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Biomedical subjects

C B Bruni

Publications and source records attributed to C B Bruni.

At least 55 records · Page 3Linked to original sources
PubMed1995

Loss of heterozygosity of imprinted genes in SV40 t/T antigen-induced hepatocellular carcinomas.

Expression of the chromosomally linked Insulin-like Growth Factor II (IGF-II) and H19 genes is regulated by parental imprinting during development, since the maternally inherited IGF-II and the paternally inherited H19 alleles are inactive in fetal tissues. Here we show that expression of IGF-II and H19 genes is activated in transgenic mice during SV40 Tag-induced hepatocarcinogenesis and that imprinting of both genes is conserved in the liver tumors. Allelic imbalances of IGF-II and H19 genes and other chromosome 7 markers were detected in one third (13/39) of the hepatocellular carcinomas analysed. A strong bias on the allele retained in the neoplasms was observed, since underrepresentation or complete loss of maternal chromosome 7 was recognised in 12/13 cases. High levels of IGF-II mRNA were expressed by all carcinomas with relative excess of paternal chromosome 7 alleles and suppressed H19 expression was found in the neoplasms lacking the maternal alleles. Overall the results indicate that expression of imprinted genes is involved in progression of experimental liver tumors and suggest that the murine chromosome 7, whose loss may possibly cause the inactivation of a growth-inhibitory gene, is preferentially retained as paternal copy in the liver tumors because of parental imprinting of IGF-II gene.

Alleles↗
PubMed1995

Expression of galectin-1 in normal human thyroid gland and in differentiated and poorly differentiated thyroid tumors.

We previously reported that galectin-1 gene expression increases up to 100-fold in oncogene-transformed rat thyroid cells compared with their normal counterparts and that the relative mRNA levels correlate with the degree of malignancy. In the present study we investigated whether galectin-1 is differentially expressed in human thyroid neoplasms, which range from well-differentiated tumors to undifferentiated anaplastic carcinomas. We analyzed 74 human thyroid specimens of neoplastic, hyperproliferative and normal tissues and several tumor cell lines. Galectin-1 mRNA and protein levels were higher in 6 thyroid carcinoma-derived cell lines than in normal thyroid primary cultures and adenoma cells. Galectin-1 mRNA levels increased in 28/40 papillary carcinomas and in 6/7 anaplastic carcinomas compared with normal or hyperplastic thyroid. Conversely, galectin-1 expression was unaffected in follicular carcinomas and benign adenomas. Immunohistochemical analysis of normal thyroid and papillary carcinoma sections revealed a higher content of galectin-1 protein in neoplastic follicular cells than in normal cells.

Base Sequence↗
PubMed1995

Co-operative regulation of endocytosis by three Rab5 isoforms.

Rab proteins are small GTPases involved in the regulation of membrane traffic. Rab5a has been shown to regulate transport in the early endocytic pathway. Here we report the isolation of cDNA clones encoding two highly related isoforms, Rab5b and Rab5c. The two proteins share with Rab5a all the structural features required for regulation of endocytosis. Rab5b and Rab5c colocalize with the both transferrin receptor and Rab5a, stimulate the homotypic fusion between early endosomes in vitro and increase the rate of endocytosis when overexpressed in vivo. These data demonstrate that three Rab5 isoforms cooperate in the regulation of endocytosis in eukaryotic cells.

Amino Acid Sequence↗
PubMed1994

Ribonuclease E provides substrates for ribonuclease P-dependent processing of a polycistronic mRNA.

The polycistronic mRNA of the histidine operon is subject to a processing event that generates a rather stable transcript encompassing the five distal cistrons. The molecular mechanisms by which such a transcript is produced were investigated in Escherichia coli strains carrying mutations in several genes for exo- and endonucleases. The experimental approach made use of S1 nuclease protection assays on in vivo synthesized transcripts, site-directed mutagenesis and construction of chimeric plasmids, dissection of the processing reaction by RNA mobility retardation experiments, and in vitro RNA degradation assays with cellular extracts. We have found that processing requires (1) a functional endonuclease E; (2) target site(s) for this activity in the RNA region upstream of the 5' end of the processed transcript that can be substituted by another well-characterized rne-dependent cleavage site; (3) efficient translation initiation of the first cistron immediately downstream of the 5' end; and (4) a functional endonuclease P that seems to act on the processing products generated by ribonuclease E. This is the first evidence that ribonuclease P, an essential ribozyme required for the biosynthesis of tRNA, may also be involved in the segmental stabilization of a mRNA.

Amino Acid Sequence↗
PubMed1994

Parental imprinting of rat insulin-like growth factor II gene promoters is coordinately regulated.

The insulin-like growth factor II (IGF-II) gene is parentally imprinted in the mouse and human species. By following the inheritance of natural polymorphisms of IGF-II mRNA, we demonstrated that the tissue-specific parental imprinting of the IGF-II gene is conserved in the rat. The expression of the paternal IGF-II allele exceeded by more than 3 orders of magnitude that of the maternal allele in livers of 3-day-old Wistar x Fisher interstrain rat crosses. In contrast, the two alleles were both expressed in the rat central nervous system, which is also the only district of the organism where this gene is active in adult rodents. We also analyzed the allelic usage of the three IGF-II promoters, which generate alternatively spliced transcripts, and showed that parental imprinting of all transcription starts sites is coordinately regulated since P1, P2, and P3 are all repressed on the maternal allele in neonatal rat liver, and all of them are activated on both alleles in the choroid plexus of the central nervous system. RNase protection assays demonstrated that the activity ratio of the three IGF-II promoters can be different in tissues that show the same imprinting mode.

Alleles↗
PubMed1994

Alternative patterns of his operon transcription and mRNA processing generated by metabolic perturbation.

Previous studies have shown that the expression of the his operon of Salmonella typhimurium is regulated at the level of transcription initiation, transcription elongation and RNA processing. We have analyzed his RNA in both prototrophic strains or strains harboring regulatory and auxotrophic mutations grown under a variety of metabolic conditions that lead to differential expression of the operon. Under some of these conditions, there is an increase in the amount of prematurely released his-specific RNA, resulting in modulation of the relative amount of full-length transcripts. Under the same metabolic conditions, there is also a modulation of RNA processing events that generate a very stable RNA species comprising the five distal cistrons. These effects appear to be due to perturbation of the translation process caused by alterations in the intracellular pool of initiator transfer RNA.

Gene Expression Regulation, Bacterial↗
PubMed1994

Rab5a is a common component of the apical and basolateral endocytic machinery in polarized epithelial cells.

In nonpolarized cells, the small GTPase Rab5a is localized to the plasma membrane, clathrin-coated vesicles, and early endosomes. Rab5a is required for early endosome fusion in vitro and regulates transport between the plasma membrane and early endosomes, in vivo. In polarized epithelial cells endocytosis occurs from separate apical and basolateral plasma membrane domains. Internalized molecules are initially delivered to distinct apical or basolateral early endosomes. In vitro, apical early endosomes can readily fuse with one another but not with the basolateral endosomes and vice versa, thereby indicating that the apical and basolateral early endocytic pathways are controlled by distinct machineries. Here, we have investigated the localization and function of Rab5a in polarized epithelial cells. Confocal immunofluorescence microscopy on mouse kidney sections revealed association of the protein with the apical and basolateral plasma membrane domains and underlying structures. In polarized Madin-Darby canine kidney I cells, endogenous and overexpressed Rab5a have the same distribution. Moreover, overexpression of the protein causes a 2-fold increase in fluid-phase uptake from both domains of the cell, thus showing that Rab5a functions in apical and basolateral endocytosis. Our data indicate that the apical and basolateral endocytic machineries of epithelial cells share common regulatory components and that Rab5a per se is not sufficient to target endocytic vesicles to apical or basolateral early endosomes.

Amino Acid Sequence↗
PubMed1994

Expression of the soluble lectin L-14 gene is induced by TSH in thyroid cells and suppressed by retinoic acid in transformed neural cells.

Expression of the soluble lectin L-14 is low in normal and very high in transformed thyroid cells. We show that L-14 gene expression is transiently induced upon thyrotropin stimulation of normal quiescent FRTL-5 rat thyroid cells. Permanent activation of L-14 gene expression is obtained in the same cells infected with a wild-type and a temperature sensitive mutant of Kirsten murine sarcoma virus, both at the permissive and non permissive temperature for transformation. We also find that L-14 mRNA is undetectable in rat brain but is abundant in rat oligodendrocytes precursors transformed by polyoma middle T oncogene. Retinoic acid treatment of these transformed cells leads to acquisition of a differentiated phenotype accompanied by a 30-fold decrease of L-14 mRNA levels. Removal of retinoic acid restores both the transformed undifferentiated phenotype and high L-14 expression. Taken together these results indicate that growth stimulation and induction of cell differentiation are accompanied by strong modulation of L-14 gene expression.

Animals↗
PubMed1994

Control of mRNA processing and decay in prokaryotes.

Post-transcriptional mechanisms operate in regulation of gene expression in bacteria, the amount of a given gene product being also dependent on the inactivation rate of its own message. Moreover, segmental differences in mRNA stability of polycistronic transcripts may be responsible for differential expression of genes clustered in operons. Given the absence of 5' to 3' exoribonucleolytic activities in prokaryotes, both endoribonucleases and 3' to 5' exoribonucleases are involved in chemical decay of mRNA. As the 3' to 5' exoribonucleolytic activities are readily blocked by stem-loop structures which are usual at the 3' ends of bacterial messages, the rate of decay is primarily determined by the rate of the first endonucleolytic cleavage within the transcripts, after which the resulting mRNA intermediates are degraded by the 3' to 5' exoribonucleases. Consequently, the stability of a given transcript is determined by the accessibility of suitable target sites to endonucleolytic activities. A considerable number of bacterial messages decay with a net 5' to 3' directionality. Two different alternative models have been proposed to explain such a finding, the first invoking the presence of functional coupling between degradation and the movement of the ribosomes along the transcripts, the second one implying the existence of a 5' to 3' processive '5' binding nuclease'. The different systems by which these two current models of mRNA decay have been tested will be presented with particular emphasis on polycistronic transcripts.

Bacteria↗
PubMed1994

Mono- and bi-allelic expression of insulin-like growth factor II gene in human muscle tumors.

Insulin-like growth factor II (IGF-II) is a mitogen for many cell types and an important modulator of muscle growth and differentiation. IGF-II gene is prevalently expressed during prenatal development and its gene activity is regulated by genomic imprinting, in that the allele inherited from the father is active and the allele inherited from the mother is inactive in most normal tissues. IGF-II expression is activated in several types of human neoplasms and an alteration of IGF-II imprinting has been described in Beckwith-Wiedemann syndrome and Wilms' tumor. Here we show that monoallelic expression of IGF-II gene is conserved in normal adult muscle tissue whereas two or more copies of active IGF-II alleles, arising by either relaxation of imprinting or duplication of the active allele, are found in 9 out of 11 (82%) rhabdomyosarcomas retaining heterozygosity at 11p15, regardless of the histological subtype. Since IGF-II has been indicated as an autocrine growth factor for rhabdomyosarcoma cells, these findings strongly suggest that acquisition of a double dosage of active IGF-II gene is an important step for the initiation or progression of rhabdomyosarcoma tumorigenesis. Among different types of muscle tumors, relaxation of imprinting seems to arise prevalently in rhabdomyosarcomas, since we have detected only one case of partial reactivation of the maternal IGF-II allele out of 7 leiomyosarcomas tested.

Adult↗
PubMed1994

Activation of the galectin-1 (L-14-I) gene from nonexpressing differentiated cells by fusion with undifferentiated and tumorigenic cells.

Expression of the galectin-1 (L-14-I) gene, elevated in most differentiated and transformed cell lines, has been studied in cell hybrid systems. Fusion of L-14-I nonproducing rat liver differentiated FAO cells with dedifferentiated rat liver BRL3A cells leads to extinction of liver-specific gene expression while L-14-I mRNA levels remain high. Interspecific hybrids produced by fusion of tumorigenic human osteosarcoma 143TK- with FAO cells show loss of both differentiated functions and tumorigenic phenotype and activation of the FAO L-14-I alleles. Increased expression of rat L-14-I alleles was also observed in human osteosarcoma x rat thyroid cells transient heterokaryons. The data presented here show that expression of the L-14-I gene is subject to dominant positive control and that it correlates with loss of differentiation-specific functions, but it is independent from tumorigenicity. L-14-I activation in FAO cells is achieved by treatment with 5-azacytidine. This result suggests that DNA demethylation is responsible or a prerequisite for L-14-I activation in hybrids.

Alleles↗
PubMed1994

Expression of insulin-like growth factor (IGF)-II and IGF-I receptor during proliferation and differentiation of CaCo-2 human colon carcinoma cells.

We have studied the expression of insulin-like growth factor type II (IGF-II) and its autocrine role during the proliferation and differentiation of the CaCo-2 colon carcinoma cell line. IGF-II RNA levels were high in proliferating cells and decreased by more than 10-fold when cells ceased to proliferate and differentiated. Immunoreactive IGF-II protein was high in the conditioned media of proliferating cells and decreased 20-fold in the media of differentiated cells. Reduced IGF-II expression was associated with a decrease in IGF-I receptor number that was high in proliferating cells (approximately 80,000 binding sites/cell) and reduced by 4-fold in differentiated cells. Exogenously added IGF-II was able to stimulate proliferation of serum-deprived cells in a dose-dependent fashion. IGF-II acted through the IGF-I receptor, since both basal and IGF-II-stimulated cell proliferation was inhibited by the monoclonal antibody alpha-IR3, which blocks the binding sites of the IGF-I receptor. The inhibition of CaCo-2 basal cell growth by the alpha-IR3 antibody suggests that IGF-II may act as an autocrine growth factor for these cells.

Antibodies, Monoclonal↗
PubMed1993

Mapping on human and mouse chromosomes of the gene for the beta-galactoside-binding protein, an autocrine-negative growth factor.

The structural gene for beta-galactoside-binding protein (Lgals-1), a cell growth regulatory molecule and cystostatic factor, is assigned to the E-region of mouse chromosome 15 and to the region q12-q13.1 of human chromosome 22. The evolutionary conservation of these two regions has been previously suggested from comparative mapping of several loci. These include the murine SIS oncogene (Pdgfb) and its human homolog, the platelet-derived growth factor-beta polypeptide, PDGFB. The findings presented here extend the genetic homology of the two regions.

Animals↗
PubMed1993

Characterization of the rho genes of Neisseria gonorrhoeae and Salmonella typhimurium.

We have cloned and sequenced the genomic regions encompassing the rho genes of Neisseria gonorrhoeae and Salmonella typhimurium. Rho factor of S. typhimurium has only three amino acid differences with respect to the Escherichia coli homolog. Northern (RNA) blots and primer extension experiments were used to characterize the N. gonorrhoeae rho transcript and to identify the transcription initiation and termination elements of this cistron. The function of the Rho factor of N. gonorrhoeae was investigated by complementation assays of rho mutants of E. coli and S. typhimurium and by in vivo transcription assays in polar mutants of S. typhimurium.

Amino Acid Sequence↗
PubMed1993

Extinction of insulin-like growth factor II gene expression in intratypic hybrids of rat liver cells.

The expression of the developmentally regulated insulin-like growth factor II (IGF-II) gene has been studied in somatic cell hybrids derived from rat liver cells. BRL3A cells, dedifferentiated variants of rat hepatocytes, producing high levels of IGF-II, were fused to BRL30E or FAO cells of the same embryonic lineage but not expressing detectable levels of IGF-II mRNA. We report here that the IGF-II gene is subject to extinction, since its specific RNA levels are decreased both in heterokaryons and stable cell hybrids. Transcriptional analysis in isolated nuclei from parental and hybrid cells showed that the IGF-II gene is transcribed at a similar rate in all cell types. Likewise, the stability of IGF-II cytoplasmic mRNA was equivalent in the high-expressing parental cells and in the hybrids. In contrast, the distribution of IGF-II mRNA between the nuclear and the cytoplasmic compartments differed markedly in parental and hybrid cell lines. The data presented show that the expression of the IGF-II gene is subject to a dominant negative control and suggest that the phenomenon involves mechanisms that operate at the posttranscriptional level.

Animals↗
PubMed1992

Regulation of insulin-like-growth-factor-II gene expression in rat liver cells.

The rat insulin-like-growth-factor-(IGF)-II gene is expressed at high levels during embryonic and fetal life and at low levels in adult animals. To study the regulation of IGF-II gene expression, we analyzed the synthesis and localization of the IGF-II transcripts in cultured rat liver cells either expressing (BRL3A cells) or not expressing (BRL30E and FAO cells) the IGF-II mRNA. The IGF-II gene is transcribed at a similar rate in expressing and non-expressing cells, whereas its nuclear and cytoplasmic RNA levels are diversely distributed in the cells. IGF-II RNA is more abundant in the cytoplasmic than in the nuclear RNA fraction of BRL3A cells and is present in the nucleus but not in the cytoplasm of the FAO cells. However, both precursor and mature IGF-II nuclear RNA levels are reduced in FAO cells. Our data indicate that the IGF-II gene expression is regulated by mechanisms affecting the subcellular distribution and the abundance of the transcripts.

Animals↗
PubMed1992

Further characterization of the histidine gene cluster of Streptomyces coelicolor A3(2): nucleotide sequence and transcriptional analysis of hisD.

We have further characterized the genomic region of Streptomyces coelicolor A3(2) that contains genes involved in the biosynthesis of histidine. A 2,357-base pair fragment contained in plasmid pSCH3328 that complemented hisD mutations has been sequenced. Computer analysis revealed an open reading frame that encodes a protein with significant homology to the Escherichia coli, Salmonella typhimurium and Mycobacterium smegmatis hisD product, Saccharomyces cerevisiae HIS4C, and Neurospora crassa his3 gene products. Two other contiguous open reading frames oriented divergently with respect to hisD did not show significant similarity with any of the his genes or to other sequences included in the gene bank. S1 nuclease mapping and primer extension experiments indicate that the transcription initiation site of the his-specific mRNA coincides with the GUG translation initiation codon of the hisD cistron.

Base Sequence↗