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F Cimino

Publications and source records attributed to F Cimino.

At least 19 recordsLinked to original sources
PubMed1992

Sp1 DNA binding efficiency is highly reduced in nuclear extracts from aged rat tissues.

To explore the role of transcriptional factors in the genesis of the senescent phenotype, nuclear extracts from 4- and 30-month-old rat brains were analyzed for the presence of DNA-binding proteins able to interact with double-stranded oligonucleotides containing recognition sites for sequence-specific DNA-binding factors. Gel shift assays revealed that the DNA-binding efficiency of Sp1 is significantly reduced in aged animals compared to young ones, whereas CTF/NF1 and AP1 from young and old rat nuclear extracts bind their DNA targets with the same efficiency. The quantitative analysis of Sp1 by immunoblotting indicated that equivalent quantities and degrees of heterogeneity of Sp1 protein are present in both nuclear extracts, suggesting that the observed difference is not due to a different expression of this transcriptional factor. DNase I footprinting of the heavy chain ferritin gene promoter, which contains a Sp1 binding site, demonstrated that the nuclear extract from 30-month-old rat brain does not protect the region involved in the regulation of the H ferritin gene by Sp1. This results in a reduction of about 50% of the expression of the H ferritin mRNA in aged rat brains. Furthermore, the Sp1 binding sites present in the SV40 early promoter are not protected in a DNase I footprinting assay where a nuclear extract from 30-month-old rat brain was used as a source of DNA binding proteins. Liver nuclear extracts prepared from young and aged rats demonstrated that a decrease of Sp1 binding efficiency is similarly present in this tissue.

Aging↗
PubMed1992

Promoter for the human ferritin heavy chain-encoding gene (FERH): structural and functional characterization.

We conducted a functional analysis of the promoter for the human ferritin heavy chain-encoding gene (pFERH) in HepG2 and HeLa cells. The activity of pFERH is equivalent in both cell types, despite their different ferritin (Fer) isotypes. Transfections of a series of 5'-deletion mutants indicate that pFERH activity is essentially dependent on two motifs. One of them, accounting for about 50% of the total transcriptional activity, is recognized by the RNA polymerase II transcription factor, Sp1, and the other by a low-affinity factor present in both the cell types analyzed.

Base Sequence↗
PubMed1991

A rat brain mRNA encoding a transcriptional activator homologous to the DNA binding domain of retroviral integrases.

We have isolated a rat cDNA, named FE65, hybridizing to an mRNA of about 2,300 nucleotides present in rat brain, undetectable in rat liver and very poorly represented in other tissues. An mRNA of the same size is present in human neuroblastoma cells and is absent from other human cell lines. The FE65 cDNA contains an open reading frame (ORF) coding for a polypeptide of 499 amino acids in which 143 residues can be aligned with the DNA binding domain of the integrases encoded by mammalian immunodeficiency viruses. The remaining part of the FE65 ORF is not homologous with the correspondent regions of the integrases; the first 206 residues of the FE65 ORF show numerous negative charges and a short sequence not dispensable for the function of the transactivating acidic domain of the jun family transcriptional factors. A plasmid which expresses FE65 amino acids 1-232 fused to the yeast GAL4 DNA binding domain was co-transfected with a plasmid containing five GAL4 binding sites upstream of a minimal Adenovirus promoter controlling the expression of the CAT gene. This experiment showed that the fused protein GAL4-FE65 is able to obtain a 30-40 fold increase of the CAT gene expression compared to the expression observed in the presence of the GAL4 DNA binding domain alone. Two types of FE65 mRNA are present in rat brain, differing only for six nucleotides. We demonstrate that this is the consequence of a neuron-specific alternative splicing of a six-nucleotide miniexon, which is also present in the human genome, in an intron/exon context very similar to that of the rat FE65 gene.

Amino Acid Sequence↗
PubMed1990

Isolation of cDNA fragments hybridizing to rat brain-specific mRNAs.

A cDNA minilibrary in pUC18 has been generated from 3-month-old rat brains. Two hundred clones were randomly selected and sequenced. Comparison with a nucleic acid and protein data bank revealed a number of cDNA fragments not homologous to any published sequence. Northern blot analyses of some of these clones yielded 5 brain-specific cDNA fragments. The full-length cDNA for one of these clones has been isolated and completely sequenced. It corresponds to an mRNA of about 1,500 nucleotides, which is present in brain and, to a lesser extent, in heart and skeletal muscle. Its expression is developmentally regulated in the rat brain from 14-day embryos to 3-month-old adults. A very recent comparison with the EMBL nucleotide sequence data bank showed that this mRNA codes for a brain-specific snRNP-associated protein.

Aging↗
PubMed1989

Expression of genes of ferritin subunits in human hepatoma cell lines.

The expression of ferritin genes has been studied in two well differentiated hepatoma cell lines, HepG2 and Hep3B, and in an undifferentiated cell line, HepPLC/PRF/5. The steady-state level of the H and L ferritin subunit mRNAs is different in the three cell lines, being most abundant in the HepG2 and Hep3B cells. The efficiency of promotion of transcription of the H and L gene promoters is not correlated with the levels of the two transcripts; moreover the half-life of the H and L mRNAs varies among the hepatoma cells. The different level of ferritin mRNAs in differentiated and undifferentiated cells appears to be mainly due to a different stability of the transcripts.

Blotting, Northern↗
PubMed1989

Comparative effects of roxithromycin and erythromycin on cellular immune functions in vitro. 1. Uptake of 3H-macrolides by human macrophages.

The degree of penetration of roxithromycin in human macrophages was evaluated and compared with that of erythromycin. The results indicated that both macrolides at concentrations of 2, 10 and 20 micrograms/ml were concentrated by macrophages, even if roxithromycin was consistently and significantly more accumulated than erythromycin. Comparison of the degree of penetration of the two drugs into dead, resident and stimulated phagocytes indicated that the process of transfer through the macrophage wall was of an active nature, related to the metabolic state of the cells. Analysis of the binding of the two macrolides to intracellular proteins showed that more binding sites were available for erythromycin than for roxithromycin.

Analysis of Variance↗
PubMed1988

Oligonucleotide-directed mutagenesis: a sequence-based screening.

A procedure to screen for mutant clones obtained by oligonucleotide-directed mutagenesis has been developed. It is based on the preparation of phage containing supernatants from a number (100 or more) of randomly chosen mutagenized M13 plaques. Aliquots from these supernatants are mixed to obtain pools, each containing 10 phages. Heterogeneous single-stranded DNA is prepared from these pools and used as template in a "single letter" sequence according to the dideoxy chain terminator method. Thus, the pool(s) containing the mutated sequence and the mutated sequence itself is identified by sequencing the single-stranded DNAs of the 10 phages present in the selected pool.

Animals↗
PubMed1987

The transcriptional efficiency of clustered tRNA genes is affected by their position within the cluster.

The transcription of a mouse genomic segment containing four tRNA genes, coding for a tRNA(Ala), a tRNA(Ile), a tRNA(Pro) and a tRNA(Lys), has been studied in a HeLa cell extract, demonstrating that differences among their transcriptional efficiencies are evident using as templates either the natural cluster or an equimolecular mixture of the four isolated genes. Nevertheless, the structure of the cluster influences the transcriptional efficiency of the clustered genes. In fact, a cis-acting inhibitory sequence has been located at about 400 bp downstream of the tRNA(Pro) coding sequence. Moreover rearrangements of the reciprocal position of the various tRNA genes within the cluster results in significant changes in the transcriptional rates of the individual transcriptional units.

Animals↗
PubMed1986

Structure and in vitro transcription of tRNA gene clusters containing the primers of MuLV reverse transcriptase.

Three genes coding for mouse tRNAPro have been isolated from a genomic library and characterized both structurally and functionally. Two of these (tPro52 and tPro53) code for the tRNA primer of reverse transcriptase of MuLV. The third one (tPro51) shows several differences (mutations and deletions) that probably prevent the folding of the matured transcript into the cloverleaf structure, and is therefore a pseudogene. This pseudogene gives rise to a RNA transcription product in vitro. tPro52 is clustered with a tRNALys gene and with a tRNAAla gene, which is strongly homologous to the rat identifier repeated sequence. tPro53 is clustered with a tRNAAsp and a tRNAGly gene. Other tRNA-hybridizing sequences are present in the lambda clones that contain tPro51 and tPro53.

Anticodon↗
PubMed1985

Pseudouridine excretion and transfer RNA primers for reverse transcriptase in tumors of retroviral origin.

To evaluate the relationship between pseudouridine increase in biological fluids and retroviral cell transformation, we have studied the effect of retrovirus infection and/or transformation on the rate of pseudouridine excretion by chick embryo fibroblasts. The results show that: pseudouridine excretion by chick embryo fibroblasts transformed by Rous sarcoma virus is several times higher than that of normal cells; this increased excretion precedes by many hours the appearance of the morphological signs of transformation and it is always present when neosynthesized infectious viral particles are released into the culture medium; and pseudouridine excretion was also increased in cells infected by a mutant of Rous sarcoma virus (RAV-1) which, lacking the src gene, does not transform the cells but replicates normally. To investigate if pseudouridine overproduction is related to an altered turnover rate of specific transfer RNA (tRNA) species which functions as primer of retrovirus reverse transcriptase, the concentration of non-acylated proline-accepting tRNA and non-acylated tryptophan-accepting tRNA, primers of reverse transcriptase of murine leukemia virus and of Rous sarcoma virus, respectively, has been measured, the former in normal and transformed AKR thymus and the latter in normal fibroblasts and in fibroblasts infected by Rous sarcoma virus or by its nontransforming mutant. The results show that in both systems a significant increase of the primer tRNA species occurs in the infected or transformed cells.

Animals↗