Influence of steroid hormones on pS2/BCEI gene expression in xenografted colon tumors.

The biological function of the hormone inducible human pS2/BCEI gene, cloned from the breast cancer cell line MCF-7, still remains unknown. Our aim was to determine the in vivo influence of steroid hormones on pS2 expression and tumour growth in colorectal tumours. We transplanted aliquots of human colon tumours into male and female nude mice and studied tumour growth and expression of the pS2/BCEI gene by immunostaining and mRNA analysis.

Tumor-specific demethylation of heterochromatic repetitive DNA in gastrointestinal cancer.

DNA methylation appears to play an important role in both physiological and experimentally modified gene expression. Alterations in DNA methylation have been described in animal tumour models, in transformed cells and tumour cell lines, usually for single copy DNA sequences. By applying the isoschizomeres HpaII and MspI the methylation status of two classes of repetitive sequences (D22Z2 - a member of the alphoid repeats and D22Z3 which belongs to the HinfI repeats) in colon and stomach carcinoma were tested.

Telomeric associations and loss of telomeric DNA repeats in renal tumors.

In a series of cytogenetically analyzed renal tumors of different histological types (chromophobe carcinoma, clear-cell carcinoma, chromophilic carcinoma, and oncocytoma) and normal renal tissue, the length of the telomeric DNA repeats was assessed using a (TTAGGG)3 oligonucleotide probe. A strong correlation was noted between pronounced telomere shortening and the appearance of telomeric associations of chromosomes. These data suggest an etiologic role of the loss of telomeric DNA repeats in the formation of telomeric associations and a possible involvement of this mechanism in the pathogenesis of chromosome aberrations in human tumors.

Application of DNA filter hybridization and PCR to distinguish between human and non-human tissues of poor quality.

The present report describes a novel approach for the identification of human or non-human specimens after long-term storage in a badly preserved state. The application of the PCR-technique (polymerase chain reaction) using human-specific primers as well as Southern blot (filter) hybridization of the sample DNA to a primate-specific DNA probe enabled us to extend the positive identification beyond the limits of conventional methods such as serological or morphological examinations.

High level functional expression of human beta 1-adrenergic receptor in baculovirus-infected cells screened by a rapid in situ procedure.

A novel screening assay for the identification of baculovirus infected cells expressing membrane receptors was developed by using a replica transfer technique. Sf9 cells were cotransfected with wild type baculoviral DNA and the transfer vector pVL941-beta 1 containing the coding region of the human beta 1-adrenergic receptor gene. Infected cells embedded in agarose were incubated with [125I]-iodocyanopindolol and transferred onto filters that were subsequently autoradiographed.

Expression of the breast cancer-associated protein pS2 in adenosquamous carcinomas of the gastrointestinal tract.

The breast cancer-associated protein pS2 is also present in many human gastrointestinal tumors. In contrast to breast carcinomas, gastrointestinal tumors do not express estrogen receptors, indicating that the expression of pS2 is not estrogen-dependent. The pS2 expression was analyzed in 14 adenosquamous tumors of the human gastrointestinal tract.

Nonradioactive southwestern analysis using chemiluminescent detection.

Replacing radioactively labeled probes by nonradioactive ones and detection by chemiluminescence instead of colorimetry allows a nonhazardous handling and offers the possibility of easily reprobing filters in Southwestern analysis. Using the described procedure, we were able to determine the molecular weight of DNA-binding proteins and achieve a high signal-to-noise ratio.

Assignment of the gene for human spasmolytic protein (hSP/SML1) to chromosome 21.

A human cDNA corresponding to the porcine pancreatic spasmolytic protein (PSP) was isolated, and the recombinant clone was originally termed hSP for human spasmolytic protein. Later, the term SML1 for spasmolysin was suggested for the human gene. This protein shows a remarkable sequence homology to pS2, a protein coded by an estrogen-induced gene isolated from the breast carcinoma cell line MCF-7.

Regional loss of the y-chromosome in urothelial carcinoma.

Specific chromosomal losses have been reported for various human tumors. We have now investigated ten cases of urothelial carcinoma and observed genomic alterations by a new method allowing detection of chromosomal losses directly in the tissue section. In 6 out of 8 male carcinomas, the Y-chromosome was lost either in single cells and isolated areas or in extended regions of the tumor sample.

A non-alphoid repetitive DNA sequence from human chromosome 21.

A non-alphoid repetitive DNA from human chromosome 22, consisting of a 48-bp motif, shows homology to both G-group chromosomes in the gorilla, thus indicating the presence of additional repeat family members on further human chromosomes. Therefore, we screened a chromosome-21-specific cosmid library using this repetitive sequence from chromosome 22 (D22Z3). Some 40-50 cosmid clones were positive in tests for hybridization.

A two-colour technique for chromosome in situ hybridization in tissue sections.

By extending non-isotopic in situ hybridization of DNA probes (targeted to metaphase chromosomes or interphase nuclei) to hybridization using tissue sections, additional topological information on the DNA structure and specific alterations can be obtained. We have established a method for the application of two different, chromosome-specific probes labelled with two colour dyes allowing simultaneous detection of two-colour signals. This method was tested in and is applicable to tissue sections of various origins.

In vitro study of a novel atypical beta-adrenoceptor agonist, SM-11044.

SM-11044 (L-threo-3-(3,4-dihydroxyphenyl)-N-[3-(4-fluorophenyl) propyl] serine pyrrolidine amide hydrobromide) stimulated the relaxation of guinea pig ileum (EC50: 3.0 x 10(-8) M), trachea (EC50: 1.3 x 10(-7) M), lung parenchyma (EC50: 2.

DNA-protein interaction analysis using polyacrylamide gel electrophoresis and a simple and sensitive UV crosslinking procedure.

A simple and reproducible technique for DNA-protein interaction analysis is described using UV crosslinking and polyacrylamide gel electrophoresis, leading to visualization of the complexes as distinct and strong signals. It avoids incorporation of bromodeoxyuridine (BrdU) into DNA and requires no special equipment. It was successfully applied to an oligonucleotide sequence from within the first intron of the mouse myb proto-oncogene and nuclear extracts from a myb-expressing cell line.

Tumor-specific methylation patterns of erbB2 (HER2/neu) sequences in gastro-intestinal cancer.

To determine whether the sporadically occurring amplification of the oncogene erbB2/HER2 in gastrointestinal carcinomas is associated with additional changes of this sequence, DNA from 17 colorectal and 5 stomach carcinomas was analyzed for copy number, sequence rearrangement and DNA methylation by Southern blot hybridization. Amplification was detected in two cases. By applying the isochizomers Hpall and Mspl we tested for alterations in the DNA methylation status.

Search for putative suppressor genes in meningioma: significance of chromosome 22.

Cytogenetic and molecular studies of various solid tumors have indicated that a series of different chromosomal regions may be deleted in the tumor genome. Usually, losses of heterozygosity are observed and, from this finding, the presence of specific genes acting as tumor suppressors has been deduced. In particular tumors, however, only a single chromosome site appears to be affected.

Association of the human spasmolytic polypeptide and an estrogen-induced breast cancer protein (pS2) with human pancreatic carcinoma.

The human pS2 gene, isolated from the breast carcinoma cell line MCF-7 and shown to be under estrogen transcriptional control in a subclass of breast cancer cells was reported to be secreted in normal stomach surface epithelial cells, whereas additional gastrointestinal tissues like pancreas and colon do not secrete pS2 at all. In porcine pancreas, a spasmolytic polypeptide (sharing domains of homology with pS2) was observed; a corresponding human gene (hSP) was shown to be active in normal stomach mucosa. hSP and pS2 gene activity in normal and neoplastic pancreas tissues was then compared.

Cross-hybridization between the avian myeloblastosis oncogene and eukaryotic 28S ribosomal RNA.

In Northern blots, avian myeloblastosis (myb) oncogene probes (genomic or cDNA) cross-hybridize to the 28S rRNA band mimicking a myb-specific transcript. A misinterpretation of the hybridization data can be avoided by using an oligodeoxyribonucleotide probe.

Molecular characterization of the mouse beta 3-adrenergic receptor: relationship with the atypical receptor of adipocytes.

The gene encoding the murine beta 3-adrenergic receptor (beta 3AR) has been isolated. It translates into a polypeptide of 388 amino acid residues which shows 82% overall homology with the human beta 3AR. In Southern blot experiments, a probe derived from the murine beta 3AR gene hybridizes to a unique restriction fragment in the murine and human genomes.

Atypical beta-adrenergic receptor in 3T3-F442A adipocytes. Pharmacological and molecular relationship with the human beta 3-adrenergic receptor.

Expression of ligand binding properties for an atypical beta-adrenergic receptor (beta-AR) subtype was studied during the adipose differentiation of murine 3T3-F442A cells and compared with that of the human beta 3-AR expressed in Chinese hamster ovary cells stably transfected with the human beta 3-AR gene (CHO-beta 3 cells) Emorine, L. J., Marullo, S.

Breast cancer-associated protein pS2 expression in tumors of the biliary tract.

Expression of pS2 (an estrogen-induced gene isolated from breast carcinoma MCF7 cells) and of the human spasmolytic peptide (hSP) gene was analyzed in 21 human biliary tract and gallbladder carcinomas and 16 non-neoplastic gallbladders at the protein level (immunochemistry) and, in a series of these cases, at the RNA level (Northern blots). Eighteen carcinomas and 14 non-neoplastic mucosae with inflammatory alterations showed pS2 activity by immunostaining or Northern blotting. In addition, in samples with pS2 expression, hSP RNA was demonstrated.

Large-scale physical mapping within the region 22q12.3-13.1 in meningioma.

The lack of physical mapping data strongly restricts the analysis of the meningioma chromosomal region that was assigned to the bands 22q12.3-qter. Recently, we reported a new marker D22S16 for chromosome 22 that was assigned to the region 22q13-qter by in situ hybridization.

A set of 4 nuclear proteins binds to a DNA sequence within the FOS promoter region.

We investigated DNA-protein-interactions occurring in the promoter region of c-fos using two-dimensional electrophoresis and south-western-blotting. When nuclear extracts from the human glioblastoma cell line HeRoSV were tested for their DNA-binding behaviour to a 650 bp-fragment within the promoter region of c-fos, we found 4 proteins designated as 120/6.6, 75/5.