Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón
The circular RNAs (circRNAs) are a new class of recently reported RNAs, and their functions are still an emerging issue. Its biogenesis has been described as a product of a process called back-splicing that occurs during the maturation of pre-mRNA6,7. In this process the donor site of the exon is li...
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Facultad de Farmacia y Bioquímica
2019
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| Acceso en línea: | http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_3170 http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_3170.dir/3170.PDF |
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I28-R145-HWA_3170 |
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dspace |
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Universidad de Buenos Aires |
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I-28 |
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R-145 |
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Repositorio Digital de la Universidad de Buenos Aires (UBA) |
| language |
Español |
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spa |
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CircRNA Back-splicing RNAsa R Smaug1 Adipogénesis Ciencias de la vida |
| spellingShingle |
CircRNA Back-splicing RNAsa R Smaug1 Adipogénesis Ciencias de la vida Arizaca Maquera, Karol Andrea Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón |
| topic_facet |
CircRNA Back-splicing RNAsa R Smaug1 Adipogénesis Ciencias de la vida |
| description |
The circular RNAs (circRNAs) are a new class of recently reported RNAs, and their functions are still an emerging issue. Its biogenesis has been described as a product of a process called back-splicing that occurs during the maturation of pre-mRNA6,7. In this process the donor site of the exon is linked to the acceptor site thanks to a complementarity generated by the formation of intronic loops, thus generate a circular molecule that can be constituted only by exons (monoexonic, multi-exonic), only introns, or both4,17. It is also known that their structure gives them greater stability and makes them resistant to degradation by some RNases such as RNAse R, unlike linear RNAs.\nTo date, different functions of the circRNAs have been postulated as: competence of splicing against the gene from which are generated, sponges of miRNAs, regulators of methylation of CpG islands, or mediators in migration of pAKT to nucleus. Thus, circular RNAs are considered important post-transcriptional regulators, essentials for regulation of cell survival, and have been used as biomarkers in cancer4,10,11,12,18,19,20,21.\nThe greater accumulation of circRNAs has been reported in neural tissue, which agrees with the low cellular regeneration in that tissue, and with the greater stability against the degradation of circRNAs respect to linear RNAs. Its abundance in neural tissue has also led to relate them with development of different pathologies of the nervous system that are shown in older people.\nThe Smaug1 protein encoded by samd4 gene in mice and samd4a in humans is a translational repressor. This protein has been described in different organisms including Hommo Sapiens, Mus Musculus, Saccharomyces cerevisiae, and Drosophila Melanogaster. Initially described in Drosophila, the Smaug protein acts as regulator of the stability and translation of maternal mRNAs during embryonic development, interrupting the interaction between the factors of initiation of translation eIF4E and eIF4G52. In this way, Smaug captures target transcripts in repression complexes called S-foci. These silencing centers are organelles without membranes that behave as dynamic structures that self-assemble and contain mRNAs wich are not being translated, together with various proteins that regulate their translation, localization and degradation25,28,29,54.\nIn the mammalian genome there are 2 homologous genes called Smaug1 and Smaug2. Using mouse hippocampal neurons, in our laboratory we demonstrated that Smaug1 is located in synapse, regulating the synaptic plasticity. It was also shown that neuronal S-foci respond to synaptic stimuli 27,29,31,34. Subsequent to our findings, it has been shown that Smaug2 is involved on regulation of neurogenesis in mouse brain36.\nIn mammals Smaug1 and Smaug2 are expressed not only in CNS, but also in other tissues such: heart, bone, muscle and adipose tissue27,48,49. In our laboratory, we described the presence of two variants of splicing from Smaug1 in different human cell lines that arise from the alternative processing of exon III. It is important to note that the isoform of human Smaug1 lacking exon III shows RNA binding capacity, and a\nrepressor activity comparable to full-length human Smaug135. The homologue of Smaug1, Smaug2, is also present in the cell lines studied. Both, Smaug1 and Smaug2 form cytosolic bodies when are transfected into cell lines. Studies from another laboratory showed that in murine neural cortex cells, Smaug2 promotes neurogenesis by acting as translational repressor of nanos1 mRNA, a second RNA binding protein, capable of regulating translation and stability of transcripts36.\nAlthough Smaug1 and Smaug2 are more abundantly expressed in the brain, in mammals the Smaug mutants reported do not present phenotypes associated with the CNS. Recently, a samd4 mutation called "Supermodel" has been identified in mice, causing thinness and kyphosis associated with myopathy, and adipocyte defects. This point mutation generates a change of histidine for proline in the amino acid position 86 of the isoform1 from samd4, which in homozygosis generates mice resistant to the development of obesity induced by a high-fat diet. These metabolic defects in Supermodel mice were associated with modulation of target activities of the rapamycin1/mTORC1 (main regulator of metabolism) target complex mediated by Smaug1, evidenced by hypophosphorylation of the start translation factor 4E-BP1 and the ribosomal protein S6 in muscle and adipose tissues of homozygous mice48. Another research group analyzed a mutation of samd4 product of a transposon in homozygous mice that confers a phenotype associated with deficiency on development of skeleton, and reproduces the supermodel phenotype with a notable reduction of adipose tissue49.\nIt is suggested, then, that the post-transcriptional pathway mediated by Smaug1 is relevant not only for development of neurons, also for adipocyte development and during osteoblastogenesis.\nObesity is a disease with a high mortality rate worldwide. It is generated due to the large accumulation of triglycerides in adipocytes, generating hypertrophy and hyperplasia of adipose tissue, and is responsible for development Metabolic Syndrome (MS), branching into: central obesity, insulin resistance, non-alcoholic fatty liver, hypertension.\nIn this work, we identified the presence of Smaug1 circRNA (mmu_cir_0000529) in a model for the study of adipocyte differentiation and simultaneously analyzed the splicing of linear isoforms from Smaug1/Samd4. |
| author2 |
Fernández-Álvarez, Ana Julia |
| author_facet |
Fernández-Álvarez, Ana Julia Arizaca Maquera, Karol Andrea |
| format |
Tesis de maestría Tesis de maestría acceptedVersion |
| author |
Arizaca Maquera, Karol Andrea |
| author_sort |
Arizaca Maquera, Karol Andrea |
| title |
Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón |
| title_short |
Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón |
| title_full |
Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón |
| title_fullStr |
Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón |
| title_full_unstemmed |
Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón |
| title_sort |
detección de rnas circulares del gen smaug1/samd4 en diferentes tejidos de ratón |
| publisher |
Facultad de Farmacia y Bioquímica |
| publishDate |
2019 |
| url |
http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_3170 http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_3170.dir/3170.PDF |
| work_keys_str_mv |
AT arizacamaquerakarolandrea detecciondernascircularesdelgensmaug1samd4endiferentestejidosderaton |
| _version_ |
1766017531363983360 |
| spelling |
I28-R145-HWA_31702022-03-03 The circular RNAs (circRNAs) are a new class of recently reported RNAs, and their functions are still an emerging issue. Its biogenesis has been described as a product of a process called back-splicing that occurs during the maturation of pre-mRNA6,7. In this process the donor site of the exon is linked to the acceptor site thanks to a complementarity generated by the formation of intronic loops, thus generate a circular molecule that can be constituted only by exons (monoexonic, multi-exonic), only introns, or both4,17. It is also known that their structure gives them greater stability and makes them resistant to degradation by some RNases such as RNAse R, unlike linear RNAs.\nTo date, different functions of the circRNAs have been postulated as: competence of splicing against the gene from which are generated, sponges of miRNAs, regulators of methylation of CpG islands, or mediators in migration of pAKT to nucleus. Thus, circular RNAs are considered important post-transcriptional regulators, essentials for regulation of cell survival, and have been used as biomarkers in cancer4,10,11,12,18,19,20,21.\nThe greater accumulation of circRNAs has been reported in neural tissue, which agrees with the low cellular regeneration in that tissue, and with the greater stability against the degradation of circRNAs respect to linear RNAs. Its abundance in neural tissue has also led to relate them with development of different pathologies of the nervous system that are shown in older people.\nThe Smaug1 protein encoded by samd4 gene in mice and samd4a in humans is a translational repressor. This protein has been described in different organisms including Hommo Sapiens, Mus Musculus, Saccharomyces cerevisiae, and Drosophila Melanogaster. Initially described in Drosophila, the Smaug protein acts as regulator of the stability and translation of maternal mRNAs during embryonic development, interrupting the interaction between the factors of initiation of translation eIF4E and eIF4G52. In this way, Smaug captures target transcripts in repression complexes called S-foci. These silencing centers are organelles without membranes that behave as dynamic structures that self-assemble and contain mRNAs wich are not being translated, together with various proteins that regulate their translation, localization and degradation25,28,29,54.\nIn the mammalian genome there are 2 homologous genes called Smaug1 and Smaug2. Using mouse hippocampal neurons, in our laboratory we demonstrated that Smaug1 is located in synapse, regulating the synaptic plasticity. It was also shown that neuronal S-foci respond to synaptic stimuli 27,29,31,34. Subsequent to our findings, it has been shown that Smaug2 is involved on regulation of neurogenesis in mouse brain36.\nIn mammals Smaug1 and Smaug2 are expressed not only in CNS, but also in other tissues such: heart, bone, muscle and adipose tissue27,48,49. In our laboratory, we described the presence of two variants of splicing from Smaug1 in different human cell lines that arise from the alternative processing of exon III. It is important to note that the isoform of human Smaug1 lacking exon III shows RNA binding capacity, and a\nrepressor activity comparable to full-length human Smaug135. The homologue of Smaug1, Smaug2, is also present in the cell lines studied. Both, Smaug1 and Smaug2 form cytosolic bodies when are transfected into cell lines. Studies from another laboratory showed that in murine neural cortex cells, Smaug2 promotes neurogenesis by acting as translational repressor of nanos1 mRNA, a second RNA binding protein, capable of regulating translation and stability of transcripts36.\nAlthough Smaug1 and Smaug2 are more abundantly expressed in the brain, in mammals the Smaug mutants reported do not present phenotypes associated with the CNS. Recently, a samd4 mutation called "Supermodel" has been identified in mice, causing thinness and kyphosis associated with myopathy, and adipocyte defects. This point mutation generates a change of histidine for proline in the amino acid position 86 of the isoform1 from samd4, which in homozygosis generates mice resistant to the development of obesity induced by a high-fat diet. These metabolic defects in Supermodel mice were associated with modulation of target activities of the rapamycin1/mTORC1 (main regulator of metabolism) target complex mediated by Smaug1, evidenced by hypophosphorylation of the start translation factor 4E-BP1 and the ribosomal protein S6 in muscle and adipose tissues of homozygous mice48. Another research group analyzed a mutation of samd4 product of a transposon in homozygous mice that confers a phenotype associated with deficiency on development of skeleton, and reproduces the supermodel phenotype with a notable reduction of adipose tissue49.\nIt is suggested, then, that the post-transcriptional pathway mediated by Smaug1 is relevant not only for development of neurons, also for adipocyte development and during osteoblastogenesis.\nObesity is a disease with a high mortality rate worldwide. It is generated due to the large accumulation of triglycerides in adipocytes, generating hypertrophy and hyperplasia of adipose tissue, and is responsible for development Metabolic Syndrome (MS), branching into: central obesity, insulin resistance, non-alcoholic fatty liver, hypertension.\nIn this work, we identified the presence of Smaug1 circRNA (mmu_cir_0000529) in a model for the study of adipocyte differentiation and simultaneously analyzed the splicing of linear isoforms from Smaug1/Samd4. Fil: Arizaca Maquera, Karol Andrea. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Buenos Aires, Argentina Fernández-Álvarez, Ana Julia Facultad de Farmacia y Bioquímica Boccaccio, Graciela Lidia Arizaca Maquera, Karol Andrea 2019-05-24 Los RNA circulares (circRNAs) son una nueva clase de RNAs recientemente reportados, de los cuales sus funciones aún son un tema emergente. Su biogénesis se ha descrito como producto de un proceso llamado back-splicing que ocurre durante la maduración del pre-mRNA6,7. En este proceso el sitio dador del exón, se une al sitio aceptor gracias a una complementariedad generada por la formación de lazos intrónicos, generando así una molécula circular que puede estar constituida sólo por exones (monoexónico, multi-exónico), sólo intrones, o ambos4,17. Se conoce también que su estructura les confiere mayor estabilidad y los hace resistentes a la degradación por algunas RNAsas como la RNAsa R, a diferencia de los RNAs lineales.\nA la fecha se han postulado distintas funciones de los circRNAs como: competencia de splicing frente al gen del que derivan, esponjas de miRNAs, reguladores de la metilación de islas CpG, o mediadores en la migración de pAKT al núcleo. Así, los RNAs circulares son considerados importantes reguladores post transcripcionales fundamentales para la regulación de la sobrevida celular, y han sido utilizados como biomarcadores en cáncer4,10,11,12,18,19,20,21.\nLa mayor acumulación de circRNAs ha sido reportada en tejido neural, lo cual concuerda con la baja regeneración celular en dicho tejido, y con la mayor estabilidad frente a la degradación de los circRNAs respecto de los RNAs lineales. Su abundancia en tejido neural también ha llevado a relacionarlos con el desarrollo de distintas patologías del sistema nervioso que se muestran en personas de mayor edad.\nLa proteína Smaug1 codificada por el gen samd4 en ratones y samd4a en humanos, es un represor traduccional. Esta proteína ha sido descrita en distintos organismos incluyendo Hommo Sapiens, Mus Musculus, Saccharomyces cerevisiae, y Drosophila Melanogaster. Inicialmente descripta en Drosophila, la proteína Smaug actúa como reguladora de la estabilidad y traducción de mRNAs maternos durante el desarrollo embrionario, interrumpiendo la interacción entre los factores de inicio de la traducción eIF4E y eIF4G52. De esta manera Smaug captura transcriptos target en complejos de represión denominados S-foci. Estos focos de silenciamiento son organelas sin membranas que se comportan como estructuras dinámicas que se auto-ensamblan y contienen mRNAs que no están siendo traducidos, junto a diversas proteínas que regulan su traducción, localización y degradación25,28,29,54. En el genoma de mamíferos existen 2 genes homólogos denominados Smaug1 y Smaug2. Empleando neuronas de hipocampo de ratón, en nuestro laboratorio se demostró que Smaug1 se localiza en la sinapsis regulando la plasticidad sináptica. Se demostró también que los S-foci neuronales responden a estímulos sinápticos 27,29,31,34. Posteriormente a nuestros hallazgos, se ha demostrado que Smaug2 está implicado en la regulación de la neurogénesis en cerebro de ratón36.\nEn mamíferos Smaug1 y Smaug2 se expresan no sólo en el SNC sino también en otros tejidos como corazón, hueso, músculo y tejido adiposo27,48,49. En nuestro laboratorio se describió la presencia de dos variantes de splicing de Smaug1 en distintas líneas\ncelulares humanas, que surgen del procesamiento alternativo del exón III. Es importante resaltar que la isoforma de Smaug1 humana que carece del exón III muestra capacidad de unión al RNA, y una actividad represora comparable con la de Smaug1 humana de longitud completa35. El homólogo de Smaug1, Smaug2, también está presente en las líneas estudiadas. Tanto Smaug1 como Smaug2 forman cuerpos citosólicos cuando se transfectan en líneas celulares. Estudios de otro laboratorio demostraron que en células de corteza neural murina, Smaug2 promueve la neurogénesis al actuar como represor traduccional del mRNA de nanos1, una segunda proteína de unión al RNA, capaz de regular la traducción y estabilidad de los transcriptos36.\nA pesar de que Smaug1 y Smaug2 se expresan con mayor abundancia en cerebro, en mamíferos los mutantes reportados de la proteína no presentan fenotipos asociados al SNC. Recientemente se ha identificado en ratones una mutación puntual de samd4 llamada ?Supermodel?, que causa delgadez y cifosis asociadas con miopatía, y defectos de adipocitos. Esta mutación puntual genera un cambio de histidina por prolina en la posición aminoacídica 86 de la isoforma1 de samd4, que en homocigosis genera ratones resistentes al desarrollo de obesidad inducida por una dieta alta en grasas. Estos defectos del metabolismo en ratones Supermodel fueron asociados a la modulación de las actividades del target del complejo rapamicina1/mTORC1 (regulador principal del metabolismo) mediada por Smaug1, evidenciado por la hipofosforilación del factor de inicio de traducción 4E-BP1 y la proteína ribosomal S6 en tejidos musculares y adiposos de ratones homocigotas48. Otro grupo de investigación analizó una mutación de samd4 producto de un transposón en ratones homocigotos que confiere un fenotipo asociado a la deficiencia en el desarrollo del esqueleto, y reproduce el fenotipo supermodel con una notable reducción del tejido adiposo49.\nSe sugiere entonces, que la vía post-transcripcional mediada por Smaug1 es relevante no sólo para el desarrollo de las neuronas sino también para el desarrollo adipocitario y durante la osteoblastogénesis.\nLa obesidad es una enfermedad con alto índice de mortalidad a nivel mundial. Se genera debido a la gran acumulación de triglicéridos en los adipocitos generando hipertrofia e hiperplasia del tejido adiposo, y es responsable del desarrollo de Síndrome Metabólico (SM), ramificándose en: obesidad central, resistencia a la insulina, hígado graso no alcohólico, hipertensión.\nEn este trabajo identificamos la presencia del circRNA Smaug1 (mmu_cir_0000529) en un modelo para el estudio de la diferenciación adipocítica y analizamos simultáneamente el splicing de las isoformas lineales de Smaug1/Samd4. application/pdf Surace, Ezequiel I. De Gaudenzi, Javier Adamo, Hugo CircRNA Back-splicing RNAsa R Smaug1 Adipogénesis spa Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-nd/2.5/ar/ Ciencias de la vida Detección de RNAs circulares del gen Smaug1/SAMD4 en diferentes tejidos de ratón info:eu-repo/semantics/masterThesis info:ar-repo/semantics/tesis de maestría info:eu-repo/semantics/acceptedVersion http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_3170 http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_3170.dir/3170.PDF |