scholarly journals Cloning of oestrogen receptor beta from Old and New World primates: identification of splice variants and functional analysis

2004 ◽  
Vol 32 (3) ◽  
pp. 703-718 ◽  
Author(s):  
JE Sierens ◽  
GA Scobie ◽  
J Wilson ◽  
PT Saunders
Keyword(s):  
Splice Variant ◽  
Oestrogen Receptor ◽  
New World ◽  
Stop Codon ◽  
Full Length ◽  
Peptide Sequence ◽  
Wild Type ◽  
New World Primates ◽  
World Primate ◽  
The Impact

Oestrogens have a major impact on reproductive function in both males and females. Two oestrogen receptor genes known as ERalpha (ESR1NR3A1) and ERbeta (ESR2NR3A2) have been cloned. Splice variant isoforms of the ERbeta gene have been identified in human, bovine and rodents and it has been suggested that the existence of these forms can influence oestrogen responsiveness. In the human, splicing of an alternative eighth exon results in the formation of a C-terminal variant called hERbetacx, or hERbeta2, but this isoform has not been identified in other species. The aim of the present study was to clone ERbeta cDNAs from primates so as to determine how closely they resembled the ERbeta isoforms found in the human. The two species studied were the stump-tailed macaque (Macaca arctoides), an Old World primate, and the common marmoset (Callithrix jacchus jacchus), a New World primate. Full length ERbeta (wild type, ERbeta1) cDNAs were cloned from macaque and marmoset; they encoded proteins of similar size to those found in human (59 and 54 kDa, long and short forms respectively) and shared significant sequence homology (97.5% in macaque and 93.8% in marmoset) with the human peptide sequence. Full length cDNAs homologous to the hERbeta2 variant were identified in both primates. Marmoset ERbeta2 was slightly shorter than that of human ERbeta2 (54 kDa compared with 55 kDa) and did not contain the peptide sequence used to raise an anti-hERbeta2 antibody. All the macaque ERbeta2 cDNAs contained 56 bp of intronic sequence which included an in-frame stop codon resulting in translation of a truncated protein ( approximately 35 kDa). In all three species, truncated, alternatively spliced mRNAs lacking exon 5 were isolated on multiple occasions from all tissue extracts. In transient transfection assays, ERbeta2-containing constructs were unable to induce transcription of an oestrogen response element (ERE) reporter plasmid in the presence of oestradiol. ERbeta1 from human, macaque and marmoset exhibited minor differences in their ability to induce transcription of the ERE reporter when incubated with different ligands (oestradiol, PPT, DPN, 5-alpha-androstane-3-beta, 17beta-diol (3betaAdiol), genistein) and this may be due to amino acid substitutions within their ligand binding domains. In conclusion, we have identified and cloned wild type ERbeta (ERbeta1) from macaque and marmoset and demonstrated that splice variant mRNAs homologous to hERbeta2 are formed in both species. The marmoset monkey, therefore, provides a suitable animal model in which to investigate the impact of ERbeta variant expression on tissue responsiveness to oestrogens.

The impact of TAS3681, a new type of androgen receptor antagonist, on aberrant AR signaling that drives tumor resistance to AR-targeted therapies by down-regulating full length and splice variant AR.

2017 ◽  
Vol 35 (6_suppl) ◽  
pp. 197-197 ◽  
Author(s):  
Daisuke Kajiwara ◽  
Kazuhisa Minamiguchi ◽  
Masanao Seki ◽  
Hiroya Mizutani ◽  
Keisuke Yamamura ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Splice Variant ◽  
Full Length ◽  
Antitumor Efficacy ◽  
Tumor Xenograft ◽  
Castration Resistant Prostate Cancer ◽  
Tumor Resistance ◽  
Down Regulation ◽  
The Impact

197 Background: The treatment of castration-resistant prostate cancer (CRPC) has changed dramatically with the use of two new therapies, enzalutamide and abiraterone, directed at the androgen receptor (AR) signaling axis. However, eventually almost all patients relapse after these agents because of acquired resistance by a variety of mechanisms such as AR overexpression, the occurrence of AR mutations, or the induction of AR splice variants that confer ligand independent AR transactivation. TAS3681 is an oral pure AR antagonist with AR down-regulating activity. In this study, we investigated the effect of TAS3681 on several issues related to resistance to current AR-targeted therapy. Methods: VCaP cells transfected with wild-type AR-expressing vector, which overexpress AR, were treated with TAS3681 in the presence of androgen. Cells were harvested and luciferase activity was determined. For the analysis of F876L mutant AR, LNCaP cells stably expressing F876L AR protein were incubated with TAS3681 in the presence of androgen. The WST-8 assay was performed to measure cell viability. Prostate cancer cells expressing AR-v7 were treated with TAS3681, and full length (FL)-AR and AR-v7 protein levels were determined by Western blot. TAS3681 was orally dosed to confirm AR down-regulation in tumor and evaluate antitumor efficacy in AR-v7 positive CRPC tumor xenograft mouse model. Results: In prostate cancer (PCa) cells which overexpress AR, in contrast to enzalutamide, TAS3681 effectively suppressed AR transactivation and cell proliferation via AR down-regulation. TAS3681 effectively antagonized F876L mutant AR which confers resistance to cell growth inhibition by enzalutamide. Moreover, TAS3681 reduced both FL-AR and AR-v7 protein level in PCa cells in vitro and in vivo, and showed strong antitumor efficacy in AR-v7 positive xenografts. Conclusions: TAS3681 disrupts the aberrant AR signaling via a novel mechanism of action: pure AR antagonism plus down-regulation of full-length and splice variant AR. These findings suggest that TAS3681 has the potential to overcome resistance to current AR pathway target drugs.

scholarly journals New Insights into Mechanisms of Erythropoietin Receptor Mutations in Primary Familial and Congenital Polycythemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 631-631 ◽  
Author(s):  
Florence Pasquier ◽  
Caroline Marty ◽  
Frédérique Verdier ◽  
Sarah Grosjean ◽  
Claude Préhu ◽  
...  
Keyword(s):  
Cell Surface ◽  
Conflicts Of Interest ◽  
Stop Codon ◽  
Cell Mass ◽  
Erythropoietin Receptor ◽  
Functional Study ◽  
Common Mechanism ◽  
Wild Type ◽  
Terminal Part ◽  
The Impact

Abstract Primary Familial and Congenital Polycythemia (PFCP) is a non-malignant pathology of the erythroid lineage, characterized by an isolated increase of the red cell mass without evolution into myelofibrosis or acutisation. Around twenty constitutive non-sense and missense mutations located in the exon 8 of the erythropoietin receptor (EPOR) gene have been described so far. They all lead to the truncation of the C-terminal part of the protein and the loss of several cytoplasmic tyrosines. The erythropoietin (EPO) hypersensitivity of the PFCP erythroid progenitors is usually explained by the disappearance of these negative signaling regulation and internalization domains (Figure 1a). Nonetheless, relatively few functional studies have been carried out. We therefore investigated the mechanism of EPOR mutations in PFCP. We identified and extensively studied a new constitutive heterozygous frameshift EPOR mutation, p.Gln434Profs*11, which generates a new 11 amino acid (AA) C-terminal tail and a STOP codon at position 444, leading to the truncation of 63 AA of the wild-type receptor (Figure 1c). The primary progenitor cells displayed a major hypersensitivity to EPO, similar to Polycythemia Vera (PV) patients, as well as a spontaneous and persistent JAK2 and STAT5 phosphorylation, compared to the control cells. To study the mechanism of this new EPOR mutant, Ba/F3 cells were transduced with different retroviruses encoding either the HA-tagged wild-type EPOR (EPOR WT)or a truncated receptor at position 444, p.Gln444* (EPOR STOP) or the frameshift EPOR p.Gln434Profs*11mutation (EPOR FS), identical to the patient's mutation (Figure 2). As observed in primary cells, EPOR FS conferred a spontaneous STAT5 phosphorylation and a 4- to 5-fold EPO hypersensitivity to Ba/F3 cells (IC50 of 0.003 U/mL vs 0.01 U/mL) compared to both EPOR WT and EPOR STOP. As expected, the loss of negative regulatory domains in the C-terminal part of the receptor induced a persistent STAT5 activation in EPOR FS and EPOR STOP Ba/F3 cells. Moreover, EPOR FS was more stable (half-life of 120 minutes vs 60 minutes) and displayed a higher level of localization at the cell surface (more than 2-fold), compared to EPOR WT and EPOR STOP. However, no modification of the EPOR FS internalization pattern was observed during 125I-EPO labeling experiments and cytometry analysis. Furthermore, a dileucine motif, known to be a potential clathrin-dependent endocytosis site, is lost in the new C-terminal tail of EPOR FS mutant, yet its abrogation in EPOR WT and EPOR STOP did not modify the phenotype of Ba/F3 cells. Therefore, unlike previous reports, the major EPO hypersensitivity induced by EPOR p.Gln434Profs*11 cannot be explained by the receptor truncation, but rather by the appearance of a new C-terminal tail that confers spontaneous signaling. We wondered if this model could be extended to other EPOR mutations already described in PFCP (Figures 1a-b and 2). We therefore measured the impact on Ba/F3 cells proliferation of the frameshift EPOR p.Pro438Metfs*6 and its non-sense mutant counterpart, p.Pro443*, which retains the tyrosine at position 426, a binding site for the negative signaling regulators SOCS3 and CIS. EPO-hypersensitivity (4- to 5-fold) was only induced by EPOR p.Pro438Metfs*6, suggesting a common mechanism for the frameshift EPOR mutations. Interestingly, two proximal non-sense mutations, EPOR p.Glu399* and p.Glu425*, lacking 7 of the 8 cytoplasmic tyrosines that compose EPOR negative regulatory and internalization domains, were also able to confer a high EPO hypersensitivity to Ba/F3 cells. To our knowledge this is the first extensive functional study of EPOR mutations in PFCP. We highlighted that this pathology is much more complex than expected, since different mechanisms are involved in the EPO hypersensitivity phenotype, according to the type of EPOR mutation. Indeed, extensive truncations are sufficient by themselves to confer the EPO hypersensitivity phenotype due to the loss of all negative regulatory and internalization domains, whereas more distal truncations induced by frameshift mutants confer EPO hypersensitivity because of the appearance of a new C-terminal tail. The latter, by increasing EPOR stability at the cell surface, may cause pre-activation of both receptor and JAK2, constitutive signaling and hypersensitivity to EPO close to that of JAK2V617F-positive PVs. Disclosures No relevant conflicts of interest to declare.

scholarly journals Novel Simian Homologues of Epstein-Barr Virus

2003 ◽  
Vol 77 (19) ◽  
pp. 10695-10699 ◽  
Author(s):  
Bernhard Ehlers ◽  
Andreas Ochs ◽  
Fabian Leendertz ◽  
Michael Goltz ◽  
Christophe Boesch ◽  
...  
Keyword(s):  
Type Species ◽  
Epstein Barr Virus ◽  
New World ◽  
New World Primates ◽  
Pcr Assay ◽  
Old World ◽  
Barr Virus ◽  
World Primate ◽  
Epstein Barr

ABSTRACT Thirty different lymphocryptoviruses (LCV), 26 of them novel, were detected in primates by a panherpesvirus PCR assay. Nineteen LCV from chimpanzees, bonobos, gorillas, and other Old World primates were closely related to Epstein-Barr virus (EBV), the type species of the genus Lymphocryptovirus. Seven LCV originating from New World primates were related to callitrichine herpesvirus 3 (CalHV-3), the first recognized New World LCV. Importantly, a second LCV from gorillas and three LCV from orangutans and gibbons were only distantly related to EBV and CalHV-3. They were tentatively assigned to a novel genogroup of Old World primate LCV. The work described in the paper may also help identify an as yet unknown human LCV.

scholarly journals Reporting social behaviours of mixed-species troops formed by Callithrix jacchus and Callithrix penicillata (Primate, Callitrichidae)

2014 ◽  
Vol 74 (3) ◽  
pp. 607-611 ◽  
Author(s):  
G Valença-Silva ◽  
FG Maciel ◽  
RL Zaganini ◽  
AS Lucindo ◽  
S Caramaschi ◽  
...  
Keyword(s):  
Social Behaviour ◽  
New World ◽  
Callithrix Jacchus ◽  
The State ◽  
New World Primates ◽  
Mixed Species ◽  
The Social ◽  
Mixed Groups ◽  
The Impact ◽  
Social Behaviours

In New World primates, mixed-species troops have been reported. Here, we analysed the performance of affiliative and agonistic behaviours of Callithrix jacchus and Callithrix penicillata living in mixed groups. For this purpose, we recorded the interaction of the individuals from two groups located in Bauru city, in the state of São Paulo (Brazil). Our data show that in both groups, affiliative behaviours appeared more frequently than agonistic ones. We concluded that there is cohesion inside the mixed-species troops observed. We suggest that a deeper knowledge about the social behaviour of mixed-species troop species certainly may be useful in projects linked with the management of the impact caused by them.

The impact of moving to a novel environment on social networks, activity and wellbeing in two new world primates

2011 ◽  
Vol 73 (8) ◽  
pp. 802-811 ◽  
Author(s):  
V. Dufour ◽  
C. Sueur ◽  
A. Whiten ◽  
H.M. Buchanan-Smith
Keyword(s):  
Social Networks ◽  
New World ◽  
New World Primates ◽  
Novel Environment ◽  
The Impact

scholarly journals Purification and Characterization of a Novel Intracellular 17β-Estradiol Binding Protein in Estrogen-Resistant New World Primate Cells

2003 ◽  
Vol 88 (1) ◽  
pp. 501-504 ◽  
Author(s):  
Hong Chen ◽  
Bing Hu ◽  
Gang-Hua Huang ◽  
Amanda G. Trainor ◽  
David H. Abbott ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
New World ◽  
Binding Protein ◽  
Target Cells ◽  
Target Tissue ◽  
New World Primates ◽  
Old World ◽  
World Primate ◽  
17Β Estradiol

Compared to Old World primates, including man, New World primates display target-tissue resistance to gonadal steroid hormones. In female New World primates this resistant phenotype is characterized by elevated concentrations of plasma estradiol and progesterone. Here we describe the discovery of an intracellular estrogen binding protein (IEBP) that acts to concentrate 17β-estradiol (E2) in the cytoplasm of New World primate target cells. IEBP was purified by E2-affinity chromatography from postnuclear extracts of the B95-8 cells established from an E2-resistant New World primate. Compared with unpurified extract, affinity-purified IEBP demonstrated a 300-fold enrichment in specific E2 binding activity; half-maximal displacement of[ 3H]E2 from affinity-purified IEBP was observed with 0.1 nM E2. Affinity-purified extracts were subjected to SDS-PAGE with isolation of a dominant 27-28 kDa protein. N-terminal sequencing of tryptic peptides of the protein showed sequence homology with human heat shock protein-27 (hsp27). By immunoblot and E2 binding capacity, IEBP was 1] 2–3-fold greater in New World than in Old World primate tissues and cell lines, 2] heat-inducible and 3] up-regulated in vivo in the presence of the functioning female gonad. In conclusion, IEBP is a specific E2-interacting heat shock protein in the hsp-27 family that is relatively overexpressed in estrogen-resistant cells.

scholarly journals Reduced Prevalence of Epstein-Barr Virus-Related Lymphocryptovirus Infection in Sera from a New World Primate

2005 ◽  
Vol 79 (15) ◽  
pp. 10069-10072 ◽  
Author(s):  
Mark H. Fogg ◽  
Angela Carville ◽  
Jennifer Cameron ◽  
Carol Quink ◽  
Fred Wang
Keyword(s):  
Epstein Barr Virus ◽  
New World ◽  
Viral Gene ◽  
Gene Repertoire ◽  
New World Primates ◽  
Common Marmosets ◽  
Old World ◽  
Barr Virus ◽  
World Primate ◽  
Epstein Barr

ABSTRACT The recent discovery of an Epstein-Barr virus (EBV)-related lymphocryptovirus (LCV) naturally infecting common marmosets demonstrated that gamma-1 herpesviruses are not limited to human and Old World nonhuman primate hosts. We developed serologic assays to detect serum antibodies against lytic- and latent-infection marmoset LCV antigens in order to perform the first seroepidemiologic study of LCV infection in New World primates. In three different domestic colonies and in animals recently captured from the wild, we found that the seroprevalence of marmoset LCV infection was not as ubiquitous as with EBV or Old World LCV. These biologic differences in LCV infection of New World versus human and Old World primate hosts correlate with the evolution of the LCV viral gene repertoire.

Subtle Impact of Akt1 and Akt3 on Exploratory Behavior in Gene Targeted Mice

2015 ◽  
Vol 223 (3) ◽  
pp. 173-180 ◽  
Author(s):  
Christina Leibrock ◽  
Michael Hierlmeier ◽  
Undine E. Lang ◽  
Florian Lang
Keyword(s):  
Forced Swimming Test ◽  
Open Field Test ◽  
Wild Type ◽  
Average Speed ◽  
Closed Area ◽  
Light Area ◽  
Swimming Test ◽  
The Impact ◽  
Center Area ◽  
Dark Transition

Abstract. The present study explored the impact of Akt1 and Akt3 on behavior. Akt1 (akt1-/-) and Akt3 (akt3-/-) knockout mice were compared to wild type (wt) mice. The akt1-/- mice, akt3-/- mice, and wt mice were similar in most parameters of the open-field test. However, the distance traveled in the center area was slightly but significantly less in akt3-/- mice than in wt mice. In the light/dark transition test akt1-/- mice had significantly lower values than wt mice and akt3-/- mice for distance traveled, number of rearings, rearing time in the light area, as well as time spent and distance traveled in the entrance area. They were significantly different from akt3-/- mice in the distance traveled, visits, number of rearings, rearing time in the light area, as well as time spent, distance traveled, number of rearings, and rearing time in the entrance area. In the O-maze the time spent, and the visits to open arms, as well as the number of protected and unprotected headdips were significantly less in akt1-/- mice than in wt mice, whereas the time spent in closed arms was significantly more in akt1-/- mice than in wt mice. Protected and unprotected headdips were significantly less in akt3-/- mice than in wt mice. In closed area, akt3-/- mice traveled a significantly larger distance at larger average speed than akt1-/- mice. No differences were observed between akt1-/- mice, akt3-/- mice and wt-type mice in the time of floating during the forced swimming test. In conclusion, akt1-/- mice and less so akt3-/ mice display subtle changes in behavior.

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