PTEN-mediated G1 cell-cycle arrest in LNCaP prostate cancer cells is associated with altered expression of cell-cycle regulators.

Background: The tumor suppressor PTEN regulates many biological processes. A well-known downstream effector of PTEN is phospho-Akt. Although PTEN is the most frequently inactivated gene in prostate cancer, its mode of action is not fully understood.

Both androgen receptor and glucocorticoid receptor are able to induce prostate-specific antigen expression, but differ in their growth-stimulating properties of LNCaP cells.

Androgen receptor-positive LNCaP cells were stably transfected with a rat glucocorticoid receptor (GR) expression plasmid. Ligand-binding studies in the generated cell lines revealed high-affinity binding of the cognate ligands to their receptors. Transfection experiments with the newly derived cell lines showed that, like androgen receptor, GR can induce activity of a prostate-specific antigen promoter fragment linked to the luciferase gene.

Androgen receptor mutations.

Male sexual differentiation and development proceed under direct control of androgens. Androgen action is mediated by the intracellular androgen receptor, which belongs to the superfamily of ligand-dependent transcription factors. At least three pathological situations are associated with abnormal androgen receptor structure and function: androgen insensitivity syndrome (AIS), spinal and bulbar muscular atrophy (SBMA) and prostate cancer.

Functional domains of the human androgen receptor.

A series of human androgen receptor (AR) deletion mutants was constructed to study the relationship between various structural domains and their different functions in the AR protein. Immunoblots of wild type AR and AR mutants expressed in COS-1 cells, revealed a doublet appearance of all AR proteins. One exception was an AR mutant lacking amino acid residues 51-211 that migrated as a single protein band, possibly due to altered post-translational modification.

The human androgen receptor: structure/function relationship in normal and pathological situations.

Discrete functions have been attributed to precise regions of the human androgen receptor (hAR) by expression of deletion mutants in COS and HeLa cells. A large C-terminal domain constitutes the hormone-binding region and a central basis, cysteine-rich domain is responsible for DNA binding. In addition, separate domains responsible for transactivation and nuclear translocation have been identified.

The promoter of the prostate-specific antigen gene contains a functional androgen responsive element.

Expression of prostate-specific antigen (PA) mRNA was tested at various time periods after incubation of the human prostate tumor cell line LNCaP with the synthetic androgen R1881. Androgen-stimulated expression was observed within 6 h after addition of R1881 to the cells. Run-on experiments with nuclei isolated from LNCaP cells showed that expression of the PA gene could be regulated by R1881 on the level of transcription.

The androgen receptor: functional structure and expression in transplanted human prostate tumors and prostate tumor cell lines.

The growth of the majority of prostate tumors is androgen-dependent, for which the presence of a functional androgen receptor is a prerequisite. Tumor growth can be inhibited by blockade of androgen receptor action. However, this inhibition is transient.

Variation of prostate-specific antigen expression in different tumour growth patterns present in prostatectomy specimens.

A series of 55 randomly chosen radical prostatectomy specimens was analyzed for expression of prostate-specific antigen (PSA) by immunohistochemical techniques. Tissue sections were selected in such a manner that in addition to glandular benign prostatic hyperplasia (BPH), one or more different prostatic tumour growth patterns were present. Four monoclonal antibodies, directed against three different PSA epitopes, and one polyclonal anti-PSA antiserum were used.

Characterization of polyclonal antibodies against the N-terminal domain of the human androgen receptor.

Antibodies against the N-terminal domain of the human androgen receptor (hAR) were prepared by two different approaches. Firstly, rabbits were immunized with a beta-galactosidase-hAR (amino acids (aa) 174-353) fusion protein. Secondly, two synthetic peptides corresponding to potentially antigenic sites located within this fragment (aa 201-222 and 301-320) were used as immunogens.

Structural organization of the human androgen receptor gene.

The complete coding region of the human androgen receptor gene has been isolated from a genomic library. The information for the androgen receptor was found to be divided over eight exons and the total length of the gene exceeded 90 kb. The sequence encoding the N-terminal region is present in one large exon.

The prostate-specific antigen gene and the human glandular kallikrein-1 gene are tandemly located on chromosome 19.

Using a prostate-specific antigen cDNA as a hybridization probe, clones containing the kallikrein genes encoding prostate-specific antigen, human glandular kallikrein-1 and pancreas/kidney kallikrein were isolated from a human genomic library. Clones containing the prostate-specific antigen gene and the human glandular kallikrein-1 gene overlap and span a region of about 36 kb. The two genes are aligned in a head to tail orientation at a mutual distance of 12 kb.

Characterization of the prostate-specific antigen gene: a novel human kallikrein-like gene.

Using Prostate-specific Antigen cDNA fragments as hybridization probes a clone containing the information for the gene encoding Prostate-specific Antigen was isolated form a human genomic DNA library. The complete gene (about 6 kb) was sequenced and shown to be composed of four introns and five exons. Two major transcription initiation sites were found.

The N-terminal domain of the human androgen receptor is encoded by one, large exon.

Using specific cDNA hybridization probes, the first coding exon of the human androgen receptor gene was isolated from a genomic library. The exon contained an open reading frame of 1586 bp, encoding an androgen receptor amino-terminal region of 529 amino acids. The deduced amino acid sequence was characterized by the presence of several poly-amino acid stretches of which the long poly-glycine stretch (16 residues) and the poly-glutamine stretch (20 residues) were most prominent.

Structure and function of the androgen receptor.

The androgen receptor in several species (human, rat, calf) is a monomeric protein with a molecular mass of 100-110 kDa. The steroid binding domain is confined to a region of 30 kDa, while the DNA-binding domain has the size of approx. 10 kDa.

The human androgen receptor: domain structure, genomic organization and regulation of expression.

The domain structure and the genomic organization of the human androgen receptor (hAR) has been studied after molecular cloning and characterization of cDNA and genomic DNA encoding the hAR. The cDNA sequence reveals an open reading frame of 2751 nucleotides encoding a protein of 917 amino acids with a calculated molecular mass of 98,845 D. The N-terminal region of the hAR is characterized by a high content of acidic amino acid residues and by several homopolymeric amino acid stretches.

Molecular cloning and characterization of novel prostate antigen cDNA's.

Three different prostate antigen cDNA's were isolated from a PC 82 prostate tumor cDNA library. PA 75 has a size of 1.4 kb and contains the almost complete information for the 35 kD prostate antigen preproprotein.

Cloning, structure and expression of a cDNA encoding the human androgen receptor.

A cDNA clone has been isolated from a library prepared of mRNA of human breast cancer T47D cells with an oligonucleotide probe homologous to part of the region encoding the DNA-binding domain of steroid receptors. The clone has a size of 1505 bp and sequence analysis revealed an open reading frame of 1356 bp. The deduced amino acid sequence displays two highly conserved regions identified as the putative DNA-binding and hormone binding domains respectively of steroid receptors.

Expression of cellular oncogenes in human prostatic carcinoma cell lines.

Prostatic cancer is one of the most frequent forms of malignancy in Western countries. Initially, growth of the majority of prostate tumors can be manipulated by endocrine therapy. However, ultimately androgen independent tumors continue to grow.

Mouse interferon alpha and beta genes are linked at the centromere proximal region of chromosome 4.

In order to determine the chromosomal localization of the murine interferon-alpha (MuIFN-alpha) and murine interferon-beta (MuIFN-beta) genes the DNAs of a panel of somatic cell hybrids were analysed by Southern blot hybridization. The hybrid cells were derived from E36 Chinese hamster cells and GRSL or GR MaTu mouse cells and retained all hamster chromosomes but segregated mouse chromosomes. The MuIFN-alpha probe used was a 0.

Isolation and characterization of subspecies of murine interferon alpha.

Interferon produced by mouse L-929 cells by incubation with poly(rI).poly(rC) is known to be composed of a mixture of MuIFN-alpha and MuIFN-beta. The alpha component was separated from the bete species by affinity chromatography over a monoclonal anti-MuIFN-beta agarose column and partially purified by gel filtration.