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X Ji

Publications and source records attributed to X Ji.

At least 55 records · Page 3Linked to original sources
PubMed2000

Residue R216 and catalytic efficiency of a murine class alpha glutathione S-transferase toward benzo[a]pyrene 7(R),8(S)-diol 9(S), 10(R)-epoxide.

Murine class alpha glutathione S-transferase A1-1 (mGSTA1-1), unlike mammalian class alpha GSTs, is the most efficient in the glutathione (GSH) conjugation of the ultimate carcinogenic metabolite of benzo[a]pyrene, (+)-anti-7,8-dihydroxy-9,10-oxy-7,8,9, 10-tetrahydrobenzo[a]pyrene [(+)-anti-BPDE] [Hu, X., Srivastava, S. K., Xia, H., Awasthi, Y. C., and Singh, S. V. (1996) J. Biol. Chem. 271, 32684-32688]. Here, we report the crystal structures of mGSTA1-1 in complex with GSH and with the GSH conjugate of (+)-anti-BPDE (GSBpd) at 1.9 and 2.0 A resolution, respectively. Both crystals belong to monoclinic space group C2 with one dimer in the asymmetric unit. The structures reveal that, within one subunit, the GSH moiety interacts with residues Y8, R14, K44, Q53, V54, Q66, and T67, whereas the hydrophobic moiety of GSBpd interacts with the side chains of F9, R14, M207, A215, R216, F219, and I221. In addition, the GSH moiety interacts with D100 and R130 from the other subunit across the dimer interface. The structural comparison between mGSTA1-1.GSH and mGSTA1-1.GSBpd reveals significant conformational differences. The movement of helix alpha9 brings the residues on the helix into direct interaction with the product. Most noticeable are the positional displacement and conformational change of R216, one of the residues located in helix alpha9. The side chain of R216, which points away from the H-site in the mGSTA1-1.GSH complex, probes into the active site and becomes parallel with the aromatic ring system of GSBpd. Moreover, the guanidinium group of R216 shifts approximately 8 A and forms a strong hydrogen bond with the C8 hydroxyl group of GSBpd, suggesting that the electrostatic assistance provided by the guanidinium group facilitates the ring-opening reaction of (+)-anti-BPDE. The structure of mGSTA1-1. GSBpd is also compared with those of hGSTP1-1[V104,A113].GSBpd, hGSPA1-1.S-benzylglutathione, and mGSTA4-4. 4-S-glutathionyl-5-pentyltetrahydrofuran-2-ol. The comparison provides further evidence that supports the functional roles of R216 and helix alpha9. The lack of mobility of helix alpha9 and/or the lack of electrostatic assistance from R216 may be responsible for the relatively lower activity of hGSTA1-1, mGSTA4-4, and hGSTP1-1 toward (+)-anti-BPDE.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed2000

Catalytic center assembly of HPPK as revealed by the crystal structure of a ternary complex at 1.25 A resolution.

BACKGROUND: Folates are essential for life. Unlike mammals, most microorganisms must synthesize folates de novo. 6-Hydroxymethyl-7, 8-dihydropterin pyrophosphokinase (HPPK) catalyzes pyrophosphoryl transfer from ATP to 6-hydroxymethyl-7,8-dihydropterin (HP), the first reaction in the folate pathway, and therefore is an ideal target for developing novel antimicrobial agents. HPPK from Escherichia coli is a 158-residue thermostable protein that provides a convenient model system for mechanistic studies. Crystal structures have been reported for HPPK without bound ligand, containing an HP analog, and complexed with an HP analog, two Mg(2+) ions, and ATP. RESULTS: We present the 1.25 A crystal structure of HPPK in complex with HP, two Mg(2+) ions, and AMPCPP (an ATP analog that inhibits the enzymatic reaction). This structure demonstrates that the enzyme seals the active center where the reaction occurs. The comparison with unligated HPPK reveals dramatic conformational changes of three flexible loops and many sidechains. The coordination of Mg(2+) ions has been defined and the roles of 26 residues have been derived. CONCLUSIONS: HPPK-HP-MgAMPCPP mimics most closely the natural ternary complex of HPPK and provides details of protein-substrate interactions. The coordination of the two Mg(2+) ions helps create the correct geometry for the one-step reaction of pyrophosphoryl transfer, for which we suggest an in-line single displacement mechanism with some associative character in the transition state. The rigidity of the adenine-binding pocket and hydrogen bonds are responsible for adenosine specificity. The nonconserved residues that interact with the substrate might be responsible for the species-dependent properties of an isozyme.

Adenosine Triphosphate↗
PubMed2000

Methanocarba analogues of purine nucleosides as potent and selective adenosine receptor agonists.

Adenosine receptor agonists have cardioprotective, cerebroprotective, and antiinflammatory properties. We report that a carbocyclic modification of the ribose moiety incorporating ring constraints is a general approach for the design of A(1) and A(3) receptor agonists having favorable pharmacodynamic properties. While simple carbocyclic substitution of adenosine agonists greatly diminishes potency, methanocarba-adenosine analogues have now defined the role of sugar puckering in stabilizing the active adenosine receptor-bound conformation and thereby have allowed identification of a favored isomer. In such analogues a fused cyclopropane moiety constrains the pseudosugar ring of the nucleoside to either a Northern (N) or Southern (S) conformation, as defined in the pseudorotational cycle. In binding assays at A(1), A(2A), and A(3) receptors, (N)-methanocarba-adenosine was of higher affinity than the (S)-analogue, particularly at the human A(3) receptor (N/S affinity ratio of 150). (N)-Methanocarba analogues of various N(6)-substituted adenosine derivatives, including cyclopentyl and 3-iodobenzyl, in which the parent compounds are potent agonists at either A(1) or A(3) receptors, respectively, were synthesized. The N(6)-cyclopentyl derivatives were A(1) receptor-selective and maintained high efficacy at recombinant human but not rat brain A(1) receptors, as indicated by stimulation of binding of [(35)S]GTP-gamma-S. The (N)-methanocarba-N(6)-(3-iodobenzyl)adenosine and its 2-chloro derivative had K(i) values of 4.1 and 2.2 nM at A(3) receptors, respectively, and were highly selective partial agonists. Partial agonism combined with high functional potency at A(3) receptors (EC(50) < 1 nM) may produce tissue selectivity. In conclusion, as for P2Y(1) receptors, at least three adenosine receptors favor the ribose (N)-conformation.

Animals↗
PubMed2000

Dissecting the nucleotide binding properties of Escherichia coli 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase with fluorescent 3'(2)'-o-anthraniloyladenosine 5'-triphosphate.

6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) catalyzes the transfer of pyrophosphate from ATP to 6-hydroxymethyl-7, 8-dihydropterin, the first reaction in the folate biosynthetic pathway. Like other enzymes in the folate pathway, HPPK is an ideal target for development of antimicrobial agents because the enzyme is essential for microorganisms but is absent from humans and animals. Using 3'(2')-o-anthraniloyladenosine 5'-triphosphate as a fluorescent probe, a fluorometric competitive binding assay has been developed for measuring the dissociation constants of various compounds that bind to the ATP site of HPPK. The fluorometric assay has been used to determine the nucleotide specificity and dissect the energetics of the binding of MgATP. The order of affinity of various nucleoside triphosphates for HPPK is MgATP>MgGTP>MgITP>MgXTP approximately MgUTP approximately MgCTP. The affinity of MgATP for HPPK (K(d)=2.6+/-0.06 microM) is 260-fold higher than that of MgGTP and more than 1000-fold higher than those of the other nucleoside triphosphates, indicating that HPPK is highly specific with respect to the base moiety of the nucleotide. The affinity of ATP for HPPK in the presence of Mg(2+) is 15 times that in the absence of Mg(2+), indicating that the metal ion is important for the binding of the nucleotide. Removal of the gamma-phosphate from MgATP reduces its affinity for HPPK by a factor of approximately 21. The affinity of AMP for HPPK is about one third that of ADP and almost the same as that of adenosine. The result suggests that among the three phosphoryl groups of MgATP, the gamma-phosphoryl group is most critical for binding to HPPK and the alpha-phosphoryl group contributes little to the binding of the nucleotide. The affinity of MgATP is 18 times that of MgdATP, indicating that the 2'-hydroxyl group of MgATP is also important for binding. van't Hoff analysis suggests that binding of MgATP is mainly driven by enthalpy at 25 degrees C and the entropy of binding is also in favor of the formation of the HPPK.MgATP complex.

Binding, Competitive↗
PubMed2000

Anilide derivatives of an 8-phenylxanthine carboxylic congener are highly potent and selective antagonists at human A(2B) adenosine receptors.

No highly selective antagonists of the A(2B) adenosine receptor (AR) have been reported; however such antagonists have therapeutic potential as antiasthmatic agents. Here we report the synthesis of potent and selective A(2B) receptor antagonists. The structure-activity relationships (SAR) of 8-phenyl-1, 3-di-(n-propyl)xanthine derivatives in binding to recombinant human A(2B) ARs in HEK-293 cells (HEK-A(2B)) and at other AR subtypes were explored. Various amide derivatives of 8-[4-[[carboxymethyl]oxy]phenyl]-1,3-di-(n-propyl)xanthine, 4a, were synthesized. A comparison of aryl, alkyl, and aralkyl amides demonstrated that simple anilides, particularly those substituted in the para-position with electron-withdrawing groups, such as nitro, cyano, and acetyl, bind selectively to human A(2B) receptors in the range of 1-3 nM. The unsubstituted anilide 12 had a K(i) value at A(2B) receptors of 1.48 nM but was only moderately selective versus human A(1)/A(2A) receptors and nonselective versus rat A(1) receptors. Highly potent and selective A(2B) antagonists were a p-aminoacetophenone derivative 20 (K(i) value 1.39 nM) and ap-cyanoanilide 27 (K(i) value 1.97 nM). Compound 27 was 400-, 245-, and 123-fold selective for human A(2B) receptors versus human A(1)/A(2A)/A(3) receptors, respectively, and 8.5- and 310-fold selective versus rat A(1)/A(2A) receptors, respectively. Substitution of the 1,3-dipropyl groups with 1,3-diethyl offered no disadvantage for selectivity, and high affinities at A(2B) receptors were maintained. Substitution of the p-carboxymethyloxy group of 4a and its amides with acrylic acid decreased affinity at A(2B) receptors while increasing affinity at A(1) receptors. 1, 3-Di(cyclohexylmethyl) groups greatly reduced affinity at ARs, although the p-carboxymethyloxy derivative 9 was moderately selective for A(2B) receptors. Several selective A(2B) antagonists inhibited NECA-stimulated calcium mobilization in HEK-A(2B) cells.

Anilides↗
PubMed2000

Influence of incubation temperature on morphology, locomotor performance, and early growth of hatchling wall lizards (Podarcis muralis).

Eggs of wall lizards (Podarcis muralis) were incubated at three temperatures approaching the upper limit of viability for embryonic development in this species (26, 29, and 32 degrees C) to assess the influence of temperature on various aspects of hatchling phenotype likely affecting fitness. The thermal environment affected size and several morphometric characteristics of hatchling lizards. Hatchlings from eggs incubated at 32 degrees C were smaller (snout-vent length, SVL) than those from 26 and 29 degrees C and had smaller mass residuals (from the regression on SVL) as well as shorter tail, head, and femur relative to SVL. Variation in the level of fluctuating asymmetry in meristic and morphometric traits associated with incubation temperatures was quite high but not clearly consistent with the prediction that environmental stress associated with the highest incubation temperatures might produce the highest level of asymmetry. When tested for locomotor capacity in trials developed at body temperatures of 32 and 35 degrees C, hatchlings from the 32 degrees C incubation treatment exhibited the worst performance in any aspect considered (burst speed, maximal length, and number of stops in the complete run). Repeated measures ANCOVAs (with initial egg mass as covariate) of snout-vent length and mass of lizards at days 0 and 20 revealed significant effects of incubation temperature only for mass, being again the hatchlings from eggs incubated at 32 degrees C those exhibiting the smallest final size. All together, our results evidenced a pervasive effect of thermal regime during incubation (and hence of nest site selection) on hatchling phenotypes. However, incubation temperature does not affect hatchling phenotypes in a continuous way; for most of the analysed traits a critical threshold seems to exist between 29 and 32 degrees C, so that hatchlings incubated at 32 degrees C exhibited major detrimental effects. J. Exp. Zool. 286:422-433, 2000.

Adaptation, Physiological↗
PubMed2000

Smad1 domains interacting with Hoxc-8 induce osteoblast differentiation.

Bone morphogenetic proteins are potent osteotropic agents that induce osteoblast differentiation and bone formation. The signal transduction of bone morphogenetic proteins has recently been discovered to involve Smad proteins. Smad1 is an essential intracellular component that is specifically phosphorylated by bone morphogenetic protein receptors and translocated into the nucleus upon ligand stimulation. Previously, we have reported that Smad1 activates osteopontin gene expression in response to bone morphogenetic protein simulation through an interaction with a homeodomain transcription factor, Hoxc-8. In the present study, the interaction domains between the two proteins were characterized by deletional analysis in both yeast two-hybrid and gel shift assays. Two regions within the amino-terminal 87 amino acid residues of Smad1 were mapped to interact with Hoxc-8, one of which binds to the homeodomain. Overexpression of recombinant cDNAs encoding the Hoxc-8 interaction domains of Smad1 effectively activated osteopontin gene transcription in transient transfection assays. Furthermore, stable expression of these Smad1 fragments in 2T3 osteoblast precursor cells stimulated osteoblast differentiation-related gene expression and led to mineralized bone matrix formation. Our data suggest that the interaction of amino-terminal Smad1 with Hoxc-8 mimics bone morphogenetic protein signaling and is sufficient to induce osteoblast differentiation and bone cell formation.

Alkaline Phosphatase↗
PubMed2000

Optimized separation of pharmacologically active anthraquinones in Rhubarb by capillary electrochromatography.

A capillary electrochromatography (CEC) method with diode-array detection has been developed for the separation of the therapeutically important anthraquinones from Rhubarb extract and commercial traditional Chinese drugs containing Rhubarb. The separation of four major anthraquinones (aloe-emodin, emodin, chrysophanol, and physcion) was optimized with respect to pH and concentration of buffers, addition of acetonitrile, applied voltage, and column temperature. Baseline separation was achieved for the four anthraquinones in less than 12 min using a background electrolyte consisting of 5 mM acetic acid (pH 4.5) with 80% acetonitrile. The possibility of CEC for the analysis of traditional Chinese medicines was discussed.

Anthraquinones↗
PubMed2000

Patterns of gene expression associated with BMP-2-induced osteoblast and adipocyte differentiation of mesenchymal progenitor cell 3T3-F442A.

The pluripotent mesenchymal stem cells give rise to osteoblasts, adipocytes, chondrocytes, and myoblasts. The differentiation of these stem cells into each of the mature functional cells may be controlled by a distinctive master gene(s) and is associated with temporal and spatial expression of diverse genes. Identification of genes that are expressed during the differentiation of the mesenchymal cells to osteoblasts is, therefore, important to obtain insights into the molecular mechanisms of osteogenesis. The murine undifferentiated mesenchymal cell 3T3-F442A, when treated with the bone morphogenetic protein 2 (BMP-2), a well-characterized inducer of mesenchymal cell differentiation, exhibited both osteoblastic and adipocytic differentiation. Using the SAGE (serial analysis of gene expression) technique, which has been shown to enable quantitative analysis of large numbers of genes in a simple and quick manner, we obtained 1600 sequence tags representing 2107 individual nucleotide sequences from control and BMP-2-treated 3T3-F442A cells, respectively. By comparing the frequency of tag occurrence, we found profiles of up- or downregulated genes associated with osteoblast or adipocyte phenotype such as type I collagen, osteonectin and OSF-2, or C/EBPbeta, aP2, fatty acid synthase, and lipoprotein lipase, respectively, in BMP-2-treated 3T3-F442A cells. Our data show that BMP-2 induces not only osteoblastic but also adipocytic differentiation in the 3T3-F442A cells. They also show that the 3T3-F442A cells have bipotentials of differentiating toward osteoblasts and adipocytes. The results, therefore, might explain the inverse correlation between trabecular bone volume and fat volume in the bone marrow cavity. The results also suggest that the SAGE may be a useful technique that allows us a fast and efficient way to generate global and local views of gene expression associated with cellular differentiation of the mesenchymal stem cells.

3T3 Cells↗
PubMed2000

Determination of the alkaloid content in different parts of some Mahonia plants by HPCE.

The contents of three quaternary alkaloids (berberine, palmatine, jatrorrhizine) in different parts of the genus Mahonia were determined by high-performance capillary electrophoresis (HPCE). The background electrolyte system composed of 0.1 M phosphate buffer (pH 7.0)-methanol (2:1 V/V) was found to be the most suitable solution for this separation. Brucine was used as internal standard. The linear calibration ranges were 0.004986-0.4986 mg ml-1 (r = 0.9990, n = 5) for berberine, 0.005049-0.5049 mg ml-1 (r = 0.9996, n = 5) for palmatine, and 0.005058-0.5058 mg ml-1 (r = 0.9984, n = 5) for jatrorrhizine. The relative standard deviations were 1.56%, 1.02%, and 1.60% for berberine, palmatine, and jatrorrhizine (n = 6), respectively. The recoveries were determined to be 96.00-101.66% for berberine, 100.15-102.97% for palmatine, and 96.68-102.44% for jatrorrhizine. By using proposed HPCE method, three alkaloids were well-separated within only 5.0 min.

Alkaloids↗
PubMed2000

Dietary 4-HPR suppresses the development of bone metastasis in vivo in a mouse model of prostate cancer progression.

The effects of the synthetic retinoid N-(4-hydroxyphenyl) retinamide (4-HPR) on prostate cancer metastasis in vivo were evaluated in the mouse prostate reconstitution (MPR) model. MPRs were produced by infection of either heterozygous (+/-) or nullizygous (-/-) p53-mutant fetal prostatic epithelial cells with the recombinant retrovirus Zipras/myc 9. Previous studies have documented that loss of p53 function potentiates metastasis in this model system. MPRs were grafted into homozygous (+/+) p53 male mice, fed a 4-HPR containing diet or a control diet and maintained until the status of tumor progression dictated sacrifice. Under these experimental conditions, treatment with 4-HPR did not have a significant effect on primary tumor wet weight for either p53 +/- or p53 -/- MPRs. For, p53 +/- MPRs the animals fed the 4-HPR diet had a slight improvement in survival and a significant reduction in the number of mesenteric metastases (P = 0.0477, t-test). Notably, in p53 +/- MPRs the incidence of metastasis to lumbar spine and sternum was 92% in the control animals compared to 54% in the 4-HPR treated animals (P = 0.035, chi2-test). In p53 -/- MPRs there was a trend toward a reduction in the number of soft tissue metastases to lung and liver in the 4-HPR group relative to the control diet group and a statistically significant reduction in the incidence of metastasis to bone was demonstrated in that 50% of control animals versus 30% of 4-HPR treated p53 -/- animals harbored bone metastases (P = 0 < 0.05, chi2-test). Cell lines were established from portions of the primary tumor and from selected metastatic deposits in each experimental group. Clonal analysis, by retroviral integration pattern, indicated increased clonal diversity in both the primary tumors and metastasis-derived cell lines from 4-HPR treated animals relative to the control animals. In vitro treatment with 4-HPR did not reveal discriminating differences between cell lines derived from primary tumors and bone metastases or control and treatment groups in regard to growth arrest or apoptotic responses. Overall these studies indicate limited anti-tumor and anti-metastatic activity in this highly aggressive in vivo mouse model of prostate cancer, yet 4-HPR treatment significantly suppressed the development of bone metastases in p53 +/- and p53 -/- MPRs revealing a novel and potentially clinically useful activity of this retinoid.

Animals↗
PubMed2000

Calibration and LOD/LOQ estimation of a chemiluminescent hybridization assay for residual DNA in recombinant protein drugs expressed in E. coli using a four-parameter logistic model.

Calibration is the process of fitting a model based on reference data points (x, y), then using the model to estimate an unknown x based on a new measured response, y. In DNA assay, x is the concentration, and y is the measured signal volume. A four-parameter logistic model was used frequently for calibration of immunoassay when the response is optical density for enzyme-linked immunosorbent assay (ELISA) or adjusted radioactivity count for radioimmunoassay (RIA). Here, it is shown that the same model or a linearized version of the curve are equally useful for the calibration of a chemiluminescent hybridization assay for residual DNA in recombinant protein drugs and calculation of performance measures of the assay.

Calibration↗
PubMed2000

Interventional treatment of huge hepatic cavernous hemangioma.

OBJECTIVE: To study the methods of interventional treatment of huge hepatic cavernous hemangioma (HCH). METHOD: A total of 14 patients with HCH were treated with lipiodol-ultrafluid (10-15 ml), bleomycin A (PYM 16-32 mg), and gelatin-sponge particles. RESULTS: DSA hepatic arteriography showed multiple vascular lakes in the early arterial phase, so-called "to hang the fruits on the branches", which persisted for a long time. CT scan showed a significant reduction in tumor size in 8 of the 14 patients after the treatment. CONCLUSION: Embolization with lipiodol-ultrafluid, PYM and gelatin sponge particles is one of the best methods for the treatment of HCH.

Adult↗
PubMed2000

[Study on the relationship between CD44v6, p53 gene mutation and ovarian carcinoma metastasis].

OBJECTIVE: To explore the relationship between CD44v6 expression, p53 gene mutation, cell ploidy and ovarian metastasis. METHODS: Eighty-five ovarian specimens (20 normal specimens, 20 benign tumors, 20 non-metastatic carcinomas and 25 metastatic carcinomas) were collected. Cell ploidy and the distribution of cell population in each stage of cell cycle were detected by flow cytometry. By using techniques such as reverse transcription-polymerase chain reaction, southern blot and image analysis, the quantitative and qualitative expression of CD44v6 were determined; the mutation of p53 gene in the ovarian specimens was also determined by polymerase chain reaction-single strand conformation polymorphism with silver staining. RESULTS: In the normal ovarian specimens, benign tumors, non-metastatic and metastatic carcinomas, the positive rates of CD44v6 expression and p53 gene mutation were 0%, 10%, 75%, 88% and 0%, 5%, 40%, 60%, respectively, both increased gradually. The mean dark density in the non-metastatic (3,820 +/- 289) and metastatic (10,132 +/- 1,521) groups was significantly different (P < 0.01). In the positive and negative CD44v6 ovarian carcinoma groups, the mean carcinoma cell percentage at the G2M stage was 5.90% and 5.06% (P > 0.05), mean aneuploidy carcinoma cell percentage was 57% and 50% (P > 0.05) respectively. In the mutated and non-mutated p53 gene groups, their mean cell population at the G2M stage was 11.15% and 5.85% (P < 0.05), mean aneuploidy carcinoma cell percentage was 74% and 36% (P < 0.05), respectively. CONCLUSIONS: The expression of CD44v6 is closely related to carcinoma metastasis. The relationship between CD44v6 expression, p53 gene mutation and carcinoma metastasis can be involved in the different mechanism. As compared to mutated p53 gene, CD44v6 appears to be more suitable as a metastatic marker.

Adult↗
PubMed2000

[Morphologic analysis of 14 cases of hepatic angiomyolipoma].

OBJECTIVE: To study the morphology of hepatic angiomyolipoma (HAML). METHODS: HE staining, PAS staining, immunohistochemistry and electronic microscopy were used to study 10 cases from our department and 4 cases from consultation. RESULTS: All 14 HAML cases showed perivascular epithelioid cells (PEC) with 4 different types of morphology. They presented characteristic PAS positivity premelanosome like structures and dense body under electronic microscopy. Immunohistochemistry demonstrated positive reaction for both actin and HMB45. CONCLUSION: Light and electronmicroscopic features and immunophenotype of HAML are characteristic of PEC related lesion.

Actins↗
PubMed2000

[Rotational analysis of fluorescence excitation spectrum of NO2 in the range of 589.3-594.0 nm].

This paper shows the measurement of high-resolution fluorescence excitation spectrum of uncooled NO2 in the range of 589.3-594 nm. The 11 vibronic bands approximately 120 lines in this region were rotationally assigned. The analysis shows that these lines are mainly arise from the parallel transition of 2B2(K' = 0)-->2A2(K" = 0). The spectroscopic parameters describing these bands are obtained. The discussion is given to the complexity of spectrum structure.

Air Pollutants↗
PubMed1999

Extinction of E-cadherin expression in breast cancer via a dominant repression pathway acting on proximal promoter elements.

Inactivation of the E-cadherin cell adhesion molecule is believed critical in the development and behavior of many epithelial cancers, though mutations in the E-cadherin gene account for inactivation in only a fraction of cases. In many breast cancer lines, E-cadherin transcription is extinguished, but the role and significance of alterations in trans-acting transcription factors, promoter hypermethylation, and chromatin changes remain unresolved. To gain further insights into mechanisms underlying E-cadherin inactivation in breast cancer, we analysed somatic cell hybrids resulting from pairwise fusions between breast cancer lines with intact E-cadherin transcription (E-cad+) and lines lacking E-cadherin transcription (E-cad-). All hybrid lines failed to express E-cadherin transcripts and protein, despite the fact that E-cadherin alleles from E-cad+ lines were present in the hybrids. Elements in the proximal 108 bp of the E-cadherin promoter, when present in reporter gene constructs, were sufficient to direct strong transcription in E-cad+ breast lines, but displayed weak activity in E-cad- parental lines and hybrids. E-cadherin expression could not be restored in E-cad- lines or hybrids by treatment with a DNA demethylating agent and/or a histone deacetylase inhibitor. Our findings suggest loss of E-cadherin expression in some breast cancers may be due to dominant repression of the trans-acting pathways that regulate E-cadherin transcription.

Base Sequence↗