PubMed Health⌕ Search

Biomedical subjects

C Saint-Ruf

Publications and source records attributed to C Saint-Ruf.

At least 19 recordsLinked to original sources
PubMed2000

Different initiation of pre-TCR and gammadeltaTCR signalling.

Lineage choice is of great interest in developmental biology. In the immune system, the alphabeta and gammadelta lineages of T lymphocytes diverge during the course of the beta-, gamma- and delta-chain rearrangement of T-cell receptor (TCR) genes that takes place within the same precursor cell and which results in the formation of the gammadeltaTCR or pre-TCR proteins. The pre-TCR consists of the TCRbeta chain covalently linked to the pre-TCRalpha protein, which is present in immature but not in mature T cells which instead express the TCRalpha chain. Animals deficient in pre-TCRalpha have few alphabeta lineage cells but an increased number of gammadelta T cells. These gammadelta T cells exhibit more extensive TCRbeta rearrangement than gammadelta T cells from wild-type mice. These observations are consistent with the idea that different signals emanating from the gammadeltaTCR and pre-TCR instruct lineage commitment. Here we show, by using confocal microscopy and biochemistry to analyse the initiation of signalling, that the pre-TCR but not the gammadeltaTCR colocalizes with the p56lck Src kinase into glycolipid-enriched membrane domains (rafts) apparently without any need for ligation. This results in the phosphorylation of CD3epsilon and Zap-70 signal transducing molecules. The results indicate clear differences between pre-TCR and gammadeltaTCR signalling.

Animals↗
PubMed1999

Pleiotropic changes controlled by the pre-T-cell receptor.

The construction of various gene-deficient mice has facilitated the understanding of the role of various receptors and signaling pathways that control the generation of alphabeta lineage cells. A predominant role is occupied by the pre-TCR, which not only generates large numbers of alphabeta lineage cells but also controls TCRbeta allelic exclusion as well as commitment to the gammadelta lineage versus the alphabeta lineage.

Alleles↗
PubMed1998

Genomic structure of the human pre-T cell receptor alpha chain and expression of two mRNA isoforms.

The pre-TCR, which is minimally composed of the TCRbeta chain, the pre-Talpha chain, and the CD3 complex, regulates early T cell development. The pre-Talpha chain is a 33-kDa type I transmembrane glycoprotein with an extracellular part similar to the constant domain of the immunoglobulin supergene family. We have sequenced (11 kb) the human pTalpha gene, which like the murine pTalpha gene consists of four exons: exon 1 encodes the 5' untranslated region, the leader peptide and the first three amino acids of the mature protein, exon 2 the extracellular immunoglobulin (Ig)-like domain, exon 3 a 15-amino acid peptide including a cysteine required for heterodimerization with TCRbeta, exon 4 the transmembrane region, the cytoplasmic tail and the 3' untranslated sequence. The human pTalpha gene is located on chromosome 6p21.3, close to the HLA-A locus. Reverse transcription-PCR studies with human thymus and leukemic cells showed that alternative splicing produces a shorter pTalpha isoform, which lacks the Ig-like domain but contains the transmembrane elements and the extracytoplasmic cystein and which could thus permit pairing with TCRbeta chain and association with CD3 molecules. The conserved splice sites suggest a yet ill-defined biological function of the short pTalpha protein.

Humans↗
PubMed1998

On the role of the pre-T cell receptor in alphabeta versus gammadelta T lineage commitment.

The role of the pre-T cell receptor (TCR) in lineage commitment to the gammadelta versus alphabeta lineage of T cells was addressed by analyzing TCRbeta chain rearrangements in gammadelta T cells from wild-type and pre-TCR-deficient mice by single cell polymerase chain reaction. Results show that the pre-TCR selects against gammadelta T cells containing rearranged Vbeta genes and that gammadelta T cell precursors but not gammadelta T cells express the pre-TCRalpha protein. Furthermore, pre-TCR-induced proliferation could not be detected in gammadelta T cells. We propose that the pre-TCR commits developing T cells to the alphabeta lineage by an instructive mechanism that has largely replaced an evolutionary more ancient stochastic mechanism of lineage commitment.

Animals↗
PubMed1998

Crucial function of the pre-T-cell receptor (TCR) in TCR beta selection, TCR beta allelic exclusion and alpha beta versus gamma delta lineage commitment.

The analysis of T-cell receptor (TCR) beta selection, TCR beta allelic exclusion and TCR beta rearrangement in gamma delta T cells from normal and pre-TCR-deficient mice has shown that the pre-TCR has a crucial role in T-lymphocyte development: The pre-TCR is by far the most effective receptor that generates large numbers of CD4+8+ T cells with productive TCR beta rearrangements. In the absence of the pre-TCR, TCR beta rearrangement proceeds in developing cells irrespective of whether they already contain a productive TCR beta gene. The pre-TCR directs developing T cells to the alpha beta lineage because gamma delta T cells from pT alpha-/- mice proceed much further in TCR beta rearrangement than gamma delta T cells from wild-type mice. It is argued that the pre-TCR commits developing T cells to the alpha beta lineage by an instructive mechanism, which has largely replaced an evolutionarily more ancient mechanism that involves stochastic alpha beta lineage commitment.

Alleles↗
PubMed1995

Cloning and comparative analysis of the human pre-T-cell receptor alpha-chain gene.

In immature T cells the T-cell receptor (TCR) beta-chain gene is rearranged and expressed before the TCR alpha-chain gene. At this stage TCR beta chain can form disulfide-linked heterodimers with the pre-T-cell receptor alpha chain (pTalpha). Using the recently isolated murine pTalpha cDNA as a probe, we have isolated the human pTalpha cDNA. The complete nucleotide sequence predicts a mature protein of 282 aa consisting of an extracellular immunoglobulin-like domain, a connecting peptide, a transmembrane region, and a long cytoplasmic tail. Amino acid sequence comparison of human pTalpha with the mouse pTalpha molecule reveals high sequence homology in the extracellular as well as the transmembrane region. In contrast, the cytoplasmic region differs in amino acid composition and in length from the murine homologue. The human pTalpha gene is expressed in immature but not mature T cells and is located at the p21.2-p12 region of the short arm of chromosome 6.

Amino Acid Sequence↗
PubMed1995

Crucial role of the pre-T-cell receptor alpha gene in development of alpha beta but not gamma delta T cells.

In T-cell precursors, the T-cell-receptor beta chain is expressed before the T-cell-receptor alpha chain and is sufficient to advance T-cell development in the absence of T-cell receptor alpha chains. In immature T cells, the T-cell-receptor beta protein can form disulphide-linked heterodimers with the pre-T-cell-receptor alpha chain and associate with signal-transducing CD3 molecules. The recently cloned pre-T-cell-receptor alpha gene encodes a transmembrane protein that is expressed in immature but not mature T cells. Here we show that alpha beta, but not gamma delta, cell development is severely hampered in pre-T-cell-receptor alpha-gene-deficient mice, which establishes a crucial role for the pre-T-cell receptor in early thymocyte development.

Animals↗
PubMed1995

Genomic structure and chromosomal location of the mouse pre-T-cell receptor alpha gene.

The mouse pre-T-cell receptor alpha (pT alpha) chain is a 33,000 M(r) glycoprotein expressed on the surface of immature thymocytes as a disulfide-linked heterodimer with the T-cell receptor beta (TCR beta) chain, and in association with proteins of the CD3 complex. The cDNA for pT alpha, isolated previously, encodes a type I transmembrane protein that is a member of the immunoglobulin (Ig) superfamily. Here we report the complete nucleotide sequence, the exon/intron structure, and the chromosomal location of the pTa gene. The gene spans about 8.4 kilobases (kb) and consists of four exons. Exon 1 encodes the 5' untranslated region, the leader peptide, and the first three amino acids of the mature protein. This exon is followed by a relatively long intron of 4.9 kb that contains many short interspersed repeats (SINEs) of the B1 and B2 family. The second exon encodes the extracellular Ig-like domain and exon 3 with just 45 base pairs the connecting peptide (CP), including the cysteine required for heterodimer formation. A similar exon/intron structure encoding corresponding parts of the mature polypeptide is found both in the Tcra and Tcrd constant region genes. The last exon encodes the transmembrane portion, the cytoplasmic tail, and about 540 nucleotides of 3' untranslated sequence, including a B2 repetitive element. In situ hybridization maps the pTa gene to the D/E1 region of mouse chromosome 17.

Animals↗
PubMed1994

Analysis and expression of a cloned pre-T cell receptor gene.

The T cell antigen receptor (TCR) beta chain regulates early T cell development in the absence of the TCR alpha chain. The developmentally controlled gene described here encodes the pre-TCR alpha (pT alpha) chain, which covalently associates with TCR beta and with the CD3 proteins forms a pre-TCR complex that transduces signals in immature thymocytes. Unlike the lambda 5 pre-B cell receptor protein, the pT alpha chain is a type I transmembrane protein whose cytoplasmic tail contains two potential phosphorylation sites and a Src homology 3 (SH3)-domain binding sequence. Pre-TCR alpha transfection experiments indicated that surface expression of the pre-TCR is controlled by additional developmentally regulated proteins. Identification of the pT alpha gene represents an essential step in the structure-function analysis of the pre-TCR complex.

Amino Acid Sequence↗
PubMed1992

Co-amplification of transcriptionally active epidermal growth factor receptor and ribosomal genes in the human hepatoma cell line Li7A.

A high level of expression of the functional product of the epidermal growth factor receptor (EGFR) gene was detected in the human hepatocarcinoma cell line Li7A and it was found to correlate with gene amplification. The karyotype was paratriploid, with 15 rearranged chromosomes, several of which contained abnormally banded regions (ABRs). The search for DNA sequences co-amplified with the EGFR gene, using the in-gel renaturation technique, allowed us to detect an amplified DNA band (La1) of about 30 kb. This DNA was used as a probe for in situ hybridization on chromosomes, to locate the amplified segment. In normal lymphocytes, the DNA of band La1 hybridized to chromosome regions in which repetitive DNAs are located, i.e. on juxtacentromeric regions, the site of alphoid and CCATT satellite DNA, and on the short arms of acrocentrics, the site of ribosomal RNA (RNR) genes. In Li7A cells, it hybridized to the same regions and, in addition, to several chromosome arms corresponding to ABRs. The same ABRs hybridized to EGFR and RNR probes, but neither Alu sequences nor various probes for other repetitive sequences were recognized. They also exhibited nucleolus organizer region staining characterizing functionally active (RNR) genes. It was concluded that transcriptionally active genes were co-amplified in the same ABRs, although they originated from different chromosomes, i.e. chromosome 7 for EGFR and acrocentrics for RNR genes.

Blotting, Northern↗
PubMed1991

Recurrent homogeneously staining regions in 8p1 in breast cancer and lack of amplification of POLB, LHRH, and PLAT genes.

In a cytogenetic study of 125 primary and untreated breast cancers, 107 were selected for the quality of their metaphases permitting detection of amplifications:homogeneously staining regions (HSRs), abnormally banded region (ABRs), and double minutes (dmins). HSRs and ABRs were detected in 62 cases (58%), but no cases of dmins were observed. The localizations of HSRs and ABRs were not random because they were observed in the 8p1 position in 14 cases. The possible amplifications of five sequences, MOS (8q1), LHRH (8p21.1), POLB (8p11.2), PLAT (8p12), and D8Z2 (8c) were investigated in three tumors with HSR on the short arm of chromosome 8. Because these sequences were not amplified, two interpretations can be proposed: 1) there is a frequent amplification of a sequence from the 8p1 region, located between the investigated sequences; and 2) the amplifications do not occur in 8p1, but HSRs or ABRs of undetermined origin have a strong tendency to be translocated onto 8p. Because cases with HSR(8p) have less complex karyotypes than with HSRs in other locations, the first interpretation is the most likely: HSRs may be formed in 8p and further translocated on other chromosomes in the course of tumor progression.

Breast Neoplasms↗
PubMed1991

A t(X;15)(q23;q25) with Xq reactivation in a lymphoblastoid cell line from Fanconi anemia.

A t(X:15)(q23;q25) was detected during cytogenetic investigation of a lymphoblastoid cell line established from a female patient with Fanconi anemia. The translocation was apparently balanced at passage 300 and unbalanced at passage 13. A chromatid exchange between both the normal and the der(15), between the centromere and band 15q25, may explain these results. Replication studies, following BrdU incorporation, indicate that the segment Xq23----qter from the der(15) is early replicating whereas segment Xpter----q23 from the der(X) is late replicating. Since the normal X was early replicating, it is concluded that the segment of the long arm of chromosome X, separated from its inactivation center by the translocation, was reactivated. This interpretation is confirmed by the methylation patterns of the hypoxanthine phosphoribosyltransferase gene (HPRT), mapped on Xq26, which corresponds to that of an active gene, whereas that of phosphoglycerate kinase (PGK1), which remained on the der(X), corresponds to that of an inactive gene. This is the first example of reactivation of a segment of the X chromosome following a structural rearrangement in somatic cells.

Cell Line↗
PubMed1991

GST pi gene is frequently coamplified with INT2 and HSTF1 proto-oncogenes in human breast cancers.

The glutathione S-transferase gene (GST pi) is located on the same chromosome band (11q13) as proto-oncogenes INT2 and HSTF1 which are frequently amplified in breast cancer. Using the Southern blot technique, we looked for the amplification of the GST pi gene in 17 fresh tumors from human mammary carcinoma. The tumors were preselected because either they had an amplification of the INT2 proto-oncogene detected by dot blot, or their karyotypes exhibited or did not exhibit homogeneously staining regions, a cytogenetic character indicating amplification. Coamplification of GST pi, HSTF1 and INT2 was observed in five tumors, and coamplification of GST pi and HSTF1 without amplification of INT2 in another tumor. We also observed coamplification of GST pi, INT2, HSTF1 in the mammary carcinoma cell line MDA/MB134, whereas GST pi alone was amplified in the mammary epithelial cell line HBL100. These results indicate that INT2, HSTF1 and GST pi belong to the same large amplicon. Since GST pi is involved in intracellular detoxication and since chemotherapeutic drugs are among its substrates, it will be of interest to study GST pi gene expression as well as the response to chemotherapy in patients presenting this amplicon.

Blotting, Southern↗
PubMed1990

Proto-oncogene amplification and homogeneously staining regions in human breast carcinomas.

Cytogenetic studies on fresh human breast cancers revealed that homogeneously staining regions (HSRs), which are assumed to represent DNA amplification, are observed in almost half of the cases. To search for a possible relationship between HSRs and proto-oncogene amplification, 16 proto-oncogenes, including ERBB2, were studied by Southern blot analysis in four tumors with two or three HSRs, and in three tumors without HSRs. Only four proto-oncogenes were found to be amplified in at least one tumor each: HST and INT2 (x3), MYC (x2-3), and FES (x greater than 10). The large sizes of the HSRs, which each corresponded to several percent of the haploid genome, were hardly compatible with the low rate of amplification, except for FES and then only if a large adjacent segment was co-amplified. This incomplete correlation was demonstrated by in situ hybridization, using biotinylated probes, which showed fluorescent spots on only one HSR for FES in one tumor and for INT2 in another one. Our results indicate that most of the large amplifications corresponding to HSRs do not involve the proto-oncogenes usually studied in breast cancer. The large amplification of FES, detected in one tumor, may be coincidental.

Blotting, Southern↗
PubMed1989

Molecular cloning and characterization of endogenous SV40 DNA from human HBL-100 cells.

The human HBL-100 cell line harbours SV40 DNA integrated in tandem at a unique site. The SV40 T-antigen expressed in these cells is defective in a function essential to the replication of the viral genome. The integrated SV40 sequences were molecularly cloned in a bacteriophage, and a subclone (plasmid pSVHBI) containing a complete SV40 DNA was isolated. As compared to SV40 wild-type strain 776, sequence analysis of pSVHBI early region revealed the presence of several DNA alterations. Among these, a point mutation at position 3199, predicting a change at amino-acid 540 of arginine to isoleucine, was shown by marker rescue to be responsible for the deficiency of T-antigen. This novel mutation further delimits one of the T-antigen domains involved in SV40 DNA replication. Transfection experiments demonstrated that the transforming activity of the SV40 genome from HBL-100 cells is still preserved. Moreover, several transformed human cell clones thus obtained could be permanently established in culture.

Base Sequence↗
PubMed1989

Acquisition of tumorigenic potential in the human myoepithelial HBL100 cell line is associated with decreased expression of HLA class I, class II and integrin beta 3 and increased expression of c-myc.

The human breast cell line HBL100 acquires the capacity to invade normal tissues and to replace them by proliferation in vitro only at high passage levels (HPL). These cells therefore are a useful model for studying tumor progression in vitro. We have analyzed the expression of cell-surface markers supposed to be involved in the control of the neoplastic process. Quantitative flow cytometry has revealed that: (1) spontaneous expression of HLA class-I antigens strongly decreases in HPL HBL100 cells vs. LPL cells, which parallels amplification and over-expression of c-myc oncogene; (2) HLA DR antigens can be induced by IFN-gamma in LPL but not in HPL HBL100 cells; (3) HBL100 cells secrete a soluble protein factor which specifically inhibits HLA DR induction by IFN-gamma even in heterologous cell systems; (4) 50% of LPL HBL100 cells express integrin beta 3, whereas HPL HBL100 cells lose this antigen; (5) this cell line is myoepithelial in origin, since 100% of HBL100 cells exhibit the CD10 antigen. Our data stress a role of HLA antigens, of some integrins and of c-myc in the acquisition of malignant potential by myoepithelial mammary cells of the HBL100 line.

Biomarkers, Tumor↗
PubMed1988

Accelerated malignant conversion of human HBL-100 cells by the v-Ki-ras oncogene.

The human epithelial HBL-100 cell line harbors SV40 genetic information and has an unlimited growth potential. Despite displaying properties characteristic of transformation since its early in vitro passages, it is capable of producing progressively growing tumors in nude mice only after long-term culture. This is a reproducible phenomenon and apparently not the consequence of a selection of preexisting malignant cells. Superinfection of early passage nontumorigenic HBL-100 cells with Kirsten murine sarcoma virus, which contains a Ki-ras oncogene having undergone multiple activating events, induces morphologic alterations and rapidly converts the cells to neoplastic cells, further supporting the hypothesis of multistep carcinogenesis. The HBL-100 cell line might be useful in defining the oncogenes representative of different families, which are able to complement SV40 in this system.

Cell Line↗