Caracterización de modificaciones epigenéticas inducidas por cortisol en genes asociados a intercambio iónico en línea celular de branquias de trucha arcoíris (Oncorhynchus mykiss)
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Fecha
2024
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Idioma
es
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Universidad Andrés Bello
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Licencia CC
Licencia CC
Resumen
La acuicultura consiste en la cría de especies acuáticas para consumo humano. Esta industria ha
crecido significativamente a nivel mundial, más de un 200% en los últimos 10 años. En Chile, la
acuicultura es una actividad económica clave, siendo a nivel mundial el segundo país productor
de la trucha arcoíris (Oncorhynchus mykiss). La trucha arcoíris es una especie anádroma, capaz
de migrar entre agua dulce y salada, proceso que requiere de adaptaciones fisiológicas mediante
la esmoltificación. Estos cambios de salinidad son claves en el organismo, por lo que se necesita
conocer en detalle las adaptaciones fisiológicas que ocurren en las branquias, el principal órgano
de adaptación a estos cambios. Tanto los cambios de salinidad como de las condiciones que
enfrentan los organismos durante el proceso de acuicultura pueden generar estrés en el
organismo, produciendo la liberación de cortisol, hormona capaz de modular la expresión génica
actuando como factor de transcripción al unirse a receptores de glucocorticoides. Existen otros
mecanismos de regulación de expresión génica, como las modificaciones epigenéticas, que
consisten en cambios superficiales y heredables a la estructura del DNA sin modificar la
secuencia misma. Dentro de estas transformaciones se encuentra la metilación de las citosinas.
Las citosinas CpG tienden a acumularse en regiones llamadas “Islas CpG”, y su metilación en
regiones promotoras está asociada con una modulación negativa en la expresión de genes, pero
el rol de la metilación en otras zonas del cuerpo génico es incierto.
Este estudio investiga la expresión de genes involucrados en el transporte iónico mediante un
cultivo celular de RT-Gills frente a una exposición a cortisol en concentraciones fisiológicas y
suprafisiológicas, y cómo podría estar siendo modulada mediante la metilación de sitios CpG
próximos al sitio de inicio de la transcripción. Para ello, se analizaron los genes slc12a2, slc9a1b,
znt1 y atp1a3. Los resultados muestran que slc12a2 disminuyó su expresión tras 72 horas de
exposición al cortisol, mientras que slc9a1b no mostró cambios significativos. El gen znt1
aumentó su expresión a las 3 horas, estabilizándose posteriormente. La expresión de atp1a3
disminuyó únicamente a las 3 horas. Se evaluó la metilación del DNA en atp1a3 y slc12a2,
revelando para atp1a3 patrones que sugieren una regulación epigenética en respuesta al estrés
inducido por cortisol. Se encontró una correlación positiva entre la metilación de sitios CpG
ubicados en el primer exón cercano al sitio de inicio de la transcripción y con la expresión génica
a las 3 horas. Además, mantuvo su perfil de metilación a las 72 horas, indicando el potencial uso
como un epimarcador de situaciones de liberación de cortisol, proporcionando una mejor
comprensión de los mecanismos biológicos que facilitan su supervivencia y bienestar en la
acuicultura.
Aquaculture involves breeding aquatic species aimed mainly for human consumption. This industry has grown significantly around the world, growing more than a 200% in the last decade. In Chile, aquaculture is a fundamental economic activity, being the world’s second largest producer of rainbow trout (Oncorhynchus mykiss). Rainbow trout is an anadromous species, can migrate from saline to fresh water, a process which requires physiological adaptations that happen during smoltification. These salinity changes are crucial to the organism, making them necessary to better understand the physiological adaptations that occur in the main organ involved, the gills. Both the salinity changes and the conditions faced by the organisms during the aquaculture process may cause stress, causing the liberation of cortisol, a hormone capable of modulating gene expression by first binding to glucocorticoid receptors. There are other regulatory mechanisms involved in gene expression, such as epigenetic modifications. These consist of superficial, temporary, and hereditary changes to the surface of the DNA sequence without altering the sequence itself. Among these modifications there is cytosine methylation. It has been described that CpG cytosines tend to be grouped in regions, called “CpG Islands.” Traditionally, methylation of CpG islands located in the promoter region is associated with a negative correlation in gene expression, however its role in other gene body elements is still unclear. This study focuses on investigating the response of genes involved in ionic transport using RT Gills, exposed to cortisol in physiological and supraphysiological concentrations, and how could this expression be regulated by methylation in CpG sites close to the transcription start site. The genes slc12a2, slc9a1b, znt1, and atp1a3 were studied. The results show that slc12a2 significantly decreased its expression after 72 hours of cortisol exposure, while slc9a1b showed no significant changes. The znt1 gene increased its expression after 3 hours, stabilizing afterwards. The expression of atp1a3 decreased only after 3 hours. Additionally, DNA methylation in atp1a3 and slc12a2 was evaluated, revealing specific patterns for atp1a3 that suggest epigenetic regulation in response to cortisol-induced stress. It was found that methylation of CpG sites located in the first exon near the transcription start site showed a positive correlation with gene expression at 3 hours and maintained its methylation profile after 72 hours, indicating the potential use as an epimarker of cortisol release situations. This provides a better understanding of the biological mechanisms that facilitate their survival and welfare in aquaculture.
Aquaculture involves breeding aquatic species aimed mainly for human consumption. This industry has grown significantly around the world, growing more than a 200% in the last decade. In Chile, aquaculture is a fundamental economic activity, being the world’s second largest producer of rainbow trout (Oncorhynchus mykiss). Rainbow trout is an anadromous species, can migrate from saline to fresh water, a process which requires physiological adaptations that happen during smoltification. These salinity changes are crucial to the organism, making them necessary to better understand the physiological adaptations that occur in the main organ involved, the gills. Both the salinity changes and the conditions faced by the organisms during the aquaculture process may cause stress, causing the liberation of cortisol, a hormone capable of modulating gene expression by first binding to glucocorticoid receptors. There are other regulatory mechanisms involved in gene expression, such as epigenetic modifications. These consist of superficial, temporary, and hereditary changes to the surface of the DNA sequence without altering the sequence itself. Among these modifications there is cytosine methylation. It has been described that CpG cytosines tend to be grouped in regions, called “CpG Islands.” Traditionally, methylation of CpG islands located in the promoter region is associated with a negative correlation in gene expression, however its role in other gene body elements is still unclear. This study focuses on investigating the response of genes involved in ionic transport using RT Gills, exposed to cortisol in physiological and supraphysiological concentrations, and how could this expression be regulated by methylation in CpG sites close to the transcription start site. The genes slc12a2, slc9a1b, znt1, and atp1a3 were studied. The results show that slc12a2 significantly decreased its expression after 72 hours of cortisol exposure, while slc9a1b showed no significant changes. The znt1 gene increased its expression after 3 hours, stabilizing afterwards. The expression of atp1a3 decreased only after 3 hours. Additionally, DNA methylation in atp1a3 and slc12a2 was evaluated, revealing specific patterns for atp1a3 that suggest epigenetic regulation in response to cortisol-induced stress. It was found that methylation of CpG sites located in the first exon near the transcription start site showed a positive correlation with gene expression at 3 hours and maintained its methylation profile after 72 hours, indicating the potential use as an epimarker of cortisol release situations. This provides a better understanding of the biological mechanisms that facilitate their survival and welfare in aquaculture.
Notas
Memoria (Ingeniero en Biotecnología)
Palabras clave
Trucha Arcoiris, Genética, Cortisol