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RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle

The autosomal Sry-related gene, Sox9, encodes a transcription factor, which performs an important role in testis differentiation in mammals. In several reptiles, Sox9 is differentially expressed in gonads, showing a significant upregulation during the thermo-sensitive period (TSP) at the male-promot...

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Autores principales: Sifuentes-Romero, Itzel, Merchant-Larios, Horacio, Milton, Sarah L., Moreno-Mendoza, Norma, Díaz-Hernández, Verónica, García-Gasca, Alejandra
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3899968/
https://www.ncbi.nlm.nih.gov/pubmed/24705165
http://dx.doi.org/10.3390/genes4020293
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author Sifuentes-Romero, Itzel
Merchant-Larios, Horacio
Milton, Sarah L.
Moreno-Mendoza, Norma
Díaz-Hernández, Verónica
García-Gasca, Alejandra
author_facet Sifuentes-Romero, Itzel
Merchant-Larios, Horacio
Milton, Sarah L.
Moreno-Mendoza, Norma
Díaz-Hernández, Verónica
García-Gasca, Alejandra
author_sort Sifuentes-Romero, Itzel
collection PubMed
description The autosomal Sry-related gene, Sox9, encodes a transcription factor, which performs an important role in testis differentiation in mammals. In several reptiles, Sox9 is differentially expressed in gonads, showing a significant upregulation during the thermo-sensitive period (TSP) at the male-promoting temperature, consistent with the idea that SOX9 plays a central role in the male pathway. However, in spite of numerous studies, it remains unclear how SOX9 functions during this event. In the present work, we developed an RNAi-based method for silencing Sox9 in an in vitro gonad culture system for the sea turtle, Lepidochelys olivacea. Gonads were dissected as soon as the embryos entered the TSP and were maintained in organ culture. Transfection of siRNA resulted in the decrease of both Sox9 mRNA and protein. Furthermore, we found coordinated expression patterns for Sox9 and the anti-Müllerian hormone gene, Amh, suggesting that SOX9 could directly or indirectly regulate Amh expression, as it occurs in mammals. These results demonstrate an in vitro method to knockdown endogenous genes in gonads from a sea turtle, which represents a novel approach to investigate the roles of important genes involved in sex determination or differentiation pathways in species with temperature-dependent sex determination.
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spelling pubmed-38999682014-03-26 RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle Sifuentes-Romero, Itzel Merchant-Larios, Horacio Milton, Sarah L. Moreno-Mendoza, Norma Díaz-Hernández, Verónica García-Gasca, Alejandra Genes (Basel) Communication The autosomal Sry-related gene, Sox9, encodes a transcription factor, which performs an important role in testis differentiation in mammals. In several reptiles, Sox9 is differentially expressed in gonads, showing a significant upregulation during the thermo-sensitive period (TSP) at the male-promoting temperature, consistent with the idea that SOX9 plays a central role in the male pathway. However, in spite of numerous studies, it remains unclear how SOX9 functions during this event. In the present work, we developed an RNAi-based method for silencing Sox9 in an in vitro gonad culture system for the sea turtle, Lepidochelys olivacea. Gonads were dissected as soon as the embryos entered the TSP and were maintained in organ culture. Transfection of siRNA resulted in the decrease of both Sox9 mRNA and protein. Furthermore, we found coordinated expression patterns for Sox9 and the anti-Müllerian hormone gene, Amh, suggesting that SOX9 could directly or indirectly regulate Amh expression, as it occurs in mammals. These results demonstrate an in vitro method to knockdown endogenous genes in gonads from a sea turtle, which represents a novel approach to investigate the roles of important genes involved in sex determination or differentiation pathways in species with temperature-dependent sex determination. MDPI 2013-06-07 /pmc/articles/PMC3899968/ /pubmed/24705165 http://dx.doi.org/10.3390/genes4020293 Text en © 2013 by the authors; licensee MDPI, Basel, Switzerland. http://creativecommons.org/licenses/by/3.0/ This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/).
spellingShingle Communication
Sifuentes-Romero, Itzel
Merchant-Larios, Horacio
Milton, Sarah L.
Moreno-Mendoza, Norma
Díaz-Hernández, Verónica
García-Gasca, Alejandra
RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle
title RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle
title_full RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle
title_fullStr RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle
title_full_unstemmed RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle
title_short RNAi-Mediated Gene Silencing in a Gonad Organ Culture to Study Sex Determination Mechanisms in Sea Turtle
title_sort rnai-mediated gene silencing in a gonad organ culture to study sex determination mechanisms in sea turtle
topic Communication
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3899968/
https://www.ncbi.nlm.nih.gov/pubmed/24705165
http://dx.doi.org/10.3390/genes4020293
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