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B Calabretta

Publications and source records attributed to B Calabretta.

179 records · Page 10Linked to original sources
PubMed1986

Coding sequence and growth regulation of the human vimentin gene.

We have established the complete coding sequence of the human vimentin gene. It had 91% homology to the coding sequence of the Syrian hamster vimentin gene (Quax et al., Cell 35:215-223, 1983) and partial homology to several other sequences coding for intermediate filament proteins. The most striking difference between the Syrian hamster and human vimentin genes was in the 3' untranslated region, which was considerably longer in the Syrian hamster. Using RNA blots and a human vimentin cDNA clone from an Okayama-Berg library, we have established that expression of the vimentin gene was growth regulated. The steady-state levels of cytoplasmic vimentin mRNA in 3T3 cells were increased by serum and platelet-derived growth factor, but not by epidermal growth factor, insulin, or platelet-poor plasma. The increase in expression of the vimentin gene that occurred when G0-phase cells were stimulated to proliferate was detected in six different cell types from four different species. The expression of the vimentin gene was also increased when HL60 cells were induced to differentiate by phorbol esters; it decreased when differentiation was induced by retinoic acid.

Amino Acid Sequence↗
PubMed1986

Growth-dependent expression of human Mr 53,000 tumor antigen messenger RNA in normal and neoplastic cells.

We have investigated the expression of Mr 53,000 protein (p53) in total RNA isolated from human peripheral blood mononuclear cells stimulated by phytohemagglutinin, in serum-stimulated human diploid fibroblasts, and in normal and tumor cells of human epithelial colon tissue. We have found that the expression of p53 messenger RNA is growth regulated in human cells following kinetics similar to that previously shown in mouse 3T3 cells, and is increased in the large majority of colon adenocarcinomas in comparison to adjacent normal mucosa and adenoma. This increased expression of p53 is accompanied by a nearly proportional increase in the expression of histone H3. As the expression of histone H3 is restricted to the S phase of the cell cycle and therefore measures the growth fraction of a given population, we suggest that the increased expression of p53 observed in the large majority of colon tumors simply reflects the increased number of cycling cells frequently found in a neoplastic tissue. At variance with these findings a true overexpression of p53 was detected in one SV40-transformed human fibroblasts cell line.

Adenocarcinoma↗
PubMed1986

HLA-DR-associated invariant chain is highly expressed in chronic lymphocytic leukemia.

Total RNA extracted from peripheral blood lymphocytes of a patient with B-cell chronic lymphocytic leukemia (CLL) and the poly (A+) RNA was purified. A cDNA library was constructed and approximately 4,000 clones were screened in order to identify genes preferentially expressed in CLL. A relatively low repetition frequency characterizes the majority of the abundant mRNA species present in CLL lymphocytes. One clone, corresponding to the mRNA encoding the HLA-DR-associated invariant chain, was selected and its expression was examined in different leukemic cell populations and in normal tissues. DNA-RNA hybridization studies showed that the invariant chain mRNA (In-mRNA) is detectable in RNA preparations from human blood cells and their precursors, whereas no In-mRNA is found in several other tissues examined. Among various normal and leukemic leukocyte populations, the highest levels of In-mRNA are found in CLL. Therefore, a role of In-chain mRNA as a marker of CLL is proposed. Our data support a relationship between high levels of invariant chain mRNA and the out of cycle condition of CLL peripheral blood lymphocytes.

Clone Cells↗
PubMed1986

Expression of growth-regulated genes in human acute leukemias.

We have investigated the expression of six growth-regulated genes (c-myc, c-myb, p53, 4F1, 2F1, and ornithine decarboxylase) and the S-phase-specific histone H3 gene in acute myeloid and lymphoid leukemic cells. We have purposely chosen three growth-regulated protooncogenes that share similar biological features and three gene sequences that have in common the cell cycle dependence of their expression in cells of different tissue and in different species. The level of expression was determined by measuring the amounts of specific RNA by Northern blot analysis. Levels of expression of the six growth-regulated genes were compared to the level of expression of the S-phase-specific H3 gene and among themselves. This method distinguishes the increased expression of a growth-regulated gene due to a true altered activation from over-expression which simply reflects an increase in the fraction of cycling cells. We have found that six of 14 patients with acute leukemias have markedly high ratios of c-myc/H3, c-myc/p53, and c-myc/c-myb expression. Two patients with altered c-myc expression have also a high ratio p53/H3. Within the group of cell cycle-dependent genes the ratios of expression seem in the overall much more regular with the clear exception of a patient with acute myelogenous leukemia in which the ratios 4F1/H3 and 2F1/H3 are significantly increased. A possible interpretation of these findings is that the fraction of noncycling leukemic cells that often constitute the majority of the entire leukemic population is in some cases in a true resting state, whereas in other cases heterogeneous degrees of growth arrest might occur. The altered expression of c-myc seems the feature most commonly associated with this putative growth arrest of leukemic cells suggesting that this gene may contribute to the impairment of proliferative control that is associated with the leukemic phenotype.

Acute Disease↗
PubMed1985

Effect of interleukin-2 on the expression of cell cycle genes in human T lymphocytes.

We have studied the expression of seven cell cycle-dependent genes in phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells, in macrophage-depleted cultures and in macrophage-depleted cultures plus Interleukin-2 (IL-2). The expression of all seven genes is increased in PHA stimulated peripheral cells. Only two (2F1 and the IL-2 receptor) are increased in PHA-stimulated macrophage depleted cultures. Addition of IL-2 to these cultures increased the RNA levels of four genes (KC-1, c-myc, beta-actin and IL-2R), but has no effect on three others (4F1, 2F1, and JE-3). The results indicate that the expression of these cell cycle genes is regulated by different components of the mitogenic stimulus.

Autoradiography↗
PubMed1985

Recent amplification of an alpha satellite DNA in humans.

A repeat sequence 682 base pairs (bp) long produced by cleavage of human DNA with Xba I restriction enzyme is composed of four tandemly arranged subunits with lengths of 171, 170, 171, and 170 bp each. The sequence organization of the 682 bp Xba I repeat bears a striking resemblance to other complex satellite DNAs of primates, including the Eco RI human alpha satellite family which also occurs as a 170 bp repeat. The Eco RI tetramer and the 682 bp Xba I repeat show a sequence divergence of 21%. The 682 bp Xba I repeat sequence is restricted to humans and is only distantly related to the previously reported 340 bp Xba human repeated DNA sequence. These finding are consistent with the concept of occasional amplifications of members or groups of members of alpha satellite DNA during human evolution. Amplifications apparently occurred after humans, apes and gibbons diverged from Old World monkeys (Eco RI satellite), after humans and apes diverged from gibbons (340 bp Xba I satellite) and after humans diverged from the great apes (682 bp Xba I satellite).

Animals↗
PubMed1985

Cell-cycle-specific genes differentially expressed in human leukemias.

Three cDNA clones isolated from Syrian hamster cells (p4F1, p2F1, and p2A9) contain sequences that are preferentially expressed in the G1 phase of the cell cycle. The expression of these sequences was investigated in human peripheral blood cells from normal individuals and from patients with leukemia. The expression of p4F1 and p2F1 is clearly dependent on the cell cycle in peripheral blood mononuclear cells stimulated to proliferate with phytohemagglutinin; the p2A9 sequences cannot be clearly detected in human lymphocytes but are expressed in a cell-cycle-dependent manner in human diploid fibroblasts (WI-38). These genes also show different levels of expression in lymphoid and myeloid leukemias. The highest level of expression for p2A9 is found in patients with chronic myelogenous leukemia, and the lowest in patients with chronic lymphocytic leukemia. For p2F1 and p4F1, the highest levels of expression are found in chronic and acute myelogenous leukemia. At least two other cell-cycle genes are not expressed at detectable levels in human leukemias. These findings suggest that the activation of cell-division-cycle genes might contribute, like cellular oncogenes, to the phenotype of human malignancies and that, perhaps, new oncogenes could be found by identifying and isolating genes whose expression is dependent on the cell cycle.

Cell Cycle↗
PubMed1985

Expression of cell-cycle-dependent genes in phytohemagglutinin-stimulated human lymphocytes.

We have investigated the expression of certain cell-cycle-dependent genes in human peripheral blood mononuclear cells (PBMC) stimulated by phytohemagglutinin (PHA). The genes studied had been previously identified as cell-cycle dependent in other cell types from different species and were induced by different mitogens. One of these genes (2F1) and the gene for the interleukin 2 receptor were induced by PHA even in cultures partially depleted of accessory cells where the lymphocytes grew in size but failed to enter S phase. The other genes (c-myc, 4F1, JE-3, and KC-1) were induced only in complete cultures of PBMC stimulated by PHA. These results confirm the dissociation between growth in size and cell DNA replication that can occur during cell-cycle progression. Moreover, the time course of appearance of detectable levels of RNA for these genes suggests that they may be used as markers of cell-cycle progression in the transition of lymphocytes from G0 to S phase.

Cell Cycle↗
PubMed1985

Expression of c-myc and other cell cycle-dependent genes in human colon neoplasia.

We have investigated the expression of certain cell cycle-dependent genes in total RNA isolated from normal and neoplastic cells of human epithelial colon tissue. The genes studied had been previously identified as cell cycle dependent in rodent and human fibroblasts. Levels of expression of G1 genes were compared to the level of expression of the S-phase-specific gene H3 in normal and adjacent neoplastic epithelial cells of six different individuals. We have found that the increase in the expression of c-myc gene detected in colon tumor cells is accompanied by a parallel increase in the expression of two G1-specific genes (p2A9 and ornithine decarboxylase) and the S-phase-specific gene histone H3. An important conclusion that can be drawn from these findings is that the increased level of a cell cycle-specific RNA in a tumor may not indicate overexpression of that gene but simply reflect the increased fraction of cycling cells, unless the ratio of expression between G1 genes and G1-S-phase genes is altered.

Adenocarcinoma↗
PubMed1981

mRNA in human cells contains sequences complementary to the Alu family of repeated DNA.

Approximately one-half of the polysomal poly(A)+RNA from CCRF-CEM human lymphoblastoid cells associates at low R0t (10 M.sec) [where R0 is the initial concentration of RNA (M) and t is time (sec)] to form branched complexes detectable by electron microscopy. The complexes typically involve 2-16 molecules associated over double-stranded regions 120 +/- 30 base pairs long. Formation of such complexes suggests that poly(A)+RNA contains repeated-sequence elements that are highly represented in the mRNA population. Hybridization of polysomal poly(A)+RNA with a recombinant human DNA plasmid, p lambda H15C, which is shown to contain at least three regions complementary to two different members of the Alu family of DNA repeat sequences, showed a total of five regions where R loops are formed. The hybridized regions comprising these groups are 260 +/- 180, 240 +/- 170, 150 +/- 70, 180 +/- 60, and 180 +/- 80 base pairs long. The relative frequencies of R loops formed at these different sites indicate that sequences in this recombinant DNA are represented in the mRNA population at different frequencies. The hybridizing sequence of the RNA molecules is located near one terminus in 13% of the R loops and internally in 53% of the R loops. Surprisingly, 35% of the R loops apparently involve RNA molecules hybridized over their entire length of only 200 +/- 110 base pairs.

Cell Line↗
PubMed1981

Presence of a highly repetitive and widely dispersed DNA sequence in the human genome.

A genomic DNA library consisting of human DNA fragments about 18 kilobases long cloned in a bacteriophage lambda vector was found to contain a specific repeated DNA segment. The repeated sequence is present in greater than 95% of the genomic library, and selected clones contain at least two copies of the sequence. Our experiments indicate that this highly repetitive sequence (approximately 400,000 copies per haploid genome) is widely distributed in the human genome and is represented in the cytoplasmic polysomal mRNA. This sequence is homologous to the 300-base-pair Alu repeat family, the predominant repeat sequence in man.

Cloning, Molecular↗
PubMed1979

Kinetics of hybridization to human DNA of heterogeneous nuclear RNA isolated from normal human lymphoblasts and acute leukemia blast cells.

Heterogeneous nuclear RNA was extracted from normal PHA-stimulated human lymphocytes and acute myeloid leukemia blast cells. Experiments were performed to determine the hybridization kinetics of these RNA's to human DNA. The best least squares solutions indicate in the hybridization reaction of both normal and leukemic RNA two main components. For leukemic cell RNA the rate constants of both components were significantly different from that of normal cell RNA. In particular, the difference between the rate constants of the second lower component suggests that the slowly hybridizing sequences in leukemic cell RNA have a degree of repetition higher than of the corresponding sequences of normal cell RNA.

Base Sequence↗