix components. The underlying molecular mechanisms of keloid pathogenesis, however, remain largely uncharacterized. Similarly, cellular signaling mechanisms, which may indicate inherent differences in the way keloid fibroblasts and normal human dermal fibroblasts interact with extracellular matrix or other cells, have not been investigated. As part of a fundamental assessment of cellular response to injury in keloid fibroblasts, phosphorylation studies were performed using three different keloid (n = 3) and normal human dermal (n = 3) fibroblast cell lines. These studies were undertaken to elucidate whether keloid and normal human dermal fibroblasts exhibit different tyrosine kinase activity. Initially, distinct tyrosine phosphorylation patterns of keloid and normal human dermal fibroblasts were demonstrated. Next, the phosphorylation patterns were correlated with known molecules that may be important to keloid pathogenesis. On the basis of molecular weight, it was hypothesized that the highly phosphorylated bands seen in keloid fibroblasts represented epidermal growth factor receptor (EGFR); discoidin domain receptor 1 (DDR1); and Shc, an adaptor protein known to bind many tyrosine kinases, including EGFR and DDR1. Individual immunoblotting using EGFR, DDR1, and Shc antibodies revealed greater expression in keloid fibroblasts compared with normal human dermal fibroblasts. These data substantiate for the first time the finding of greater phosphorylation by the above-mentioned molecules, which may be important in keloid pathogenesis.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Integrated temporal regulation of the photorespiratory pathway. Circadian regulation of two Arabidopsis genes encoding serine hydroxymethyltransferase.

The photorespiratory pathway is comprised of enzymes localized within three distinct cellular compartments: chloroplasts, peroxisomes, and mitochondria. Photorespiratory enzymes are encoded by nuclear genes, translated in the cytosol, and targeted into these distinct subcellular compartments. One likely means by which to regulate the expression of the genes encoding photorespiratory enzymes is coordinated temporal control. We have previously shown in Arabidopsis that a circadian clock regulates the expression of the nuclear genes encoding both chloroplastic (Rubisco small subunit and Rubisco activase) and peroxisomal (catalase) components of the photorespiratory pathway. To determine whether a circadian clock also regulates the expression of genes encoding mitochondrial components of the photorespiratory pathway, we characterized a family of Arabidopsis serine hydroxymethyltransferase (SHM) genes. We examined mRNA accumulation for two of these family members, including one probable photorespiratory gene (SHM1) and a second gene expressed maximally in roots (SHM4), and show that both exhibit circadian oscillations in mRNA abundance that are in phase with those described for other photorespiratory genes. In addition, we show that SHM1 mRNA accumulates in light-grown seedlings, although this response is probably an indirect consequence of the induction of photosynthesis and photorespiration by illumination.

Amino Acid Sequence↗
PubMed2000

Cadherin repertoire determines partner-specific gap junctional communication during melanoma progression.

Reduced gap junction activity has long been implicated in tumorigenesis. To elucidate the potential role of intercellular communication in melanoma development, we examined gap junctional capability of melanocytic cells from various stages of tumor progression in coculture models using dye transfer assays. Normal melanocytes coupled with keratinocytes by gap junctional formation, whereas melanoma cells did not. Instead, melanoma cells communicated among themselves and with fibroblasts. This switch in communication partners coincided with a shift from E-cadherin to N-cadherin expression during melanoma development. Forced expression of E-cadherin by adenoviral gene transfer in N-cadherin-expressing melanoma cells restored gap junctional compatibility with keratinocytes. Our data suggest that (1) melanocyte transformation is associated with loss of the pre-existing gap junctional activity with keratinocytes but a concomitant gain of communication with a newly juxtaposed cell type, the fibroblasts, (2) the specificity of gap junctional formation during melanoma development is determined by the cadherin profile on the melanocytic cells and (3) the overall gap junctional activity of melanocytic cells is not reduced with transformation.

Adenoviridae↗
PubMed1999

Distribution of salicylic acid in human stratum corneum following topical application in vivo: a comparison of six different formulations.

Distribution of salicylic acid in human stratum corneum from treatment of six different formulations was assessed by quantitation of drug content in sequentially tape-stripped stratum corneum after a single 2-h dose was applied unoccluded to skin on the ventral forearm of four female subjects. The profile and total amounts of stratum corneum removed in 20 tape-strips varied among different types of formulations. With or without normalization by the total stratum corneum weights removed, the extent of drug delivery to the stratum corneum decreased in the following order: SA (5%) > > SAC (10%), Duofilm (16.7%) > TSSS (2%) > SAO (10%), Salic (2.5%), the percentage in parentheses indicating the salicylic acid concentration in each formulation. The greatest topical bioavailability was observed for the alcoholic solution containing glycerol (SA). The 10% collodion formulation (SAC) was found to deliver an amount of salicylic acid into the stratum corneum 2-fold greater than 10% ointment formulation (SAO). Use of absorption ointment (TSSS) also increased the uptake of salicylic acid into the stratum corneum in comparison with formulations based on simple ointment (SAO) and oil in water (o/w) cream (Salic). The partitioning of salicylic acid from collodion formulations (SAC and Duofilm) appeared to be concentration-independent. The results of this study indicate that topical bioavailability of salicylic acid in the stratum corneum varies substantially among different formulations.

Adhesives↗
PubMed1999

Function and sequence analyses of tumor suppressor gene p53 of CHO.K1 cells.

The tumor suppressor gene p53 plays an important role in guarding genomic integrity. When induced in response to environmental results, the gene product of p53 functions as a transcription factor to transactivate genes involved in arresting the cell cycle and as a facilitator of DNA repair. In contrast, the status of p53 in Chinese hamster ovary (CHO) cells, commonly used as a model system for various studies including those involving the cell cycle and transformation, remains an enigma. In this study, the function and sequence of p53 in CHO.K1 cells were investigated. The level of p53 proteins was elevated on ultraviolet (UV) irradiation of the cells, and the proteins formed specific complexes as probed with DNA containing p53-binding sequences. Its activities toward responsive promoters were inducible by UV in a dose-dependent manner. Although p53 in CHO.K1 contained a single missense mutation at codon 211, the mutation apparently had no effect on the functional properties of the protein. The CHO.K1 cells on X-ray irradiation failed to arrest at G1 phase even when the cells were transfected with a wildtype human p53 gene, indicating that the failure probably was not caused by dysfunction of its p53, but by some other mechanism. This result is consistent with the finding that p21(Waf1/Cip1) is undetectable in UV-treated CHO.K1 cells, whereas Gadd45 is induced by UV light in the cells.

3T3 Cells↗