Pirkko Vihko

SEVERAL GENES ENCODING RIBOSOMAL PROTEINS ARE OVER-EXPRESSED IN PROSTATE-CANCER CELL LINES: CONFIRMATION OF L7a AND L37 OVER-EXPRESSION IN PROSTATE-CANCER TISSUE SAMPLES

A cDNA library specific for mRNA over‐expressed in prostate cancer was generated by subtractive hybridization of transcripts originating from prostatic hyperplasia and cancer tissues. cDNA encoding ribosomal proteins L4, L5, L7a, L23a, L30, L37, S14 and S18 was found to be present among 100 analyzed clones. Levels of ribosomal mRNA were significantly higher at least in one of the prostate‐cancer cell lines, LNCaP, DU‐145 and PC‐3, than in hyperplastic tissue, as determined by slot‐blot hybridization. Furthermore, L23a‐ and S14‐transcript levels were significantly elevated in PC‐3 cells as compared with those in the normal prostate epithelial cell line PrEC. Generally, dramatic changes in the mRNA content of the ribosomal proteins were not detected, the most evident over‐expression being that of L37 mRNA, which was 3.4 times more abundant in LNCaP cells than in hyperplastic prostate tissue. The over‐expression of L7a and L37 mRNA was confirmed in prostate‐cancer tissue samples by in situ hybridization. Elevated cancer‐related expression of L4 and L30 has not been reported, but levels of the other ribosomal proteins are known to be increased in several types of cancers. These results therefore suggest that prostate cancer is comparable with other types of cancers, in that a larger pool of some ribosomal proteins is gained during the transformation process, by an unknown mechanism. Int. J. Cancer 78:27–32, 1998.© 1998 Wiley‐Liss, Inc.

The TMPRSS2 gene encoding transmembrane serine protease is overexpressed in a majority of prostate cancer patients: Detection of mutated TMPRSS2 form in a case of aggressive disease

The serine protease TMPRSS2 gene expression was studied by in situ hybridization using benign prostatic hyperplasia and prostate cancer tissue samples from 32 patients. Expression of TMPRSS2 gene was higher in cancer cells than that in benign cells in 84% of the specimens containing both benign and malignant tissues. The TMPRSS2 mRNA level was significantly increased in poorly differentiated (p = 0.014, n = 7) and untreated (p = 0.022, n = 13) primary prostate adenocarcinomas compared to benign tissues. In addition, androgen‐deprivation therapy significantly decreased the expression of TMPRSS2 in benign prostate tissue (p = 0.07), which is in accordance with the androgen‐inducible expression of the gene. The gene copy number of TMPRSS2, analyzed by competitively differential PCR, was duplicated in the malignant cells of about 38% of the prostate cancer patients analyzed. Thus, the increase in the gene copy number is probably not the primary reason for the detected overexpression of the TMPRSS2 gene. Mutations in the TMPRSS2 gene were screened using DNA isolated from paraffin‐embedded prostate cancer tissues from 9 patients with aggressive prostate cancer and from 9 patients with nonaggressive disease. Thirteen exons covering the coding region were checked using enzymatic mutation detection and direct sequencing. One patient with aggressive prostate cancer carried a deletion and a stop codon in exon 11, leading to inactivation of the serine protease domain in TMPRSS2.

Characterization of Molecular and Catalytic Properties of Intact and Truncated Human 17β-Hydroxysteroid Dehydrogenase Type 2 Enzymes: Intracellular Localization of the Wild-Type Enzyme in the Endoplasmic Reticulum1

Human 17beta-hydroxysteroid dehydrogenase (17HSD) type 2 is a widely distributed enzyme that primarily converts the highly active 17beta-hydroxysteroids to their inactive keto forms. In the present study, full-length human 17HSD type 2 was localized in the endoplasmic reticulum using a double immunofluorescence labeling technique. As a consequence of its strong membrane interaction, full-length human 17HSD type 2 could not be solubilized as a biologically active form in vitro. However, by deleting the first 29 amino acids from the N-terminus, we were able to purify a catalytically active enzyme from the cytosolic fraction of Sf9 insect cells. Biochemical and catalytic properties of the purified truncated human 17HSD type 2 protein confirm its suitability for structure-function analyses of the enzyme. Both intact and truncated 17HSD type 2 enzymes efficiently catalyzed the oxidation of estradiol, testosterone, dihydrotestosterone, androstenediol, and 20alpha-dihydroprogesterone. The oxidation of estradiol brought about by human 17HSD type 2 was effectively inhibited by several other steroidal compounds, such as 2-hydroxyestradiol, 5beta-androstan-3alpha,17beta-diol, 5alpha-androstan-3alpha,17beta-diol, and 5alpha-androstan-3beta,17beta-diol. The broad substrate specificity of human 17HSD type 2 together with its predominant oxidative activity and intracellular location, as observed in this study, indicate the physiological role of the enzyme to be primarily an inactivator of highly active 17beta-hydroxysteroids.

Expression of transmembrane serine protease TMPRSS2 in mouse and human tissues.

The purpose of this study was to clarify the expression of TMPRSS2 in mice during development and to compare the tissue distribution of the transcripts in adult mouse and human tissues. Mouse TMPRSS2u2009cDNA was cloned; the predicted amino acid sequence contains 490 residues sharing 81.4% similarity with human TMPRSS2. According to northern blots, mouse TMPRSS2 is expressed mainly in the prostate and kidney, while human TMPRSS2 is expressed in the prostate, colon, stomach, and salivary gland. In situ hybridization analyses of mouse embryos and adult tissues revealed that TMPRSS2 was expressed in the epithelia of the gastrointestinal, urogenital, and respiratory tracts. Expression was very selective and constant after the gene was turned on during development. Expression of TMPRSS2 was localized in the luminal epithelial cells of the mouse and human prostate. The information presented here will be useful in further studies regarding the function and physiological significance of TMPRSS2. Copyright

Expression of Mouse 17β-Hydroxysteroid Dehydrogenase/17-Ketosteroid Reductase Type 7 in the Ovary, Uterus, and Placenta: Localization from Implantation to Late Pregnancy1

Rodent 17β-hydroxysteroid dehydrogenase/17-ketosteroid reductase type 7 (17HSD/KSR7) catalyzes the conversion of estrone (E1) to estradiol (E2) and is abundantly expressed in the ovaries of pregnant animals in particular. In the present work we demonstrate cell-specific expression of 17HSD/KSR7 in the ovaries, uteri, and placentas of pregnant and nonpregnant mice using in situ hybridization. The results show that mouse 17HSD/KSR7 (m17HSD/KSR7) messenger RNA is distinctly and exclusively expressed in a proportion of corpora lutea (CLs). During pregnancy, expression of m17HSD/KSR7 is most abundant around embryonic day 14.5 (E14.5), when the ovaries are filled with CLs expressing 17HSD/KSR7. In the uterus, m17HSD/KSR7 is first detected on E5.5, when expression surrounds the implantation site on the antimesometrial side. As gestation progresses, m17HSD/KSR7 is expressed in the decidua capsularis on E8 and E9.5, disappearing thereafter from the antimesometrial decidua. On E9 onward, m17HSD/KSR7 messenger RNA e...

Comparison of human prostate specific glandular kallikrein 2 and prostate specific antigen gene expression in prostate with gene amplification and overexpression of prostate specific glandular kallikrein 2 in tumor tissue

There is a need for specific markers that can indicate the different stages of prostate carcinoma. There is ongoing speculation concerning the value of prostate specific glandular kallikrein (hK2) in this regard.

Identification of Essential Subelements in the hHSD17B1 Enhancer: Difference in Function of the Enhancer and That of the hHSD17BP1 Analog Is due to −480C and− 486G

The function of the gene encoding human 17β-hydroxysteroid dehydrogenase (17HSD) type 1, the hHSD17B1 gene, is regulated by a cell-specific enhancer at position −662 to −392. The adjacent hHSD17BP1 gene, whose function is not known, contains an analogous region in its 5′-flanking region. The identity between the hHSD17B1 enhancer and the hHSD17BP1 equivalent is as high as 98%, i.e. they differ by only five nucleotides. Results from reporter gene analyses showed that the hHSD17BP1 analog, a pseudoenhancer, has only 10% the activity of the hHSD17B1 enhancer. Furthermore, the results indicate that the reduced function of the pseudoenhancer is a consequence of the presence of G and A at positions −480 and −486, whereas the hHSD17B1 enhancer contains −480C and −486G. In addition, three protected areas were localized to regions −495/−485 (FP1), −544/−528 (FP2), and −589/−571 (FP3) in deoxyribonuclease I footprinting analysis of the hHSD17B1 enhancer. Replacement of the footprinted regions with a nonsense sequen...