Olli Jänne - Academia.edu (original) (raw)

Papers by Olli Jänne

Experimental Cell Research, Dec 1, 1995

Changes in cell shape occur during the cell cycle and influence cell proliferation and differenti... more Changes in cell shape occur during the cell cycle and influence cell proliferation and differentiation. In order to study how altered cell proliferation and cell shape are interrelated, we have studied ornithine decarboxylase (ODC) regulation in cultured normal human epidermal keratinocytes (NHEK). Cytoskeletal disruptors have been reported to modulate regulation of ODC; the products of ODC, the polyamines, influence actin polymerization rates in vitro, and polyamine auxotrophs have profoundly disrupted cytoskeletons. Therefore, altered ODC levels could be involved in signaling changes in cell shape and an intact cytoskeleton could transduce signals to regulate ODC levels. We had previously observed that the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), which profoundly alters cell shape, markedly suppresses ODC biosynthesis in NHEK solely at posttranscriptional/protein synthesis levels. TPA treatment caused NHEK to rapidly assume a rounded morphology that was accompanied by a change in actin organization, as determined by rhodamine-phalloidin labeling. Immunolocalization of ODC showed a perinuclear/nuclear distribution in untreated NHEK and a more diffuse pattern after TPA treatment that was apparent within 15-30 min. Changes in ODC enzyme activity are not significant until 60 min after TPA treatment. NHEK treated with cytochalasin B or D to inhibit actin polymerization exhibited a diffuse ODC localization that could be reversed by removal of the cytochalasin; inhibition of ODC by alpha-difluoromethylornithine caused a diffuse ODC localization. All treatments resulted in cytoskeletal remodeling. These data are the first evidence for a distinct subcellular localization for ODC and suggest that changes in ODC localization may be an initial step in regulation of ODC activity. Furthermore, changes in ODC activity cause an altered cytoskeleton, suggesting one means by which growth regulatory signals can be transduced to the cytoskeleton from various signaling pathways.

Molecular Endocrinology, May 1, 2001

Molecular and Cellular Biology, Dec 1, 2003

Modification by acetylation occurs at-amino lysine residues of histones and transcription factors... more Modification by acetylation occurs at-amino lysine residues of histones and transcription factors. Unlike phosphorylation, a direct link between transcription factor acetylation and cellular growth or apoptosis has not been established. We show that the nuclear androgen receptor (AR), a DNA-binding transcriptional regulator, is acetylated in vivo. The acetylation of the AR is induced by ligand dihydrotestosterone and by histone deacetylase (HDAC) inhibitors in living cells. Direct AR acetylation augmented p300 binding in vitro. Constructs mimicking neutral polar substitution acetylation (AR K630Q , AR K630T) enhanced p300 binding and reduced N-CoR/HDAC/Smad3 corepressor binding, whereas charged residue substitution (AR K630R) reduced p300 binding and enhanced corepressor binding. The AR acetylation mimics promoted cell survival and growth of prostate cancer cells in soft agar and in nude mice and augmented transcription of a subset of growth control target gene promoters. Thus, transcription factor acetylation regulates coactivator/corepressor complex binding, altering expression of specific growth control genes to promote aberrant cellular growth in vivo.

Journal of Cell Science, 2000

The androgen receptor (AR) is a transcription factor that mediates androgen action. We have used ... more The androgen receptor (AR) is a transcription factor that mediates androgen action. We have used the green fluorescent protein (GFP) technique to investigate dynamics of nuclear trafficking of human AR in living cells. In the absence of ligand, the GFP-AR fusion protein is distributed between cytoplasm and nuclei. Androgen exposure leads to a rapid and complete import of GFP-AR to nuclei of CV-1 cells (≥90% nuclear in 30 minutes), whereas a pure antiandrogen, Casodex, elicits a slower (<40% nuclear in 30 minutes) and incomplete transfer. Unliganded ARs with mutations in the basic amino acids of the bipartite nuclear localization signal (NLS) within the second zinc finger and the hinge region are predominantly cytoplasmic and their androgen-dependent nuclear import is severely compromised (≤20% nuclear in 30 minutes). Interestingly, substitutions of the Leu residues flanking the bipartite NLS lead to inefficient nuclear transfer in response to androgen (≤20% nuclear in 30 minutes)...

Molecular and Cellular Biology, 1998

Using the DNA-binding domain of androgen receptor (AR) as a bait in a yeast two-hybrid screening,... more Using the DNA-binding domain of androgen receptor (AR) as a bait in a yeast two-hybrid screening, we have identified a small nuclear RING finger protein, termed SNURF, that interacts with AR in a hormone-dependent fashion in both yeast and mammalian cells. Physical interaction between AR and SNURF was demonstrated by coimmunoprecipitation from cell extracts and by protein-protein affinity chromatography. Rat SNURF is a highly hydrophilic protein consisting of 194 amino acid residues and comprising a consensus C 3 HC 4 zinc finger (RING) structure in the C-terminal region and a bipartite nuclear localization signal near the N terminus. Immunohistochemical experiments indicated that SNURF is a nuclear protein. SNURF mRNA is expressed in a variety of human and rat tissues. Overexpression of SNURF in cultured mammalian cells enhanced not only androgen, glucocorticoid, and progesterone receptor-dependent transactivation but also basal transcription from steroid-regulated promoters. Mutat...

Gene Regulation by Steroid Hormones III, 1987

Physiological and synthetic androgens bring about their actions in murine kidney via androgen rec... more Physiological and synthetic androgens bring about their actions in murine kidney via androgen receptor-mediated mechanisms in a fashion similar to that of other steroid hormones through their respective receptor proteins in a variety of target tissues. Unlike some other steroid-responsive tissues or cell lines, in which large amounts of a few proteins are induced (O’Malley and Means, 1974; O’Malley et al., 1979; Tata and Smith, 1979), androgen action in mouse kidney involves induction of many proteins that are of low abundancy, with none predominating as a percentage of total protein concentration. For example, two-dimensional gel electrophoretic analysis of soluble renal proteins could not reveal any major differences in their profile in untreated and androgen-induced mice (Swank et al., 1978). These considerations, along with well-established mouse genetics, render the murine kidney an attractive model system to study steroid hormone action and, in particular, to investigate regulation of a number of gene products within a single cell type and by a single steroid hormone.

Molecular and Cellular Biology, 2002

PIAS (protein inhibitor of activated STAT) proteins interact with and modulate the activities of ... more PIAS (protein inhibitor of activated STAT) proteins interact with and modulate the activities of various transcription factors. In this work, we demonstrate that PIAS proteins xα, xβ, 1, and 3 interact with the small ubiquitin-related modifier SUMO-1 and its E2 conjugase, Ubc9, and that PIAS proteins themselves are covalently modified by SUMO-1 (sumoylated). PIAS proteins also tether other sumoylated proteins in a noncovalent fashion. Furthermore, recombinant PIASxα enhances Ubc9-mediated sumoylation of the androgen receptor and c-Jun in vitro. Importantly, PIAS proteins differ in their abilities to promote sumoylation in intact cells. The ability to stimulate protein sumoylation and the interaction with sumoylated proteins are dependent on the conserved PIAS RING finger-like domain. These functions are linked to the activity of PIASxα on androgen receptor-dependent transcription. Collectively, our results imply that PIAS proteins function as SUMO-1-tethering proteins and zinc finge...

Molecular Endocrinology, 2004

GnRH controls expression of the LH subunit genes, α and LHβ, with the LHβ subunit regulated most ... more GnRH controls expression of the LH subunit genes, α and LHβ, with the LHβ subunit regulated most dramatically. Two enhancer regions, distal and proximal, on the rat LHβ gene promoter cooperate for full basal expression and GnRH stimulation. It has been hypothesized that the transcription factors binding to these regions, Sp1, Egr-1, and steroidogenic factor 1 (SF-1), may interact directly or indirectly via a coactivator. One such coactivator may be small nuclear RING finger protein (SNURF), which is expressed in pituitary tissue and the LβT2 gonadotrope cell line. In transfection experiments in LβT2 cells, SNURF stimulated basal expression of LHβ and increased overall GnRH stimulation. SNURF specifically stimulated LHβ, with no effect on the α-subunit promoter. SNURF interacts with Sp1 and SF-1, but not Egr-1, in pull-down experiments. Point mutations or deletions of SNURF functional domains demonstrated that Sp1 and SF-1 interactions with SNURF are required for SNURF stimulatory ef...

Molecular Endocrinology, 1993

A series of deletion mutants was constructed for the rat androgen receptor (AR) to delineate sequ... more A series of deletion mutants was constructed for the rat androgen receptor (AR) to delineate sequences involved in transcriptional activation. Using transient expression conditions in CV-1 cells and in vitro DNAbinding studies, the amino-terminal domain of the receptor was shown to contain a region (residues 147-296) that is mandatory for trans-activation. Receptors with deletions (residues 147-408) in the N-terminal domain, but with intact DNA-and ligandbinding domains, interacted in vitro with androgenresponsive elements albeit with affinities lower than that of the wild type receptor. Coexpression of Nterminal deletion mutants (1146-408 and A38-296) with the wild type AR blunted trans-activation by the latter protein in a dominant fashion.

Molecular Endocrinology, 1997

Androgen, glucocorticoid, and progesterone receptors (ARs, GRs, and PRs) often can regulate trans... more Androgen, glucocorticoid, and progesterone receptors (ARs, GRs, and PRs) often can regulate transcription via composite hormone response elements in target genes. We have used artificial and natural mutant ARs from patients with androgen resistance to study their effects on dominant negative activity on wild type AR, GR, and PR function on mouse mammary tumor virus (MMTV) and tyrosine aminotransferase (TAT) promoters. Artificial ARs that contained internal deletions within the amino-terminal region had minimal transcriptional activity but blocked ligand-mediated transcription by wild type AR. Mutants containing deletions of the DNA-binding and ligand-binding domains had minimal or weak dominant negative activity. We then tested the ability of wild type and mutant ARs to modulate GRand PR-mediated transcriptional activity. The amino-terminal deletion mutants exerted dominant negative effects on GRand PR-mediated activity, both in the absence and presence of testosterone. Surprisingly, wild type AR, which had approximately 20% of the maximal transcriptional activity of GR on the MMTV promoter, also had dominant negative activity on dexamethasone-regulated transcription mediated by GR. This dominant negative activity likely involves DNA binding because a point mutation in the DNAbinding domain abrogated such activity of an aminoterminal deletion mutant. Additionally, natural human AR mutants from patients with androgen resistance, which do not bind either DNA or ligand, did not block dexamethasone-mediated transcription. In summary, these studies suggest that mutant and wild type ARs can display dominant negative activity on other steroid hormone receptors that bind to a composite hormone response element. This cross-regulation may be important in regulating maximal transcriptional activity in tissues where these receptors are coexpressed and may contribute to the phenotype of patients with steroid hormone resistance.

Molecular Endocrinology, 2007

An adenosine triphosphatase of the sucrose nonfermenting 2 protein family, androgen receptorinter... more An adenosine triphosphatase of the sucrose nonfermenting 2 protein family, androgen receptorinteracting protein 4 (ARIP4), modulates androgen receptor activity. To elucidate receptor-dependent and-independent functions of ARIP4, we have analyzed Arip4 gene-targeted mice. Heterozygous Arip4 mutants were normal. Arip4 is expressed mainly in the neural tube and limb buds during early embryonic development. Arip4 ؊/؊ embryos were abnormal already at embryonic d 9.5 (E9.5) and died by E11.5. At E9.5 and E10.5, almost all major tissues of Arip4-null embryos were proportionally smaller than those of wild-type embryos, and the neural tube was shrunk in some Arip4 ؊/؊ embryos. Dramatically reduced cell proliferation and increased apoptosis were observed in E9.5

Molecular and Cellular Biology, 2008

To elucidate SUMO-1 functions in vivo, we targeted by homologous recombination the last three exo... more To elucidate SUMO-1 functions in vivo, we targeted by homologous recombination the last three exons of the murine Sumo-1 gene. Sumo-1 mRNA abundance was reduced to one-half in heterozygotes and was undetectable in Sumo-1 −/− mice, and SUMO-1-conjugated RanGAP1 was detectable in wild-type mouse embryo fibroblasts (MEFs) but not in Sumo-1 −/− MEFs, indicating that gene targeting yielded Sumo-1 -null mice. Sumo-1 mRNA is expressed in all tissues of wild-type mice, and its abundance is highest in the testis, brain, lungs, and spleen. Sumo-2 and Sumo-3 mRNAs are also expressed in all tissues, but their abundance was not upregulated in Sumo-1 -null mice. The development and function of testis are normal in the absence of Sumo-1 , and Sumo-1 − / − mice of both sexes are viable and fertile. In contrast to a previous report (F. S. Alkuraya et al., Science 313:1751, 2006), we did not observe embryonic or early postnatal demise of Sumo-1 -targeted mice; genotypes of embryos and 21-day-old mice...

Laboratory Investigation, 2003

Androgen action is mediated through androgen receptor (AR), which appears to undergo structural a... more Androgen action is mediated through androgen receptor (AR), which appears to undergo structural and functional alterations during prostate cancer (CaP) progression. AR mutations have been infrequently reported in CaP before hormonal therapy, but in untreated, advanced tumors AR mutations are suggested to be more common. To investigate the frequency of AR mutations in aggressive CaP before hormonal therapy, we have analyzed AR coding region for aberrations in 21 paraffin-embedded prostate carcinoma samples (14 primary tumors, 7 metastases) of poor histologic differentiation. Singlestranded conformational polymorphism and sequencing analyses revealed AR missense mutations in 29% (4/14) of the primary tumors and in one (14%) metastasis. Mutations resided in the transactivation domain and in the hinge region. One of the hinge region mutants, Ser646Phe, that was identified in a patient with short endocrine therapy response, exhibited a markedly increased transcriptional activity on single androgen response element-containing promoters. In conclusion, AR mutations are frequent in high-grade CaP before initiation of hormonal therapy, and these mutations may play a role in poor therapy response and emergence of hormone-refractory CaP in some cases.

The Journal of Clinical Endocrinology & Metabolism, 2004

We have examined the relationship between serum androgen bioactivity, as measured with a recombin... more We have examined the relationship between serum androgen bioactivity, as measured with a recombinant cell bioassay, and progression of puberty in 14 boys with constitutional delay of puberty. Six boys were followed up without treatment (control group), and eight boys received low-dose (1 mg/kg) testosterone enanthate im for 0-6 months together with an aromatase inhibitor, letrozole, 2.5 mg orally once a day for 0-12 months (treatment group). In the control group, serum androgen bioactivity increased during the course of puberty (P < 0.001). During 0-12 months of the study, the boys in the treatment group had higher androgen bioactivity levels (P < 0.05) and faster rate of pubic hair growth than the control boys (P < 0.05). Overall, the average serum androgen bioactivity during 12 months of follow-up correlated strongly with the concomitant changes in Tanner genital (r S ‫؍‬ 0.89; n ‫؍‬ 13; P < 0.005) and pubic hair stages (r S ‫؍‬ 0.79; n ‫؍‬ 13; P < 0.01). In conclusion, our results suggest that circulating androgen bioactivity mediates the tempo of pubertal maturation and that the combination of testosterone and letrozole given to boys with constitutional delay of puberty accelerates puberty.

Journal of Biological Chemistry, 2002

The steroid receptor coactivator (SRC) proteins comprise a well-characterized family of nuclear r... more The steroid receptor coactivator (SRC) proteins comprise a well-characterized family of nuclear receptor (NR) coactivators that increase transcriptional activation by NRs via covalent modification of chromatin proteins and recruitment of other coactivators. We have recently shown that the SRC family member GRIP1 interacts with a class of SUMO-1 (small ubiquitin-like modifier 1) E3 ligases, the PIAS proteins, and that the coactivator is subjected to SUMO-1 modifications (sumoylation). In this work, we demonstrate that lysine residues 239, 731, and 788 of GRIP1 serve as principal attachment sites for SUMO-1. Lys-731 and Lys-788 are located in the NR interaction domain (NID), and their substitution by arginines impairs the ability of GRIP1 to colocalize with androgen receptor (AR) in nuclei. Likewise, Lys-731 and Lys-788 mutants of GRIP1 have attenuated ability to enhance AR-dependent transcription and fail to synergize with PIASx␤-mediated activation of AR function, indicating that sumoylation modifies the ability of GRIP1 to function as a steroid receptor coactivator. The Lys-731 sumoylation site is conserved in SRC-3 and SRC-1, and the NIDs of the latter coactivators harbor one or two additional sites matching with the consensus sites for SUMO-1 attachment, respectively, suggesting a more general role for the modification in the regulation of SRC protein activity.

Journal of Biological Chemistry, 2000

Journal of Biological Chemistry, 1996

Cross-modulation between androgen receptor (AR) and NF-B/Rel proteins was studied using various a... more Cross-modulation between androgen receptor (AR) and NF-B/Rel proteins was studied using various androgen-and NF-B-regulated reporter genes under transient transfection conditions. In COS-1 cells, elevated expression of RelA (p65) repressed AR-mediated transactivation in a dose-dependent manner, whereas NFB1 (p50), another major member of the NF-B family, did not influence transactivation. The repression of AR appeared to involve the N-terminal region of the protein between residue 297 and the DNA-binding domain. RelA-mediated transrepression could not be overcome by increasing the amount of AR. Transcriptional interference between RelA and AR was mutual in that cotransfected AR was able to attenuate transactivation by RelA in a dose-and steroid-dependent fashion. An excess of RelA was able to rescue the repression to some extent. Immunological analyses of RelA and AR protein levels indicated that transrepression was not due to reciprocal decrease in their amounts. Neither did AR increase the concentration of IB␣, which can sequester and inactivate RelA. Electrophoretic mobility shift assays using extracts from cotransfected cells and purified recombinant proteins showed that AR and RelA did not significantly influence each other's DNA binding activity. Nevertheless, protein-protein interaction experiments demonstrated a weak association between AR and RelA. Collectively, these data suggest that the mutual repression in intact cells is due to formation of AR-RelA complexes that are held together by another partner or to competition for a coactivator required for transcription. Androgen receptor (AR) 1 belongs to the superfamily of ligand-activated transcription factors, the nuclear receptors (1, 2). AR regulates the development, differentiation, and maintenance of male reproductive functions. In addition, androgens are involved in the regulation of other sexually dimorphic

Journal of Biological Chemistry, 1999

We have characterized a novel partner for androgen receptor (AR), termed ARIP3, that interacts wi... more We have characterized a novel partner for androgen receptor (AR), termed ARIP3, that interacts with the DNA-binding domain/zinc finger region of AR and is predominantly expressed in the testis. Rat ARIP3 is a nuclear protein comprising 572 amino acids. It modulates AR-dependent but not basal transcription, suggesting that ARIP3 acts as an AR transcriptional coregulator. Except for the C-terminal AR-interacting domain, ARIP3 contains distinct regions that are also present in two recently described proteins, a protein inhibitor of activated Stat3 and an RNA helicase II-interacting protein (Gu/RH-II binding protein). Conserved structural features of these proteins indicate the existence of a gene family involved in the regulation of various transcription factors. Collectively, ARIP3 belongs to a novel nuclear protein family and is perhaps the first tissuespecific coregulator of androgen receptor.

FEBS Letters, 2002

We have used confocal microscopy to elucidate the e¡ects of antiandrogens on nuclear localization... more We have used confocal microscopy to elucidate the e¡ects of antiandrogens on nuclear localization of the androgen receptor (AR) with its transcriptional coactivator GRIP1. We show that the agonist-activated AR recruits GRIP1 to colocalize with the receptor in the nucleoplasm. By contrast, AR complexed to the antiandrogens hydroxy£utamide and bicalutamide fails to in£uence nuclear distribution of GRIP1. Likewise, the non-steroidal antiandrogens prevent the agonist-induced ARĜ RIP1 colocalization from occurring. Androgen antagonists a¡ect nuclear redistribution of AR^GRIP1 in a fashion that parallels their e¡ects on the transcriptional activity of AR, in that the pure antagonists block GRIP1-dependent activation of AR function, whereas the mixed antagonist/agonist cyproterone acetate promotes both AR-driven redistribution of GRIP1 and activation of AR by GRIP1.