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Query: UMLS:C0027651 (tumor)
 685,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have studied the mechanisms that regulate the expression of the mouse gene encoding steroid 11 beta-hydroxylase (11 beta-OHase), a steroidogenic cytochrome P450 enzyme that is expressed only in the adrenal cortex. DNase I footprinting and gel-mobility shift analyses revealed potential regulatory elements at -370 and -310 in the 11 beta-OHase promoter region. To determine the contributions of these elements to expression, we altered their sequences by site-selected mutagenesis and studied promoter activity after transfection into Y1 mouse adrenocortical tumor cells. Mutation of either element markedly decreased basal promoter activity but did not affect the response to treatment with 8-bromo cAMP. These experiments thus document the functional roles of these elements, within the context of the intact promoter, in constitutive expression of 11 beta-OHase. Moreover, addition of either of these elements to p-40GH, a 5'-deletion plasmid containing 11 beta-OHase sequences from -40 to +8 upstream of a growth hormone reporter gene, significantly increased promoter activity but did not confer cAMP responsiveness. Finally, increased expression was seen after transfection of Y1 derivatives deficient in cAMP-dependent protein kinase, indicating that neither element required cAMP-dependent protein kinase activity. These studies thus define two regulatory elements that play important roles in 11 beta-OHase expression.
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PMID:Identification and characterization of two upstream elements that regulate adrenocortical expression of steroid 11 beta-hydroxylase. 223 42

Steroid 21-hydroxylase (21-OHase) is specifically expressed at high levels in the adrenal cortex, where it is required for the synthesis of mineralocorticoids and glucocorticoids. In this study, we have investigated the regulatory elements in the 21-OHase promoter region which contribute to the expression of this gene in Y1 adrenocortical cells. Eight potential regulatory elements in the 5'-flanking region of the 21-OHase gene were identified by DNase I footprinting and gel mobility shift experiments. Some of these footprints were produced by nuclear extracts from many cell lines, whereas other interactions were seen only when using nuclear extracts from Y1 adrenocortical and MA-10 Leydig tumor cells. Mutation of most of the elements markedly decreased the expression of a 21-OHase gene transfected into Y1 cells, thus documenting their functional importance for expression. Moreover, oligonucleotides containing the sequences of two related elements at -65 and -210, which share the heptamer AGGTCAG, increased the activity of a heterologous promoter in a Y1 cell-specific manner. Collectively, these results demonstrate that expression of 21-OHase in Y1 adrenocortical cells requires interactions among multiple cis-acting elements and regulatory proteins.
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PMID:Multiple regulatory elements determine adrenocortical expression of steroid 21-hydroxylase. 233 16

The cholesterol side-chain cleavage enzyme (SCC) catalyzes the initial and rate-limiting step in the synthesis of steroid hormones. The mouse gene encoding SCC was cloned and the nucleotide sequence of its 5'-flanking region determined. This sequence includes an AP-1 motif at -319 and two motifs, AGGTCA at -70 and AGCCTTG at -40, that match elements proposed to be important in the expression of steroid 21-hydroxylase. When transfected into mouse Y1 adrenocortical tumor cells, 1.5 kilobase pairs of 5'-flanking region of the SCC gene directed high levels of expression of a growth hormone reporter gene; treatment of the transfected Y1 cells with 8-bromo-cAMP increased this expression by 5-fold. In contrast, transfected mouse MA-10 Leydig cells showed appreciably lower expression, suggesting that SCC expression in Leydig cells requires additional elements not contained in the 5'-flanking region of the SCC gene used in these experiments. Deletion experiments showed that 424 base pairs of 5'-flanking sequences were sufficient for regulated expression in Y1 cells and mapped two regulatory regions: one from -424 to -327 and a second from -219 to -77. DNase I footprinting and gel mobility shift analyses of these 424 base pairs defined several interactions between nuclear proteins and the SCC promoter, including footprints centered over the AP-1 motif, over a sequence at -120, and over the sequences (-70 and -40) that resemble 21-hydroxylase promoter elements. Finally, site-selected mutagenesis of the potential elements at -40, -70, or -120 decreased SCC promoter activity in transfected Y1 adrenocortical cells, thus establishing their importance in SCC expression.
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PMID:Analysis of the promoter region of the gene encoding mouse cholesterol side-chain cleavage enzyme. 236 94

The murine thyrotropic MGH101A tumor is characterized by absent thyrotropin (TSH) beta gene expression and altered thyroid hormone (T3) regulation of the alpha-subunit. Comparison of the promoter structures of both alpha and TSH beta subunit genes from MGH101A with the promoter in expressing TtT-97 thyrotropes revealed no detectable differences. Transfection of the TSH beta promoter from MGH101A linked to luciferase showed minimal expression in primary or cloned MGH101A cells, or L-cells. However, a 6- to 10-fold increase in expression was exhibited in transfected thyrotropes. For the alpha gene, promoter activity was highest in thyrotropes and in cloned MGH101A cells, 5-fold lower in MGH101A tumors, and 10-fold lower in L-cells. Both promoters were not substantially affected by T3 treatment in MGH101A cells. In thyrotropes, promoter activity was inhibited 62.5% and 57.7% by 10 nM T3 treatment for the TSH beta and alpha genes, respectively. DNase I protection showed that factors from TtT-97 but not from MGH101A cells interacted with regions in the TSH beta promoter, while nuclear extracts from each tumor demonstrated at least one protein-DNA interaction with the alpha-subunit promoter. These studies suggest that the molecular defects in the MGH101A tumor are related to the absence of trans-acting factors and are not a result of altered primary gene structure.
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PMID:TSH subunit gene promoters from a murine alpha-subunit producing tumor function normally. 237 87

We have examined the chromatin structure of the 5'-flanking region of the albumin and alpha-fetoprotein (Afp) genes in different developing rat tissues and cloned cell lines that display various functional states of these genes. Nuclease-hypersensitive sites were probed with DNase I, using an indirect end-labeling technique. In albumin-producing rat cells two major DNase I-hypersensitive sites were found near the promoter region and one additional site was located approximately 3 kilobases (kb) upstream. Similarly, in Afp-producing rat tissues and cell lines we mapped one DNase I-hypersensitive region close to the promoter region and two cleavage sites further upstream at approximately 2.2 and approximately 3.8 kb from the cap site. The DNase I-hypersensitive sites of both genes were absent in nonhepatic rat cells and therefore appear to be tissue specific. Loss of specific sets of DNase I-hypersensitive sites accompanies the cessation of transcription for the Afp gene in adult rat liver and in a "dedifferentiated" hepatoma cell line. Likewise, specific sets of DNase I-hypersensitive sites disappear during the inactivation of the albumin gene in hepatoma cells. The distal upstream sites of the Afp and albumin genes display the same DNase I sensitivity in expressing and potentially expressible states. These findings suggest that reversible changes in short chromatin regions may be involved in the actual transcription of the albumin and Afp genes, while more permanent tissue-specific changes at other sites correlate with the capacity of these genes to be expressed during hepatic differentiation and neoplasia.
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PMID:Specific sets of DNase I-hypersensitive sites are associated with the potential and overt expression of the rat albumin and alpha-fetoprotein genes. 243 25

As in tumors with c-myc chromosomal translocations, c-myc retrovirus-induced monocyte tumors constitutively express an activated form of c-myc (the proviral gene), whereas the normal endogenous c-myc genes are transcriptionally silent. Treatment of these retrovirus-induced tumor cells with a number of bioactive chemicals and growth factors that are known to induce c-myc expression in cells of the monocyte lineage failed to induce the endogenous c-myc gene. In contrast, the same treatments induced the c-fos gene in both tumors and a control macrophage line. To investigate c-myc suppression further, a normal copy of the human c-myc gene was introduced into tumor and control cell lines by using a retrovirus with self-inactivating long terminal repeats. This transduced normal gene was expressed at equivalent levels in all cells, regardless of the state of endogenous c-myc gene expression, and was strongly induced by agents that induce the normal gene in the control cells. These results indicate that the signal transduction pathways that normally activate the c-myc gene are functional in myc-induced tumor cells and suggest that endogenous c-myc is actively suppressed. An examination of the c-myc locus itself showed that the lack of transcriptional activity correlated with the absence of several prominent DNase I-hypersensitive sites in the 5'-flanking region of the gene but without loss of general DNase sensitivity. Furthermore, analysis of 22 methylation-sensitive restriction enzyme sites in the 5'-flanking region, first exon, and first intron indicated that the silent c-myc genes remained in the same unmethylated state as did actively expressed genes. Thus, c-myc suppression does not appear to result from the most frequently described mechanisms of gene inactivation.
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PMID:Germ line c-myc is not down-regulated by loss or exclusion of activating factors in myc-induced macrophage tumors. 247 87

The fibrosarcoma IC9 is deficient in the expression of the major histocompatibility complex class I genes Kb, Kk, and Dk and expresses only the Db molecule. Because class I deficiency may enable tumor cells to escape the immune response by cytotoxic T lymphocytes, we investigated why the class I genes are not expressed. Expression of the silent class I genes could not be induced, but all known DNA-binding factors specific for class I genes could be detected in nuclear extracts of IC9 cells. After cloning of the silent Kb gene from the IC9 cells and subsequent transfection of this cloned Kb gene into LTK- and IC9 cells, normal Kb antigens were expressed on the cell surface of both cell lines. Digestion of the chromatin of IC9 cells with micrococcal nuclease and DNase I showed a decreased nuclease sensitivity of the silent class I genes in comparison with active genes and the absence of DNase I hypersensitive sites in the promoter region of the silent Dk gene. These findings demonstrate that class I expression is turned off by a cis-acting regulatory mechanism at the level of the chromatin structure.
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PMID:Major histocompatibility complex class I genes in murine fibrosarcoma IC9 are down regulated at the level of the chromatin structure. 250 38

MHC class I antigens play a crucial role in immunological functions, e.g. transplant and tumor rejection and antigen presentation. Whereas class I antigens are normally expressed on most adult tissues, albeit in varying amounts, embryonic as well as many tumor cells are characterized by the absence of major histocompatibility complex class I antigens on their cell surfaces. In this study the mechanism controlling the lack of class I expression was analyzed at the level of the chromatin structure. Five DNase I hypersensitive sites were determined at the H-2 D-locus of cell lines constitutively expressing class I genes. Two of them (DH1, located at the TATA box/transcription start site, and DH2, located at the enhancer/interferon response sequence) were absent in the fibrosarcoma IC9 in which expression of the silent Dk class I gene was not inducible. DH1 and DH2 remained absent even after fusion with class-I-positive cells. However, transfected Dk genes were expressed in IC9, and both DH1 and DH2, which were probably derived from the transfected gene, became detectable. In tumor cells expressing class I genes only after treatment with IFN-gamma (e.g. the lung carcinoma CMT64.5) or after in vitro differentiation (F9 embryonal carcinoma cells), DH1 and DH2 were already present before induction of class I expression. However, the intensity of the band indicative of DH2 was reduced in undifferentiated and differentiated F9 cells and in untreated CMT cells.
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PMID:Altered regulation of MHC class I genes in different tumor cell lines is reflected by distinct sets of DNase I hypersensitive sites. 250 17

In Y1 mouse adrenocortical tumor cells, expression of steroid 11 beta-hydroxylase (11 beta-OHase) is stimulated by cAMP following a delay of 4-6 h. Our results demonstrate that a cAMP-responsive element (CRE) within the 11 beta-OHase promoter region is a major determinant of this induction. The 5'-flanking sequences from the mouse 11 beta-OHase gene were placed in front of a human growth hormone reporter gene and transfected into Y1 cells. Treatment of transfected cells with 8-bromo-cAMP increased expression directed by the 11 beta-OHase 5'-flanking region by 3.8-fold. In 5'-deletion analyses, 123 base pairs of 5'-flanking sequences were sufficient for cAMP induction, whereas cAMP treatment did not affect expression of a plasmid with only 40 base pairs of 5'-flanking sequence. Within these 123 base pairs, a region from -56 to -49 matched 7 of 8 bases comprising the consensus sequence for the CRE. 11 beta-OHase 5'-flanking sequences from -65 to -42, including the CRE-like sequence, conferred cAMP inducibility to promoters from the thymidine kinase and chorionic gonadotropin alpha-subunit genes. DNase I footprinting and Southwestern blotting analyses demonstrated that the protein which interacted with the CRE in the 11 beta-OHase promoter region was similar to the CRE-binding protein associated with other cAMP-regulated genes. Together, these results suggest that an interaction between the 11 beta-OHase CRE and CRE-binding protein mediates cAMP induction of the 11 beta-OHase gene.
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PMID:A cAMP-responsive element regulates expression of the mouse steroid 11 beta-hydroxylase gene. 254 79

In Agrobacterium tumefaciens, a cis-active 24-base-pair sequence adjacent to the right border of the T-DNA, called overdrive, stimulates tumor formation by increasing the level of T-DNA processing. Recent results from our laboratory have suggested that the virC operon which enhances T-DNA processing probably does so because the VirC1 protein interacts with overdrive (N. Toro, A. Datta, M. Yanofsky, and E. W. Nester, Proc. Natl. Acad. Sci. USA 85:8558-8562, 1988). We report here the purification of the VirC1 protein from cells of Escherichia coli harboring a plasmid containing the coding sequences of the virC locus of the octopine Ti plasmid. By gel mobility shift and DNase I footprinting assays, we showed that this purified virC1 gene product binds to overdrive but not to the right border of T-DNA.
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PMID:The Agrobacterium tumefaciens virC1 gene product binds to overdrive, a T-DNA transfer enhancer. 259 51


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