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Biomedical subjects

J Shipley

Publications and source records attributed to J Shipley.

At least 73 records · Page 4Linked to original sources
PubMed1996

Interphase fluorescence in situ hybridization and reverse transcription polymerase chain reaction as a diagnostic aid for synovial sarcoma.

Identification of the t(X;18)(p11.2;q11.2) that is associated with a high proportion of synovial sarcoma can be a useful diagnostic aid. The translocation results in fusion of the SYT gene on chromosome 18 to either the SSX1 or the SSX2 gene, two homologous genes within Xp11.2. Two-color interphase fluorescence in situ hybridization and reverse transcription polymerase chain reaction were assessed as approaches to identify the rearrangement in well characterized cases. The presence of the translocation, and the specific chromosome X gene disrupted, were inferred from the configuration of signals from chromosome-specific centromere probes, paints, and markers flanking each gene in preparations of interphase nuclei. Rearrangement was found in two cell lines and eight of nine tumor samples, including analysis of five touch imprints. This was consistent with cytogenetic data in four cases and reverse transcription polymerase chain reaction analysis using primers known to amplify both SYT-SSX1 and SYT-SSX2 transcripts. The transcripts were distinguished by restriction with LspI and SmaI. Contrary to previous suggestions, there was no obvious correlation between histological subtype and involvement of the SSX1 or SSX2 gene. These approaches could also be applied to the identification of tumor-free margins and metastatic disease.

Adolescent↗
PubMed1995

Fusion of SYT to two genes, SSX1 and SSX2, encoding proteins with homology to the Kruppel-associated box in human synovial sarcoma.

We demonstrate that the cytogenetically defined translocation t(X;18)(p11.2;q11.2) found in human synovial sarcoma results in the fusion of the chromosome 18 SYT gene to either of two distinct genes, SSX1 or SSX2, at Xp11.2. The SSX1 and SSX2 genes encode closely related proteins (81% identity) of 188 amino acids that are rich in charged amino acids. The N-terminal portion of each SSX protein exhibits homology to the Kruppel-associated box (KRAB), a transcriptional repressor domain previously found only in Kruppel-type zinc finger proteins. PCR analysis demonstrates the presence of SYT-SSX1 or SYT-SSX2 fusion transcripts in 29 of 32 of the synovial sarcomas examined, indicating that the detection of these hybrid transcripts by PCR may represent a very useful diagnostic method. Sequence analysis has demonstrated heterogeneity in the fusion transcripts with the formation of two distinct SYT-SSX1 fusion junctions and two distinct SYT-SSX2 fusion junctions.

Adolescent↗
PubMed1995

Pez: a novel human cDNA encoding protein tyrosine phosphatase- and ezrin-like domains.

We have isolated cDNAs from normal human breast tissue and breast tumour cells that encode a protein (pez) with features of a novel non-receptor tyrosine phosphatase possessing N-terminal sequence homology to the ezrin-band 4.1-merlin-radixin protein family. Northern blot analysis indicates that pez is expressed in a variety of human tissues including kidney, skeletal muscle, lung and placenta. Fluorescence in situ hybridization has mapped pez to chromosome 1 region q32.2-41. Sequence identity to a characterized polymorphic marker confirms this localization.

Amino Acid Sequence↗
PubMed1995

Characterization of a t(10;11)(p13-14;q14-21) in the monoblastic cell line U937.

Previous analysis of the monoblastic cell line U937 has shown that several sublines contain a rearranged chromosome arm 11q. In order to determine the true nature of the rearrangement, fluorescence in situ hybridization (FISH) was carried out with various combinations of single copy anonymous markers, clones containing genes, a chromosome 10 paint, and an 11 centromere specific sequence. The rearrangement was deduced to be a reciprocal translocation between chromosomes 10 and 11 described as t(10;11)(p13-14;q14-21). The breakpoint on chromosome 11 is telomeric to the INT2 gene and the pHS11 probe at 11q13, and centromeric to the marker D11S36 localized to 11q14.3-q22.1 and the MLL gene at 11q23. Similar translocations have been reported in various acute leukemias, principally of the monocytic lineage, and also in T-cell precursor acute lymphocytic leukemias. Further characterization of the genetic rearrangements in U937 may lead to the isolation of genes important in leukemogenesis and provide an in vitro system for their study.

Chromosome Mapping↗
PubMed1994

Molecular analysis of simple variant translocations in acute promyelocytic leukemia.

The primary cytogenetic abnormality in acute promyelocytic leukemia (APL; FAB M3) is a reciprocal translocation, t(15;17)(q22;q12), which serves to fuse the PML gene on chromosome 15 to the retinoic acid receptor alpha (RARA) gene on chromosome 17. A PML-RARA fusion message transcribed from the der(15) is thought to mediate leukemogenesis. Two APL patients with simple variants of this translocation, t(3;15)(q21;q22) and t(X;15)(p11;q22), have previously been reported who lack cytogenetic involvement of chromosome 17, although their breakpoint positions on chromosome 15 still suggest the involvement of the PML gene. Here we report on a combined analysis by molecular genetics and in situ hybridization of these two patients, in which we wanted to determine whether the PML gene has alternative fusion partners or whether cryptic rearrangement of the RARA locus has occurred instead. A cryptic involvement of RARA was demonstrated in both patients by a combination of Southern analysis, reverse transcription coupled to PCR (RT-PCR), and fluorescence in situ hybridization. The results indicate an absolute requirement for the rearrangement of the RARA gene in the pathogenesis of APL and underline the importance of RARA during normal myeloid differentiation.

Adolescent↗
PubMed1994

Mapping of the human SAP1 (SRF accessory protein 1) gene and SAP2, a gene encoding a related protein, to chromosomal bands 1q32 and 12q23, respectively.

SAP1, SAP2, and ELK1 form a related subgroup of ETS-domain proteins that can form ternary complexes with the transcription factor SRF at the c-fos serum response element (SRE). SAP1 was identified by a genetic screen for proteins interacting with SRF expressed in yeast, and SAP2 by its homology with SAP1; ELK1 was previously identified by its homology to the ETS domain. cDNA probes were used to isolate cosmid and phage clones harboring genes encoding SAP1 and SAP2. These clones were subsequently used to map the genes to 1q32 and 12q23, respectively, by fluorescence in situ hybridization.

Chromosome Mapping↗
PubMed1994

Identification of novel genes, SYT and SSX, involved in the t(X;18)(p11.2;q11.2) translocation found in human synovial sarcoma.

Human synovial sarcomas contain a recurrent and specific chromosomal translocation t(X;18)(p11.2;q11.2). By screening a synovial sarcoma cDNA library with a yeast artificial chromosome spanning the X chromosome breakpoint, we have identified a hybrid transcript that contains 5' sequences (designated SYT) mapping to chromosome 18 and 3' sequences (designated SSX) mapping to chromosome X. An SYT probe detected genomic rearrangements in 10/13 synovial sarcomas. Sequencing of cDNA clones shows that the normal SYT gene encodes a protein rich in glutamine, proline and glycine, and indicates that in synovial sarcoma rearrangement of the SYT gene results in the formation of an SYT-SSX fusion protein. Both SYT and SSX failed to exhibit significant homology to known gene sequences.

Amino Acid Sequence↗
PubMed1994

Application of molecular genetics and cytogenetics to breast cancer and soft tissue sarcomas.

BACKGROUND: Over the last decade there have been considerable advances in understanding of the molecular events involved in the initiation and progression of a wide range of solid tumours. In many instances, cytogenetic abnormalities have been the first indication that there is a mutated gene at a particular locus. In colonic polyposis, for example, the loss of 5q material in a man with the disease indicated the importance of the region and led to the subsequent cloning of the gene. Interestingly, loss of this tumour suppressor gene is also seen in approximately 20%-50% of sporadic colorectal carcinomas, which suggests that it may be important to study these inherited syndromes to identify genes that may be more widely involved in carcinogenesis. It is also important to realize that the order in which different molecular events occur is unlikely to fit into predictable chronological patterns, and that there will be great variation in the molecular defects of tumours that otherwise appear morphologically similar and have a common pattern of clinical presentation and progression. This is because established tumours are probably composed of a number of subclones at the molecular level, and clonal selection will only take place if a particular mutation provides a selective growth advantage or it is necessary for a stage in progression, such as invasion or metastasis. Multiple sites of loss of heterozygosity (LOH) have been identified in a range of tumours, many of which may be sites of as yet unidentified tumour suppressor genes.(ABSTRACT TRUNCATED AT 250 WORDS)

Breast Neoplasms↗
PubMed1992

Inappropriate electrical countershocks by an automated external defibrillator.

A 79-year-old man who was in normal sinus rhythm with a palpable pulse was inappropriately shocked twice by a fully automated external defibrillator. The second shock resulted in ventricular tachycardia. The device then countershocked a third time, restoring normal sinus rhythm. The problem occurred primarily because the rescuers did not follow the device's instructions warning against applying the device in analysis mode to a patient with a palpable pulse, and they attempted to analyze the rhythm in a moving vehicle. Motion artifact and T waves that were relatively tall compared with the QRS amplitude were misinterpreted as ventricular fibrillation by the device. This example of inappropriate countershocks delivered by rescuers using a fully automated device illustrates the importance of proper training and continuing education.

Aged↗
PubMed1992

Identification of region-specific yeast artificial chromosomes using pools of Alu element-mediated polymerase chain reaction probes labeled via linear amplification.

The ability to identify large numbers of yeast artificial chromosomes (YACs) specific to any given genomic region rapidly and efficiently enhances both the construction of clone maps and the isolation of region-specific landmarks (e.g., polymorphic markers). We describe a method of preparing region-specific single-stranded hybridization probes from Alu element-mediated polymerase chain reaction (Alu-PCR) products of somatic cell hybrids for YAC library screening. Pools of up to 50 cloned Alu-PCR products from an irradiation-reduced hybrid containing 22q11.2-q13.1 were labeled to high specific activity by linear amplification using a single vector primer. The resulting single-stranded probes were extensively competed to remove repetitive sequences, while retaining the full complexity of the probe. Extensive coverage of the region by YACs using multiple probe pools was demonstrated as many YACs were detected more than once. In situ analysis using chosen YACs confirmed that the clones were specific for the region. Thus, this pooled probe approach constitutes a rapid method to identify large numbers of YACs relevant to a large chromosomal region.

Base Sequence↗
PubMed1991

A novel and rapid method for isolating sequences adjacent to rare cutting sites and their use in physical mapping.

We describe a simple PCR based technique which can be used to isolate sequences adjacent to rare cutter sites and can subsequently be employed for the construction of long range physical maps. The method involves the ligation of an adaptor to rare cutter sequences and its use as a target for forward priming in PCR. Primers to Alu repeat elements initiate synthesis of the reverse strand. Using this technique any rare cutter site which has a repeat element within amplification range can be cloned. We have isolated six unique sequences around NotI sites from an irradiation reduced hybrid containing a fragment of human chromosome 22 and are using these for physical mapping around the Ewing's sarcoma translocation breakpoint on chromosome 22.

Base Sequence↗
PubMed1989

Deletion of c-ets1 and T3 gamma loci from the 11q- chromosome in the human monoblastic cell line U937.

The 11q- chromosome in the human monoblastic cell line U937 appears to be derived from an interstitial deletion in bands 11q21-23/24 or a translocation with an unknown chromosome. We show here by in situ hybridisation that this chromosome has lost the c-ets1 and T3 gamma loci. C-ets1 and T3 gamma sequences were not detected on any chromosomes besides the normal 11, indicating either that they were lost from the genome or that the chromosome to which they were translocated was not present in a high enough proportion of the cells to be detected by in situ hybridisation. No DNA rearrangements were found with three different restriction enzymes in the c-ets1, N-CAM, Thy-1 and c-sea genes detected by our probes. There was also no detectable rearrangement in the c-fms gene which was shown to be translocated from chromosome 5 to chromosome 1 in a subline of U937. The size of the c-fms and c-ets1 messages were normal. The possible significance of these findings is discussed.

Blotting, Southern↗
PubMed1985

Molecular and cytologic analysis of DNA amplification in retinoblastoma.

Amplification of two distinct genomic DNA segments is observed in homogeneously staining regions in two sets of retinoblastoma cell lines derived from two different patients. One DNA segment was known to have sequence homology to the c-myc oncogene, and both DNA segments had previously been shown to be amplified in neuroblastoma cells. The absolute degree of amplification differed in all cytogenetically distinct retinoblastoma cell lines tested. Also, the relative amplification of these two DNA segments was unequal within a given cell line. Minimal amplification of both DNA segments was also detected in DNA directly isolated from one primary retinoblastoma. Based on these and previous results, it is concluded that assembly of amplifiable, relocatable units in many human retinoblastoma and neuroblastoma cells may involve a complex process of differential recruitment of separate DNA segments that are located on human chromosome #2.

Cell Line↗