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1.
Genesis ; 57(11-12): e23335, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31513344

RESUMO

Dead-End 1 (DND1) encodes an RNA binding protein critical for viable primordial germ cells in vertebrates. When introduced into cancer cell lines, DND1 suppresses cell proliferation and enhances apoptosis. However, the molecular function of mammalian wild-type DND1 has mostly been studied in cell lines and not verified in the organism. To facilitate study of wild-type DND1 function in mammalian systems, we generated a novel transgenic mouse line, LSL-FM-DND1 flox/+ , which conditionally expresses genetically engineered, FLAG-tagged and myc-tagged DND1 in a cell type-specific manner. We report that FLAG-myc-DND1 is indeed expressed in specific tissues of the mouse when LSL-FM-DND1 flox/+ is combined with mouse strains expressing Cre-recombinase. LSL-FM-DND1 flox/+ mice are fertile with no overt health effects. We expressed FLAG-myc-DND1 in the pancreas and found that chronic, ectopic expression of FLAG-myc-DND1 led to increase in fasting glucose levels in older mice. Thus, this novel LSL-FM-DND1 flox/+ mouse strain will facilitate studies on the biological and molecular function of wild-type DND1.

2.
BMC Genet ; 15: 65, 2014 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-24886204

RESUMO

BACKGROUND: Complex genetic factors underlie testicular germ cell tumor (TGCT) development. One experimental approach to dissect the genetics of TGCT predisposition is to use chromosome substitution strains, such as the 129.MOLF-Chr 19 (M19). M19 carries chromosome (Chr) 19 from the MOLF whereas all other chromosomes are from the 129 strain. 71% of M19 males develop TGCTs in contrast to 5% in 129 strain. To identify and map tumor loci from M19 we generated congenic strains harboring MOLF chromosome 19 segments on 129 strain background and monitored their TGCT incidence. RESULTS: We found 3 congenic strains that each harbored tumor promoting loci that had high (14%-32%) whereas 2 other congenics had low (4%) TGCT incidences. To determine how multiple loci influence TGCT development, we created double and triple congenic strains. We found additive interactions were predominant when 2 loci were combined in double congenic strains. Surprisingly, we found an example where 2 loci, both which do not contribute significantly to TGCT, when combined in a double congenic strain resulted in greater than expected TGCT incidence (positive interaction). In an opposite example, when 2 loci with high TGCT incidences were combined, males of the double congenic showed lower than expected TGCT incidence (negative interaction). For the triple congenic strain, depending on the analysis, the overall TGCT incidence could be additive or could also be due to a positive interaction of one region with others. Additionally, we identified loci that promote bilateral tumors or testicular abnormalities. CONCLUSIONS: The congenic strains each with their characteristic TGCT incidences, laterality of tumors and incidence of testicular abnormalities, are useful for identification of TGCT susceptibility modifier genes that map to Chr 19 and also for studies on the genetic and environmental causes of TGCT development. TGCTs are a consequence of aberrant germ cell and testis development. By defining predisposing loci and some of the locus interactions from M19, this study further advances our understanding of the complex genetics of TGCTs, which is the most common cancer in young human males.


Assuntos
Loci Gênicos , Predisposição Genética para Doença , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Testiculares/genética , Animais , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Congênicos
3.
PLoS One ; 9(4): e93311, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24691397

RESUMO

Exposure to radiation during fetal development induces testicular germ cell tumors (TGCT) and reduces spermatogenesis in mice. However, whether DNA damaging chemotherapeutic agents elicit these effects in mice remains unclear. Among such agents, cyclophosphamide (CP) is currently used to treat breast cancer in pregnant women, and the effects of fetal exposure to this drug manifested in the offspring must be better understood to offer such patients suitable counseling. The present study was designed to determine whether fetal exposure to CP induces testicular cancer and/or gonadal toxicity in 129 and in 129.MOLF congenic (L1) mice. Exposure to CP on embryonic days 10.5 and 11.5 dramatically increased TGCT incidence to 28% in offspring of 129 mice (control value, 2%) and to 80% in the male offspring of L1 (control value 33%). These increases are similar to those observed in both lines of mice by radiation. In utero exposure to CP also significantly reduced testis weights at 4 weeks of age to ∼ 70% of control and induced atrophic seminiferous tubules in ∼ 30% of the testes. When the in utero CP-exposed 129 mice reached adulthood, there were significant reductions in testicular and epididymal sperm counts to 62% and 70%, respectively, of controls. In female offspring, CP caused the loss of 77% of primordial follicles and increased follicle growth activation. The results indicate that i) DNA damage is a common mechanism leading to induction of testicular cancer, ii) increased induction of testis cancer by external agents is proportional to the spontaneous incidence due to inherent genetic susceptibility, and iii) children exposed to radiation or DNA damaging chemotherapeutic agents in utero may have increased risks of developing testis cancer and having reduced spermatogenic potential or diminished reproductive lifespan.


Assuntos
Antineoplásicos Alquilantes/efeitos adversos , Ciclofosfamida/efeitos adversos , Exposição Materna , Folículo Ovariano/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal , Espermatogênese/efeitos dos fármacos , Neoplasias Testiculares/etiologia , Animais , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Folículo Ovariano/efeitos da radiação , Ovário/efeitos dos fármacos , Ovário/metabolismo , Ovário/patologia , Ovário/efeitos da radiação , Gravidez , Contagem de Espermatozoides , Espermatogênese/efeitos da radiação , Testículo/efeitos dos fármacos , Testículo/metabolismo , Testículo/patologia , Testículo/efeitos da radiação
4.
Can J Urol ; 20(4): 6860-7, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23930614

RESUMO

INTRODUCTION: A small subset of young men die from seminoma. Studying these high risk, clinically atypical seminomas (CASs)-aggressive tumors with visceral metastases and chemotherapy resistance-may provide clues to the nature of drug resistance and the origin of testicular cancers. We explored the possibility that these seminomas are a unique clinical and biologic entity with intrinsic yolk sac tumor (YST) features. MATERIALS AND METHODS: We assayed available archived tissue samples (n = 22) for chemotherapy-resistance markers found in YSTs. Specifically, we analyzed tissues and clinical histories from patients with CASs (those who had visceral metastases and recurrent disease), classical seminomas, and mixed germ-cell tumors containing YST. By using immunohistochemical testing, we evaluated the expression of bone morphogenetic protein 2, alpha fetoprotein, and glutathione S-transferase (pi) [GST (pi)]. RESULTS: GST (pi) expression significantly predicted for overall survival (p = .036). In addition, according to the results of GST (pi) immunohistochemical staining, the CASs appeared to resemble YSTs more than they did classical seminomas (p = 0.043). Less-advanced tumors, both those that expressed GST (pi) and those that were negative for GST (pi), were more amenable to local therapies, and the patients who had those tumors had better clinical outcomes. CONCLUSIONS: Results from this exploratory study suggest that certain CASs that express GST (pi) are more similar to YST than they are to classical seminomas, and that GST (pi) expression may be able to be used as a prognosticator of disease-specific survival. Such CASs thus may have a unique biologic origin that differs from that of classical seminomas. Additional studies are needed to determine the natural history and therapeutic implications of these CASs.


Assuntos
Biomarcadores Tumorais/metabolismo , Tumor do Seio Endodérmico/metabolismo , Seminoma/metabolismo , Neoplasias Testiculares/metabolismo , Proteína Morfogenética Óssea 2/metabolismo , Tumor do Seio Endodérmico/diagnóstico , Tumor do Seio Endodérmico/patologia , Glutationa Transferase/metabolismo , Humanos , Imuno-Histoquímica/métodos , Masculino , Prognóstico , Estudos Retrospectivos , Seminoma/diagnóstico , Seminoma/patologia , Neoplasias Testiculares/diagnóstico , Neoplasias Testiculares/patologia , alfa-Fetoproteínas/metabolismo
5.
BMC Mol Biol ; 14: 16, 2013 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-23890083

RESUMO

The RNA binding protein DEAD-END (DND1) is one of the few proteins known to regulate microRNA (miRNA) activity at the level of miRNA-mRNA interaction. DND1 blocks miRNA interaction with the 3'-untranslated region (3'-UTR) of specific mRNAs and restores protein expression. Previously, we showed that the DNA cytosine deaminase, APOBEC3 (apolipoprotein B mRNA-editing enzyme, catalytic polypeptide like 3), interacts with DND1. APOBEC3 has been primarily studied for its role in restricting and inactivating retroviruses and retroelements. In this report, we examine the significance of DND1-APOBEC3 interaction. We found that while human DND1 inhibits miRNA-mediated inhibition of P27, human APOBEC3G is able to counteract this repression and restore miRNA activity. APOBEC3G, by itself, does not affect the 3'-UTR of P27. We found that APOBEC3G also blocks DND1 function to restore miR-372 and miR-206 inhibition through the 3'-UTRs of LATS2 and CX43, respectively. In corollary experiments, we tested whether DND1 affects the viral restriction function or mutator activity of APOBEC3. We found that DND1 does not affect APOBEC3 inhibition of infectivity of exogenous retrovirus HIV (ΔVif) or retrotransposition of MusD. In addition, examination of Ter/Ter;Apobec3-/- mice, lead us to conclude that DND1 does not regulate the mutator activity of APOBEC3 in germ cells. In summary, our results show that APOBEC3 is able to modulate DND1 function to regulate miRNA mediated translational regulation in cells but DND1 does not affect known APOBEC3 function.


Assuntos
Citidina Desaminase/metabolismo , Regulação para Baixo , MicroRNAs/metabolismo , Proteínas de Neoplasias/metabolismo , Regiões 3' não Traduzidas , Desaminase APOBEC-3G , Animais , Inibidor de Quinase Dependente de Ciclina p27/genética , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Citidina Desaminase/genética , Regulação da Expressão Gênica , Humanos , Camundongos , Camundongos Knockout , MicroRNAs/genética , Proteínas de Neoplasias/genética , Ligação Proteica
6.
PLoS One ; 7(2): e32064, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22348147

RESUMO

The prevalence of testicular germ cell tumors (TGCT), a common solid tissue malignancy in young men, has been annually increasing at an alarming rate of 3%. Since the majority of testicular cancers are derived from germ cells at the stage of transformation of primordial germ cell (PGC) into gonocytes, the increase has been attributed to maternal/fetal exposures to environmental factors. We examined the effects of an estrogen (diethylstilbestrol, DES), an antiandrogen (flutamide), or radiation on the incidence of testicular germ cell tumors in genetically predisposed 129.MOLF-L1 (L1) congenic mice by exposing them to these agents on days 10.5 and 11.5 of pregnancy. Neither flutamide nor DES produced noticeable increases in testis cancer incidence at 4 weeks of age. In contrast, two doses of 0.8-Gy radiation increased the incidence of TGCT from 45% to 100% in the offspring. The percentage of mice with bilateral tumors, weights of testes with TGCT, and the percentage of tumors that were clearly teratomas were higher in the irradiated mice than in controls, indicating that irradiation induced more aggressive tumors and/or more foci of initiation sites in each testis. This radiation dose did not disrupt spermatogenesis, which was qualitatively normal in tumor-free testes although they were reduced in size. This is the first proof of induction of testicular cancer by an environmental agent and suggests that the male fetus of women exposed to radiation at about 5-6 weeks of pregnancy might have an increased risk of developing testicular cancer. Furthermore, it provides a novel tool for studying the molecular and cellular events of testicular cancer pathogenesis.


Assuntos
Feto/efeitos da radiação , Efeitos Tardios da Exposição Pré-Natal , Neoplasias Testiculares/etiologia , Antagonistas de Androgênios/toxicidade , Animais , Dietilestilbestrol/toxicidade , Estrogênios não Esteroides/toxicidade , Feminino , Flutamida/toxicidade , Predisposição Genética para Doença , Masculino , Exposição Materna , Camundongos , Gravidez , Efeitos Tardios da Exposição Pré-Natal/genética
7.
Urol Oncol ; 30(4): 494-501, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-20822932

RESUMO

OBJECTIVES: Certain patients with seminoma and clinically atypical phenotypes--visceral metastases, elevated levels of ß human chorionic gonadotropin (ßHCG), and/or recurrent disease--have a poor prognosis. The primary goal of this pilot study was to characterize the clinical characteristics and treatment profile of these rare patients. We also wished to test whether these tumors expressed any specific biomarkers that might distinguish them as a unique subtype of seminoma. MATERIALS AND METHODS: We retrospectively identified 25 patients with a history of seminoma plus visceral metastases, ßHCG levels >200 mU/ml, and/or recurrent disease. We reviewed these patients' histories for treatment efficacy and clinical outcome. Tissue samples were available from 6 of those patients, and we studied them for expression of the markers OCT 3/4, PLAP, CD30, TRA-1-60, c-kit, and gp200. We compared our results with the expression of those markers in tissue samples from mixed seminoma/embryonal carcinomas and classic seminomas. RESULTS: Our analysis suggested that certain chemotherapeutic regimens (such as ifosfamide, paclitaxel, and cisplatin) are efficacious for the treatment of patients with these atypical seminomas. Further, specimens from the atypical seminomas generally had staining profiles that resembled those of classic seminomas and the seminoma components in mixed germ-cell tumors, but the profiles differed from those of the embryonal carcinoma components in the same mixed germ-cell tumors. CONCLUSIONS: Although these atypical seminomas tend to be resistant to chemotherapy, they may still respond to certain chemotherapeutic regimens. Our pilot immunohistochemical study also suggested that the unique phenotypes associated with these atypical seminomas do not result from any relationship with embryonal carcinomas. More study is needed to confirm these initial findings.


Assuntos
Recidiva Local de Neoplasia , Seminoma/patologia , Neoplasias Testiculares/patologia , Vísceras/patologia , Adulto , Idoso , Antígenos de Superfície/metabolismo , Antineoplásicos/uso terapêutico , Gonadotropina Coriônica Humana Subunidade beta/metabolismo , Humanos , Imuno-Histoquímica , Antígeno Ki-1/metabolismo , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Fator 3 de Transcrição de Octâmero/metabolismo , Projetos Piloto , Proteínas/metabolismo , Proteoglicanas/metabolismo , Proteínas Proto-Oncogênicas c-kit/metabolismo , Estudos Retrospectivos , Seminoma/tratamento farmacológico , Seminoma/metabolismo , Sialoglicoproteínas/metabolismo , Neoplasias Testiculares/tratamento farmacológico , Neoplasias Testiculares/metabolismo , Resultado do Tratamento , Vísceras/metabolismo , Adulto Jovem
8.
BMC Mol Biol ; 12: 37, 2011 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-21851623

RESUMO

BACKGROUND: The RNA binding protein, DEAD END (DND1), is essential for maintaining viable germ cells in vertebrates. It is also a testicular germ cell tumor susceptibility factor in mice. DND1 has been shown to interact with the 3'-untranslated region (3'-UTR) of mRNAs such as P27 and LATS2. Binding of DND1 to the 3'-UTRs of these transcripts blocks the inhibitory function of microRNAs (miRNA) from these transcripts and in this way DND1 helps maintain P27 and LATS2 protein expression. We found that DND1 is also expressed in embryonic stem (ES) cells. Because ES cells share similar gene expression patterns as germ cells, we utilized ES cells to identify additional candidate mRNAs that associate with DND1. RESULTS: ES cells are readily amenable to genetic modification and easier to culture in vitro compared to germ cells. Therefore, for the purpose of our study, we made a genetically modified, stable, human embryonic stem (hES) cell line that expresses hemagluttinin (HA)-tagged DND1 in a doxycycline (dox) regulatable manner. This line expresses modest levels of HA-DND1 and serves as a good system to study DND1 function in vitro. We used this stable cell line to identify the transcripts that physically interact with DND1. By performing ribonucleoprotein immunoprecipitation (RIP) followed by RT-PCR, we identified that transcripts encoding pluripotency factors (OCT4, SOX2, NANOG, LIN28), cell cycle regulators (TP53, LATS2) and apoptotic factors (BCLX, BAX) are specifically associated with the HA-DND1 ribonucleoprotein complex. Surprisingly, in many cases, bioinformatics analysis of the pulled-down transcripts did not reveal the presence of known DND1 interacting motifs. CONCLUSIONS: Our results indicate that the inducible ES cell line system serves as a suitable in vitro system to identify the mRNA targets of DND1. The RIP-RT results hint at the broad spectrum of mRNA targets that interact with DND1 in ES cells. Based on what is known about DND1 function, our results suggest that DND1 may impose another level of translational regulation to modulate expression of critical factors in ES cells.


Assuntos
Células-Tronco Embrionárias/fisiologia , Proteínas de Neoplasias/metabolismo , RNA Mensageiro/metabolismo , Regiões 3' não Traduzidas , Animais , Linhagem Celular , Células-Tronco Embrionárias/citologia , Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos , Proteínas de Neoplasias/genética , RNA Mensageiro/genética , Ribonucleoproteínas/genética , Ribonucleoproteínas/metabolismo , Proteínas Supressoras de Tumor/metabolismo
9.
Cancer Res ; 70(18): 7264-72, 2010 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-20736371

RESUMO

Testicular germ cell tumors (TGCT) originate from germ cells. The 129-Ter and M19 (129.MOLF-Chr19 consomic) mouse strains have extremely high incidences of TGCTs. We found that the expression levels of Sf1-encoded splicing factor 1 (SF1) can modulate the incidence of TGCTs. We generated mice with inactivated Sf1. Sf1 null mice (Sf1-/-) died before birth. Mice with one intact allele of Sf1 (Sf1+/-) were viable but expressed reduced levels of Sf1. When Sf1-deficient mice (Sf1+/-) were crossed to the 129-Ter and M19 strains, we observed decreased incidence of TGCTs in Sf1+/-;Ter and Sf1+/-;M19/+ mice compared with that in control cohorts. Therefore, Sf1 deficiency protects against TGCT development in both strains. Sf1 is expressed in the testes. We found that Sf1 levels vary significantly in the testes of inbred strains such as 129 and MOLF, and as such Sf1 is an oncogenic tumor-susceptibility factor from 129. Our results also highlight the complications involved in evaluating Sf1 levels and TGCT incidences. When a large number of tumor-promoting factors are present in a strain, the protective effect of lower Sf1 levels is masked. However, when the dosage of tumor-promoting factors is reduced, the protective effect of lower Sf1 levels becomes apparent. SF1 is involved in splicing of specific pre-mRNAs in cells. Alternate splicing generates the complex proteosome in eukaryotic cells. Our data indicate that Sf1 levels in mouse strains correlate with their incidences of TGCTs and implicate the importance of splicing mechanisms in germ cell tumorigenesis.


Assuntos
Transformação Celular Neoplásica/genética , Proteínas de Ligação a DNA/deficiência , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Testiculares/genética , Fatores de Transcrição/deficiência , Processamento Alternativo , Animais , Transformação Celular Neoplásica/metabolismo , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/genética , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neoplasias Embrionárias de Células Germinativas/metabolismo , Fatores de Processamento de RNA , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Neoplasias Testiculares/metabolismo , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética
10.
Proc Natl Acad Sci U S A ; 106(52): 22323-8, 2009 Dec 29.
Artigo em Inglês | MEDLINE | ID: mdl-20007774

RESUMO

Dmrt1 (doublesex and mab-3 related transcription factor 1) is a conserved transcriptional regulator of male differentiation required for testicular development in vertebrates. Here, we show that in mice of the 129Sv strain, loss of Dmrt1 causes a high incidence of teratomas, whereas these tumors do not form in Dmrt1 mutant C57BL/6J mice. Conditional gene targeting indicates that Dmrt1 is required in fetal germ cells but not in Sertoli cells to prevent teratoma formation. Mutant 129Sv germ cells undergo apparently normal differentiation up to embryonic day 13.5 (E13.5), but some cells fail to arrest mitosis and ectopically express pluripotency markers. Expression analysis and chromatin immunoprecipitation identified DMRT1 target genes, whose missexpression may underlie teratoma formation. DMRT1 indirectly activates the GDNF coreceptor Ret, and it directly represses the pluripotency regulator Sox2. Analysis of human germ cell tumors reveals similar gene expression changes correlated to DMRT1 levels. Dmrt1 behaves genetically as a dose-sensitive tumor suppressor gene in 129Sv mice, and natural variation in Dmrt1 activity can confer teratoma susceptibility. This work reveals a genetic link between testicular dysgenesis, pluripotency regulation, and teratoma susceptibility that is highly sensitive to genetic background and to gene dosage.


Assuntos
Células-Tronco Fetais/citologia , Células-Tronco Fetais/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Espermatogênese/genética , Espermatogênese/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/fisiologia , Animais , Diferenciação Celular , Proliferação de Células , Dosagem de Genes , Expressão Gênica , Genes Supressores de Tumor , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Embrionárias de Células Germinativas/metabolismo , Neoplasias Embrionárias de Células Germinativas/patologia , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Fenótipo , Proteínas Proto-Oncogênicas c-ret/genética , Proteínas Proto-Oncogênicas c-ret/metabolismo , Teratoma/genética , Teratoma/metabolismo , Teratoma/patologia , Neoplasias Testiculares/genética , Neoplasias Testiculares/metabolismo , Neoplasias Testiculares/patologia
11.
Genesis ; 47(9): 617-27, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19548313

RESUMO

Germ cells are essential for the propagation of individual species. Studies on germ cell development in mice highlight important biological paradigms. Beginning with their first appearance around embryonic day 7 (E7), germ cells undergo specific cellular changes at different stages of their embryonic and adult development. Germ cells migrate through the hind-regions of the embryo to eventually home into the developing gonads. Further differentiation and development of germ cells differ in males and females. The processes involved in germ cell development and their eventual differentiation into sperm and oocytes have been under extensive investigation in recent years. Studies on germ cells have shed light on the cellular and molecular processes involved in their specification, migration, proliferation, death, and differentiation. These studies have also revealed much about maintenance of stem cell populations and fertility. Here we review the genetic tools that are at present available to study germ cells in the mouse.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento/genética , Técnicas Genéticas , Células Germinativas/citologia , Células Germinativas/crescimento & desenvolvimento , Modelos Animais , Animais , Biomarcadores/metabolismo , Diferenciação Celular/fisiologia , Movimento Celular/fisiologia , Proliferação de Células , Cruzamentos Genéticos , Células Germinativas/metabolismo , Integrases/metabolismo , Camundongos , Camundongos Transgênicos
12.
PLoS Genet ; 4(8): e1000171, 2008 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-18769712

RESUMO

The thymus is a vertebrate-specific organ where T lymphocytes are generated. Genetic programs that lead to thymus development are incompletely understood. We previously screened ethylnitrosourea-induced medaka mutants for recessive defects in thymus development. Here we report that one of those mutants is caused by a missense mutation in a gene encoding the previously uncharacterized protein WDR55 carrying the tryptophan-aspartate-repeat motif. We find that WDR55 is a novel nucleolar protein involved in the production of ribosomal RNA (rRNA). Defects in WDR55 cause aberrant accumulation of rRNA intermediates and cell cycle arrest. A mutation in WDR55 in zebrafish also leads to analogous defects in thymus development, whereas WDR55-null mice are lethal before implantation. These results indicate that WDR55 is a nuclear modulator of rRNA synthesis, cell cycle progression, and embryonic organogenesis including teleost thymus development.


Assuntos
Ciclo Celular , Proteínas de Peixes/metabolismo , Proteínas Nucleares/metabolismo , Oryzias/crescimento & desenvolvimento , Oryzias/metabolismo , RNA Ribossômico/biossíntese , Timo/crescimento & desenvolvimento , Sequência de Aminoácidos , Animais , Nucléolo Celular/genética , Nucléolo Celular/metabolismo , Proteínas de Peixes/genética , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Células NIH 3T3 , Proteínas Nucleares/genética , Oryzias/genética , Fenótipo , Processamento Pós-Transcricional do RNA , RNA Ribossômico/genética , Alinhamento de Sequência , Timo/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento , Peixe-Zebra/metabolismo
13.
PLoS One ; 3(5): e2315, 2008 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-18509452

RESUMO

BACKGROUND: The dead-end (Dnd1) gene is essential for maintaining the viability of germ cells. Inactivation of Dnd1 results in sterility and testicular tumors. The Dnd1 encoded protein, DND1, is able to bind to the 3'-untranslated region (UTR) of messenger RNAs (mRNAs) to displace micro-RNA (miRNA) interaction with mRNA. Thus, one function of DND1 is to prevent miRNA mediated repression of mRNA. We report that DND1 interacts specifically with APOBEC3. APOBEC3 is a multi-functional protein. It inhibits retroviral replication. In addition, recent studies show that APOBEC3 interacts with cellular RNA-binding proteins and to mRNA to inhibit miRNA-mediated repression of mRNA. METHODOLOGY/PRINCIPAL FINDINGS: Here we show that DND1 specifically interacts with another cellular protein, APOBEC3. We present our data which shows that DND1 co-immunoprecipitates APOBEC3 from mammalian cells and also endogenous APOBEC3 from mouse gonads. Whether the two proteins interact directly remains to be elucidated. We show that both DND1 and APOBEC3 are expressed in germ cells and in the early gonads of mouse embryo. Expression of fluorescently-tagged DND1 and APOBEC3 indicate they localize to the cytoplasm and when DND1 and APOBEC3 are expressed together in cells, they sequester near peri-nuclear sites. CONCLUSIONS/SIGNIFICANCE: The 3'-UTR of mRNAs generally encode multiple miRNA binding sites as well as binding sites for a variety of RNA binding proteins. In light of our findings of DND1-APOBEC3 interaction and taking into consideration reports that DND1 and APOBEC3 bind to mRNA to inhibit miRNA mediated repression, our studies implicate a possible role of DND1-APOBEC3 interaction in modulating miRNA-mediated mRNA repression. The interaction of DND1 and APOBEC3 could be one mechanism for maintaining viability of germ cells and for preventing germ cell tumor development.


Assuntos
Citidina Desaminase/metabolismo , Células Germinativas/metabolismo , Proteínas de Neoplasias/metabolismo , Regiões 3' não Traduzidas , Animais , Citidina Desaminase/genética , DNA Complementar , Masculino , Camundongos , Proteínas de Neoplasias/genética , Ligação Proteica , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Testículo/metabolismo
14.
Mamm Genome ; 18(12): 832-8, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18049836

RESUMO

Germ cell tumor development in humans has been proposed to be part of testicular dysgenesis syndrome (TDS), which manifests as undescended testes, sterility, hypospadias, and, in extreme cases, as germ cell tumors. Males of the Ter mouse strain show interesting parallels to TDS because they either lack germ cells and are sterile or develop testicular germ cell tumors. We found that these defects in Ter mice are due to mutational inactivation of the Dead-end (Dnd1) gene. Here we report that chromosome X modulates germ cell tumor development in Ter mice. We tested whether the X or the Y chromosome influences tumor incidence. We used chromosome substitution strains to generate two new mouse strains: 129-Ter/Ter that carry either a C57BL/6J (B6)-derived chromosome (Chr) X or Y. We found that Ter/Ter males with B6-Chr X, but not B6-Chr Y, showed a significant shift in propensity from testicular tumor development to sterile testes phenotype. Thus, our studies provide unambiguous evidence that genetic factors from Chr X modulate the incidence of germ cell tumors in mice with inactivated Dnd1.


Assuntos
Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Testiculares/genética , Cromossomo X , Cromossomo Y , Animais , Cruzamentos Genéticos , Genótipo , Humanos , Masculino , Camundongos , Camundongos Mutantes , Neoplasias Embrionárias de Células Germinativas/patologia , Neoplasias Testiculares/patologia , Testículo/patologia
15.
Ann N Y Acad Sci ; 1120: 181-6, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17905939

RESUMO

Testicular germ-cell tumors occur in human males of all age groups, from infants to men over 50 years old. Most commonly, germ-cell tumors (generally known as testicular cancer) occur in young males between the ages of 15 to 35 years. The tumor tissues are usually histologically diverse, and testicular tumors that occur in the different age groups tend to be of specific histological subtypes. Most germ-cell tumors originate from primordial germ cells during embryonic development, although the progression and eventual detection of the disease occurs decades later in humans. Mouse strains spontaneously develop a specific subtype of testicular germ-cell tumors, the type I germ-cell tumors, and these tumors are similar to the germ-cell tumors (or teratomas) that occur in human infants. Some mouse strains, such as the 129-Ter strain, have extremely high germ-cell tumor incidences, making such strains ideal for genetic and biological studies of germ cell-tumor development. Here a brief overview of the recently identified genetic defect in the Ter strain, inactivation of the dead-end (Dnd1) gene, and the ongoing studies on Dnd1 to understand its role in germ-cell and germ cell-tumor development, are provided.


Assuntos
Proteínas de Neoplasias/fisiologia , Neoplasias Embrionárias de Células Germinativas/etiologia , Neoplasias Testiculares/etiologia , Animais , Diferenciação Celular/genética , Progressão da Doença , Células Germinativas/fisiologia , Humanos , Masculino , Camundongos , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Testiculares/genética
16.
Mamm Genome ; 18(8): 584-95, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17671812

RESUMO

Chromosome substitution strains (CSS or consomic strains) are useful for mapping phenotypes to chromosomes. However, huge efforts are needed to identify the gene(s) responsible for the phenotype in the complex context of the chromosome. Here we report the identification of candidate disease genes from a CSS by using a combination of genetic and genomic approaches and by using knowledge about the germ cell tumor disease etiology. We used the CSS 129.MOLF-Chr19 chromosome substitution strain, in which males develop germ cell tumors of the testes at an extremely high rate. We were able to identify three protein-coding genes and one microRNA on chromosome 19 that have previously not been implicated to be testicular tumor susceptibility genes. Our findings suggest that changes in gene expression levels in the gonadal tissues of multiple genes from Chr 19 likely contribute to the high testicular germ cell tumor (TGCT) incidence of the 129.MOLF-Chr19 strain. Our data advance the use of CSS to identify disease susceptibility genes and demonstrate that the 129.MOLF-Chr19 strain serves as a useful model to elucidate the genetics and biology of germ cell transformation and tumor development.


Assuntos
Cromossomos de Mamíferos , Predisposição Genética para Doença , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Testiculares/genética , Animais , Animais Recém-Nascidos , Embrião de Mamíferos , Regulação Neoplásica da Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos , MicroRNAs/genética , Modelos Biológicos , Neoplasias Embrionárias de Células Germinativas/patologia , Linhagem , Neoplasias Testiculares/patologia
17.
Biochem Biophys Res Commun ; 355(1): 194-9, 2007 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-17291453

RESUMO

Inactivation of the dead-end (Dnd1) gene in the Ter mouse strain results in depletion of primordial germ cells (PGCs) so that mice become sterile. However, on the 129 mouse strain background, loss of Dnd1 also increases testicular germ cell tumor incidence in parallel to PGC depletion. We report that inactivation of Dnd1 also affects embryonic viability in the 129 strain. Mouse Dnd1 encodes two protein isoforms, DND1-isoform alpha (DND1-alpha) and DND1-isoform beta (DND1-beta). Using isoform-specific antibodies, we determined DND1-alpha is expressed in embryos and embryonic gonads whereas DND1-beta expression is restricted to germ cells of the adult testis. Our data implicate DND1-alpha isoform to be necessary for germ cell viability and therefore its loss in Ter mice results in PGC depletion, germ cell tumor development and partial embryonic lethality in the 129 strain.


Assuntos
Desenvolvimento Embrionário/fisiologia , Células Germinativas/fisiologia , Proteínas de Neoplasias/genética , Animais , Anticorpos , Sobrevivência Celular , Células Cultivadas , DNA Complementar/genética , Perda do Embrião , Desenvolvimento Embrionário/genética , Feminino , Células Germinativas/citologia , Masculino , Camundongos , Camundongos Endogâmicos , Proteínas de Neoplasias/deficiência , Proteínas de Neoplasias/imunologia , Gravidez , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiologia , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células de Sertoli/citologia , Células de Sertoli/fisiologia
18.
Cell Cycle ; 4(9): 1136-8, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16082220

RESUMO

Testicular germ cell tumors (TGCTs) are the first tumors where the cell of origin, and the time of transformation were precisely defined. TGCTs in mice originate from primordial germ cells (PGCs) and develop within the testis during fetal development. TGCTs occur at an appreciable frequency (5%) only in the 129 family of inbred strains of laboratory mice, suggesting strong genetic control. These developmental and genetic properties make the 129 strains an exceptional model system to dissect TGCT pathogenesis. Ter is one of the most potent cancer modifiers genes known; it is a single gene mutation that causes progressive loss of PGCs on all inbred strain backgrounds and dramatically increased susceptibility to spontaneous TGCTs only on the 129 background. We recently showed that inactivation of the Dead end gene causes the Ter phenotype. Sequence analysis of Dead end encoded protein indicates it is homologous to factors involved in gene editing. The identity of Ter as Dead end and its function in PGCs will help clarify the role of editing in PGC biology and elucidate the causes of TGCTs in mice and humans.


Assuntos
Regulação Neoplásica da Expressão Gênica , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Testiculares/genética , Animais , Predisposição Genética para Doença , Células Germinativas/metabolismo , Humanos , Masculino , Camundongos , Mutação , Neoplasias Embrionárias de Células Germinativas/etiologia , Fenótipo , Análise de Sequência de DNA , Neoplasias Testiculares/etiologia , Testículo/embriologia
19.
Nature ; 435(7040): 360-4, 2005 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-15902260

RESUMO

In mice, the Ter mutation causes primordial germ cell (PGC) loss in all genetic backgrounds. Ter is also a potent modifier of spontaneous testicular germ cell tumour (TGCT) susceptibility in the 129 family of inbred strains, and markedly increases TGCT incidence in 129-Ter/Ter males. In 129-Ter/Ter mice, some of the remaining PGCs transform into undifferentiated pluripotent embryonal carcinoma cells, and after birth differentiate into various cells and tissues that compose TGCTs. Here, we report the positional cloning of Ter, revealing a point mutation that introduces a termination codon in the mouse orthologue (Dnd1) of the zebrafish dead end (dnd) gene. PGC deficiency is corrected both with bacterial artificial chromosomes that contain Dnd1 and with a Dnd1-encoding transgene. Dnd1 is expressed in fetal gonads during the critical period when TGCTs originate. DND1 has an RNA recognition motif and is most similar to the apobec complementation factor, a component of the cytidine to uridine RNA-editing complex. These results suggest that Ter may adversely affect essential aspects of RNA biology during PGC development. DND1 is the first protein known to have an RNA recognition motif directly implicated as a heritable cause of spontaneous tumorigenesis. TGCT development in the 129-Ter mouse strain models paediatric TGCT in humans. This work will have important implications for our understanding of the genetic control of TGCT pathogenesis and PGC biology.


Assuntos
Células Germinativas/metabolismo , Células Germinativas/patologia , Mutação/genética , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias Testiculares/genética , Neoplasias Testiculares/patologia , Alelos , Animais , Sequência de Bases , Peso Corporal , Mapeamento Cromossômico , Cromossomos de Mamíferos/genética , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Teste de Complementação Genética , Hibridização In Situ , Masculino , Camundongos , Neoplasias Embrionárias de Células Germinativas/genética , Neoplasias Embrionárias de Células Germinativas/metabolismo , Neoplasias Embrionárias de Células Germinativas/patologia , Tamanho do Órgão , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Neoplasias Testiculares/metabolismo , Testículo/embriologia , Testículo/metabolismo , Testículo/patologia
20.
Hum Mol Genet ; 12(4): 389-98, 2003 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-12566386

RESUMO

Testicular germ cell tumors (TGCTs) are the most common solid cancers affecting young men. Although the evidence for genetic predisposition to TGCTs in humans is compelling, the genetic control of susceptibility is poorly understood. The 129S1/SvImJ (129/Sv) inbred strain of mice is an excellent model for studying TGCT susceptibility. We previously reported a new mouse strain, the 129.MOLF-Chr19 chromosome substitution strain, which develops spontaneous TGCTs at a high frequency (70-80%) as compared with the much lower rate in the 129/Sv strain (5%). To characterize the genetic control of TGCT susceptibility, we created a panel of single- and double-congenic strains derived from 129.MOLF-Chr19. The frequency of TGCTs in these strains suggests that several genes with additive and epistatic effects located at distinct sites on chromosome 19 control susceptibility. However, an alternative interpretation involving epigenesis is based on a striking correlation between TGCT frequency and the length of the MOLF-derived congenic segment, regardless of their chromosomal location on Chr 19 in each congenic strain. We also show that bilateral TGCT cases result from the coincidental co-occurrence of unilateral TGCTs rather than from the action of distinct genes that control susceptibility to bilateral versus unilateral TGCT cases. Finally, we propose that these TGCTs result from disrupted testicular and spermatogenic developmental programs.


Assuntos
Predisposição Genética para Doença , Neoplasias Testiculares/genética , Animais , Mapeamento Cromossômico , Cruzamentos Genéticos , Genótipo , Humanos , Masculino , Camundongos , Camundongos Congênicos , Fenótipo , Espermatogênese , Neoplasias Testiculares/patologia
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