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Biomedical subjects

A C Cato

Publications and source records attributed to A C Cato.

At least 19 recordsLinked to original sources
PubMed2001

Glucocorticoids inhibit MAP kinase via increased expression and decreased degradation of MKP-1.

Glucocorticoids inhibit the proinflammatory activities of transcription factors such as AP-1 and NF-kappa B as well as that of diverse cellular signaling molecules. One of these signaling molecules is the extracellular signal-regulated kinase (Erk-1/2) that controls the release of allergic mediators and the induction of proinflammatory cytokine gene expression in mast cells. The mechanism of inhibition of Erk-1/2 activity by glucocorticoids is unknown. Here we report a novel dual action of glucocorticoids for this inhibition. Glucocorticoids increase the expression of the MAP kinase phosphatase-1 (MKP-1) gene at the promoter level, and attenuate proteasomal degradation of MKP-1, which we report to be triggered by activation of mast cells. Both induction of MKP-1 expression and inhibition of its degradation are necessary for glucocorticoid-mediated inhibition of Erk-1/2 activation. In NIH-3T3 fibroblasts, although glucocorticoids up-regulate the MKP-1 level, they do not attenuate the proteasomal degradation of this protein and consequently they are unable to inhibit Erk-1/2 activity. These results identify MKP-1 as essential for glucocorticoid-mediated control of Erk-1/2 activation and unravel a novel regulatory mechanism for this anti-inflammatory drug.

Animals↗
PubMed2001

BAG-1 family of cochaperones in the modulation of nuclear receptor action.

BAG-1 is a family of cochaperones consisting of at least four polypeptides BAG-1L, BAG-1M/RAP46, BAG-1 and p29. These proteins are translated from the same mRNA at alternative translation initiation sites. They possess conserved carboxy-terminal sequences which enable them to bind and inhibit the action of the molecular chaperone Hsp70/Hsc70. BAG-1 was the first member in the family of the BAG-1 proteins to be isolated. It was identified as an anti-apoptotic protein because of its ability to bind and augment the activity of the anti-death protein, Bcl-2. Since then other BAG-1 proteins have been identified and shown to interact with several cellular factors including nuclear receptors. Recent findings show that the effect of the BAG-1 proteins on nuclear receptors ranges from inhibition to enhancement of the transactivation functions of the receptors. Available data on the negative regulation of glucocorticoid receptor (GR) action by the BAG-1 proteins identify two modes of action: inhibition of the hormone binding activity of the GR and a more direct nuclear action at the level of regulation of the transactivation function of the receptor. In the latter case, the BAG-1 proteins repress DNA binding by the GR in a process that requires prior binding of Hsp70/Hsc70 to the receptor. Positive regulatory action of the BAG-1 proteins on nuclear receptors has also been reported which may involve yet other mechanisms. This review puts together recent findings on the action the BAG-1 proteins and presents them as a novel group of regulators of action of nuclear receptor.

Animals↗
PubMed2001

Expression level-dependent contribution of glucocorticoid receptor domains for functional interaction with STAT5.

The action of the glucocorticoid receptor (GR) on beta-casein gene transcription serves as a well-studied example of a case where the action of the GR is dependent on the activity of another transcription factor, STAT5. We have investigated the domain-requirement of the GR for this synergistic response in transfection experiments employing GR mutants and CV-1 or COS-7 cells. The results were influenced by the expression levels of the GR constructs. At low expression, STAT5-dependent transactivation by mutants of the GR DNA binding domain or N-terminal transactivation domain was impaired and the antiglucocorticoid RU486 exhibited a weak agonistic activity. When the N-terminal region of the GR was exchanged with the respective domain of the progesterone receptor, STAT5-dependent transactivation was reduced at low and high expression levels. Only at high expression levels did the GR exhibit the properties of a coactivator and enhanced STAT5 activity in the absence of a functional DNA binding domain and of GR binding sites in the proximal region of the beta-casein gene promoter. Furthermore, at high GR expression levels RU486 was nearly as efficient as dexamethasone in activating transcription via the STAT5 dependent beta-casein gene promoter. The results reconcile the controversial issue regarding the DNA binding-independent action of the GR together with STAT5 and provide evidence that the mode of action of the GR depends not only on the type of the particular promoter at which it acts but also on the concentration of the GR. GR DNA binding function appears to be mandatory for beta-casein gene expression in mammary epithelial cells, since the promoter function is completely dependent on the integrity of GR binding sites in the promoter.

Amino Acid Sequence↗
PubMed2001

Alteration of the stability of Bag-1 protein in the control of olfactory neuronal apoptosis.

Normal apoptosis occurs continuously in the olfactory neuroepithelium of adult vertebrates, making it a useful model for studying neuronal apoptosis. Here we demonstrate that overexpression of the anti-apoptotic Bag-1 gene in olfactory neuronal cells confers a strong resistance to apoptosis. Conversely decreased levels of Bag-1 were found to precede a massive wave of olfactory neuronal apoptosis triggered by synaptic target ablation. We show that the decrease is brought about by ubiquitination and subsequent degradation of the Bag-1 protein. The ring finger protein Siah-2 is a likely candidate for the ubiquitination reaction since Siah-2 mRNA accumulated in lesioned olfactory neuroepithelium and overexpression of Siah-2 stimulated Bag-1 ubiquitination and degradation in transient expression assays. These results together identify destabilization of Bag-1 as a necessary step in olfactory neuronal apoptosis.

Amino Acid Sequence↗
PubMed2000

Hsp70-RAP46 interaction in downregulation of DNA binding by glucocorticoid receptor.

Receptor-associating protein 46 (RAP46) is a cochaperone that regulates the transactivation function of several steroid receptors. It is transported into the nucleus by a liganded glucocorticoid receptor where it downregulates DNA binding and transactivation by this receptor. The N- and C-termini of RAP46 are both implicated in its negative regulatory function. In metabolic labelling experiments, we have shown that the N-terminus of RAP46 is modified by phosphorylation, but this does not contribute to the downregulation of glucocorticoid receptor activity. However, deletion of a sequence that binds 70 kDa heat shock protein (Hsp70) and the constitutive isoform of Hsp70 (Hsc70) at the C-terminus of RAP46 abrogated its negative regulatory action. Surface plasmon resonance studies showed that RAP46 binds the glucocorticoid receptor only when it has interacted with Hsp70/Hsc70, and confocal immunofluorescence analyses revealed a nuclear transport of Hsp70/Hsc70 by the liganded receptor. Together these findings demonstrate an important contribution of Hsp70/Hsc70 in the binding of RAP46 to the glucocorticoid receptor and suggest a role for this molecular chaperone in the RAP46-mediated downregulation of glucocorticoid receptor activity.

Amino Acid Sequence↗
PubMed2000

Cytoplasmic localization and the choice of ligand determine aggregate formation by androgen receptor with amplified polyglutamine stretch.

Polyglutamine tract expansion in androgen receptor is a recognized cause of spinal and bulbar muscular atrophy (SBMA), an X-linked motor neuronopathy. Similar mutations have been identified in proteins associated with other neurodegenerative diseases. Recent studies have shown that amplified polyglutamine repeat stretches form cellular aggregates that may be markers for these neurodegenerative diseases. Here we describe conditions that lead to aggregate formation by androgen receptor with polyglutamine stretch amplification. In transfection experiments, the mutant, compared with the wild-type receptor, was delayed in its cytoplasmic-nuclear translocation and formed large cytoplasmic aggregates in the presence of androgen. The cytoplasmic environment appears crucial for this aggregation, since retention of both the wild-type and mutant receptors in this cellular compartment by the deletion of their nuclear localization signals resulted in massive aggregation. Conversely, rapid nuclear transport of both receptors brought about by deletion of their ligand binding domains did not result in aggregate formation. However, androgen antagonists that altered the conformation of the ligand binding domain and promoted varying rates of cytoplasmic-nuclear translocation all inhibited aggregate formation. This demonstrates that in addition to the cytoplasmic localization, a distinct contribution of the ligand binding domain of the receptor is necessary for the aggregation. The finding that antiandrogens inhibit aggregate formation may provide the basis for in vivo determination of the role of these structures in SBMA.

Animals↗
PubMed2000

Isoform-specific expression of BAG-1 in mouse development.

BAG-1 is a family of proteins with diverse activities that range in cultured cells from protection against programmed cell death through to regulation of steroid hormone action. At least three proteins (BAG-1L, BAG-1M and BAG-1) are encoded by the Bag-1 mRNA through the use of alternative translation-initiation sites. To assess the in vivo function of these factors, we have used in situ hybridization and immunohistochemical techniques to determine the distribution of Bag-1 transcript and proteins during mouse development. Bag-1 mRNA was identified in several organs with cartilaginous tissues showing the highest expression levels. The level of expression at some of these sites was downregulated during the course of development. In the immunohistochemical studies, antibodies directed against the BAG-1 proteins stained all the sites identified in the in situ hybridization studies although isoform-specific differences were observed. BAG-1L specific antibody showed ubiquitous staining as early as day 10.5 post-coitum but there was a progressive restriction during subsequent stages of embryogenesis. On the contrary, an antibody that preferentially recognized the other isoforms only stained the mouse myocardium in the early developmental stages before finally recognizing additional organs later on in development. These results demonstrate a stage- and site-specific expression of the BAG-1 isoforms during mouse development.

Amino Acid Sequence↗
PubMed2000

BAG-1M: a potential specificity determinant of corticosteroid receptor action.

BAG-1M is a eukaryotic cochaperone that associates with several proteins, including the glucocorticoid receptor (GR). It down-regulates GR-mediated transactivation by a mechanism that requires its prior recruitment by the liganded receptor from cytoplasm into the nucleus. In the nucleus, it uses a repeated sequence motif ([EEX4]8) at its NH2 terminus to inhibit DNA binding, as well as transactivation functions of the receptor. The mineralocorticoid receptor (MR), a structural and functional homologue of the GR, is unable to translocate BAG-1M into the nucleus, and its transactivation function is also not affected by this protein. This differential regulation of GR and MR activity could be relevant in classic mineralocorticoid tissues such as the kidney in which GR activity needs to be repressed to allow the MR to exert its action. In in situ hybridization studies, we show that BAG-1M is expressed in the kidney. Its expression pattern, especially in the developing kidney, correlated well with that of the GR. We therefore postulate that BAG-1M may be a specificity determinant in GR and MR action, and may feature prominently in the control of GR activity in kidney development.

Aging↗
PubMed1999

Rapid signalling by androgen receptor in prostate cancer cells.

Androgens are important growth regulators in prostate cancer. Their known mode of action in target cells requires binding to a cytoplasmic androgen receptor followed by a nuclear translocation event and modulation of the expression of specific genes. Here, we report another mode of action of this receptor. Treatment of androgen responsive prostate cancer cells with dihydrotestosterone leads to a rapid and reversible activation of mitogen-activated protein kinases MAPKs (also called extracellular signal-regulated kinases or Erks). Transient transfection assays demonstrated that the androgen receptor-mediated activation of MAP kinase results in enhanced activity of the transcription factor Elk-1. This action of the androgen receptor differs from its known transcriptional activity since it is rapid and insensitive to androgen antagonists such as hydroxyflutamide or casodex. Biochemical studies as well as analyses with dominant negative mutants showed the involvement of kinases such as MAPK/Erk kinase, phosphatidyl-inositol 3-kinase and protein kinase C in the androgen receptor-mediated activation of MAP kinase. These results demonstrate a novel regulatory action of the androgen receptor and prove that in addition to its known transcriptional effects, it also uses non-conventional means to modulate several cellular signalling processes.

Adenocarcinoma↗
PubMed1999

A nuclear action of the eukaryotic cochaperone RAP46 in downregulation of glucocorticoid receptor activity.

RAP46 is a eukaryotic cochaperone that associates with several proteins, including the heat shock protein hsp70/hsc70 and the glucocorticoid receptor (GR). Here we show a downregulation of GR-mediated transactivation by RAP46 via a mechanism independent of a cytoplasmic action of this cochaperone. We demonstrate a specific cytoplasmic-nuclear recruitment of RAP46 by the liganded GR that results in inhibition of the transactivation function of the receptor. A repeated sequence motif [EEX(4)](8) at the NH(2) terminus of RAP46 or BAG-1L, a larger isoform of RAP46, is responsible for this downregulation of GR activity. BAG-1, a shorter isoform with only a duplication of the [EEX(4)] sequence, does not inhibit GR activity. The [EEX(4)](8) motif, when linked to an otherwise unrelated protein, abrogated the inhibitory action of endogenous RAP46 on GR-mediated transactivation. The nuclear effects of RAP46 and BAG-1L are specific since GR-mediated inhibition of AP-1 activity was not affected. These studies identify the [EEX(4)](8) sequence as a signature motif for inhibition of GR-mediated transactivation and demonstrate a specific nuclear action of a eukaryotic cochaperone in the regulation of GR activity.

Amino Acid Sequence↗
PubMed1999

The interaction between the forkhead thyroid transcription factor TTF-2 and the constitutive factor CTF/NF-1 is required for efficient hormonal regulation of the thyroperoxidase gene transcription.

The forkhead thyroid-specific transcription factor TTF-2 is the main mediator of thyrotropin and insulin regulation of thyroperoxidase (TPO) gene expression. This function depends on multimerization and specific orientation of its DNA-binding site, suggesting that TTF-2 is part of a complex interaction network within the TPO promoter. This was confirmed by transfection experiments and by protein-DNA interaction studies, which demonstrated that CTF/NF1 proteins bind 10 base pairs upstream of the TTF-2-binding site to enhance its action in hormone-induced expression of the TPO gene. GST pull-down assays showed that TTF-2 physically interacts with CTF/NF1 proteins. In addition, we demonstrate that increasing the distance between both transcription factors binding sites by base pair insertion results in loss of promoter activity and in a drastic decrease on the ability of the promoter to respond to the hormones. CTF/NF1 is a family of transcription factors that contributes to constitutive and cell-type specific gene expression. Originally identified as factors implicated in the replication of adenovirus, this group of proteins (CTF/NF1-A, -B, -C, and -X) is now known to be involved in the regulation of several genes. In contrast to other reports regarding the involvement of these proteins in inducible gene expression, we show here that members of this family of transcription factors are regulated by hormones. With the use of specific CTF/NF1 DNA probes and antibodies we demonstrate that CTF/NF1-C is a thyrotropin-, cAMP-, and insulin-inducible protein. Thus CTF/NF1 proteins do not only mediate hormone-induced gene expression cooperating with TTF-2, but are themselves hormonally regulated. All these findings are clearly of important value in understanding the mechanisms governing the transcription regulation of RNA polymerase II promoters, which often contain binding sites for multiple transcription factors.

Base Sequence↗
PubMed1999

Regulation of E-cadherin gene expression during tumor progression: the role of a new Ets-binding site and the E-pal element.

A new regulatory region (-108 to -86), named CE, containing potential CRE- and Ets-binding sites has been identified in the murine E-cadherin promoter. The Ets-binding site (at -97 position) negatively modulates the activity of the E-cadherin promoter in expressing keratinocyte cell lines and was responsible for the specific retarded complexes obtained with the CE region. Analysis of the methylation status of the endogenous E-cadherin promoter indicated that silencing of E-cadherin expression in malignant keratinocytes cannot be explained by hypermethylation mechanisms. Furthermore, treatment with 5'-aza-2'-deoxycytidine was unable to induce the expression of E-cadherin in deficient keratinocytes. However, in vivo footprinting analysis of the endogenous E-cadherin promoter showed a very distinct pattern in expressing and nonexpressing keratinocytes. Extensive interactions in the previously postulated proximal regulatory elements and in the CE region were detected in expressing cells, while only some nucleotides of the E-pal element and of the CE region were protected in nonexpressing keratinocytes. These results indicate a complex regulation of the mouse E-cadherin promoter and support a model where the combination of positive (CCAAT-box and GC-rich region) and negative (E-pal element and CE region) cis-acting elements contribute to the final level of E-cadherin gene expression. In addition, our results show that downregulation of E-cadherin expression in transformed epidermal keratinocytes is mainly exerted through the interaction of repressor factor(s) with the E-pal element and to the lack of interaction of positive acting factors with the proximal regions.

3T3 Cells↗
PubMed1999

Mechanism of androgen receptor activation and possible implications for chemoprevention trials.

Androgens are pivotal regulators of prostate cell growth, differentiation and function, and their actions are believed to be involved in prostate cancer development. The androgen-signaling pathway in the prostate gland is therefore one of the possible sites of intervention in prostate cancer prevention efforts. The central element of androgen signaling in the cell is the androgen receptor (AR), a member of the superfamily of nuclear receptors. Binding of androgen to its ligand-binding domain transforms the receptor to an active transcription factor that regulates gene expression by interacting with specific regulatory elements in the promoters of genes. In addition to this genomic action, the AR also interacts with other signaling pathways through protein-protein interaction, for example with AP-1 or Ets transcription factors. It is not only the action of androgenic hormones, but also the interactions with growth factor and protein kinase A-signaling pathways that can induce activation of AR. Moreover, these ligand-independent activators act synergistically together with low concentrations of androgens. The effects of long-term androgen deprivation on androgen signaling have been investigated in the LNCaP cell culture system. Long-term culture in a steroid-free medium results in a subline showing a hyperreactive AR characterized by increased AR expression and enhanced AR transcriptional activity in an environment with low levels of androgen hormones. It is not yet clear if similar changes also occur in normal or premalignant prostate epithelial cells and are thus relevant for prevention trials which interfere with androgen hormone signaling.

Antineoplastic Agents↗
PubMed1998

RAP46 is a negative regulator of glucocorticoid receptor action and hormone-induced apoptosis.

RAP46 was first identified by its ability to bind the glucocorticoid receptor. It has since been reported to bind several cellular proteins, including the anti-apoptotic protein Bcl-2, but the biological significance of these interactions is unknown. Here we show that RAP46 binds the hinge region of the glucocorticoid receptor and inhibits DNA binding and transactivation by the receptor. We further show that overexpression of RAP46 in mouse thymoma S49.1 cells inhibits glucocorticoid-induced apoptosis. Conversely, glucocorticoid-induced apoptosis and transactivation were enhanced after treating S49.1 cells with the immunosuppressant rapamycin, which down-regulates cellular levels of BAG-1, the mouse homolog of RAP46. The effect of rapamycin can, however, be overcome by overexpression of RAP46. These results together identify RAP46 as a protein that controls glucocorticoid-induced apoptosis through its negative regulatory action on the transactivation property of the glucocorticoid receptor.

Animals↗
PubMed1998

Expression, structure, and function of androgen receptor in advanced prostatic carcinoma.

BACKGROUND: Endocrine therapy for prostate cancer aims to reduce the levels of circulating androgen or to inhibit androgen action by blocking the androgen receptor in the prostate, or both. Studies in various animal and human prostate cancer models suggested that there may be a downregulation of androgen receptor during prostate cancer progression. Recent work, however, showed androgen receptor expression in all stages of prostate cancer. The presence of mutant androgen receptors in a portion of prostate cancers and receptor activation in the absence of androgen or in the presence of low androgen concentrations is discussed within this context. METHODS: This review attempts to summarize the literature on androgen receptor expression in vitro and in vivo, as well as structural and functional alterations and communication between androgen signal transduction cascade and other signaling pathways. CONCLUSIONS: Prostate tumors adapt to an environment with low androgen supply by using a hyperactive androgen receptor. The mechanisms involved are mutations of the androgen receptor generating receptors with broadened activation spectrum, increased receptor expression, and activation by interaction with other signaling pathways.

Androgen Antagonists↗
PubMed1998

Recent developments in molecular action of antihormones.

Antihormones are by definition antagonists of steroid hormone action. They interact with the ligand binding domains of steroid hormone receptors and competitively inhibit the action of the receptors by mechanisms that are not quite understood. In certain cases antihormones also exhibit agonistic activity especially in connection with certain naturally occurring receptor mutants. These observations together with findings of indiscriminate interaction of antihormones with several classes of steroid receptors have necessitated a search of more effective and reliable antihormones. Recent advances in the resolution of the crystal structure of the ligand binding domains of certain members of the steroid receptor family and identification of non-liganded activation of steroid receptors have produced considerable information that can be harnessed into a fruitful search for a new generation of antihormones.

Animals↗
PubMed1998

Expression of androgen receptor mRNA during mouse embryogenesis.

Androgen receptor (AR) is a member of the nuclear receptor superfamily which acts as a ligand-dependent transcription factor (Beato, M., Herrlich, P., Schütz, 1989. Steroid hormone receptors: many actors in search of a plot. Cell 83, 851-857). It plays a pivotal role in sexual development and reproduction (Wilson, J.D., Griffin, J.E., George, F.W., Leshin, M., 1981. The role of gonadal steroids in sexual differentiation. Rec. Prog. Horm. Res. 37, 1-39; Jost, A., 1990. Hormonal control of the masculinization of the body. In: Baulieu, E.E., Kelly, D.A., (Eds.), Hormones, from Molecules to Disease. Chapman and Hall, New York and London, pp. 439-442.). Mutations in the AR sequence cause a number of physiological disorders, such as partial and complete androgen insensitivity syndromes, that lead to abnormal sexual development (Patterson, M.N., McPhaul, M.J., Hughes, I.A., 1994. Androgen insensitivity syndrome. Ballière's Clin. Endocrinol. Metab. 8, 379-404.). There are indications that AR may also have other functions. For example, structural alterations of the AR sequence have been implicated in prostate cancer (Visakorpi, T., Huytinen, E., Koivisto, P., Tanner, M., Keinänen, R., Palmberg, C., Palotie, A., Tammela, T., Isola, J., Kallioniemi, O.-P., 1995. In vivo amplification of the androgen receptor gene and progression of human prostate cancer. Nature Genet. 9, 401-406.) and in the development of spinal and bulbar muscular atrophy, a neurodegenerative disease (Kennedy, W.R., Alter, M., Sung, J.H., 1968. Progressive proximal spinal and bulbar muscular atrophy of late onset: a sex-linked recessive trait. Neurology 18, 671-680.). Here, we have investigated the spatial and temporal expression of AR during mouse organogenesis by in situ hybridisation. We demonstrate that AR transcripts occur in the developing external genitalia, pituitary, adrenals, kidneys and musculus levator ani, in addition to the known expression sites in the Wolffian ducts and its derivatives and during development of the mammary glands.

Animals↗