Multiple alternative splicing of Dmrt1 during gonadogenesis in Indian mugger, a species exhibiting temperature-dependent sex determination

Alladi, Anand; Patel, Minarbha; Lalremruata, Albert; Singh, Ajeet; Agrawal, Raman; Singh, Lalji; Aggarwal, Ramesh K.

doi:10.1016/j.gene.2008.08.005

Cited by 33 publications

(32 citation statements)

References 32 publications

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“…Our previous study showed isolation of the DMRT1 homologue from Crocodylus palustris , along with its partial genomic locus. DMRT1 exhibits transcriptional diversity as seen in GSD species [Anand et al, 2008]. The karyotype analysis from the early passages of actively proliferating cultures showed only 30 homomorphic chromosomes without any distinguishable sex chromosome pair as reported earlier [Singh and Ray-Chaudhuri, 1973].…”

Section: Discussionsupporting

confidence: 75%

Cytogenetic Characterization and Fluorescence in situ Hybridization of (GATA)10 Repeats on Established Primary Cell Cultures from Indian Water Snake (Natrix piscator) and Indian Mugger (Crocodylus palustris) Embryos

Rao

Raseswari

Patel

et al. 2009

Cytogenet Genome Res

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Sex determination among reptiles has continued to draw the attention of geneticists and the mechanisms involved have been extensively studied and documented in the past 3 decades. The setting up of primary cell lines of reptilian tissues is an important tool in the present study which is a unique aspect not applied in earlier studies. Establishing the cell lines from various species of reptiles would help in our understanding of the mechanisms of evolution and differentiation of sex chromosomes. Therefore, in the present study, we have established for the first time primary cell cultures from Indian water snake (Natrix piscator) and Indian mugger (Crocodylus palustris) embryos. In the preliminary growth stage, 2 types of cells, fibroblast- and epithelial-like, were found to be attached and proliferating in vitro. These fibroblast-like cell cultures were later overtaken by epithelial cells. The cell lines were grown in minimal essential medium supplemented with fetal bovine serum and subcultured for a period of 8–10 months. The morphology of cell types was kept under constant observation microscopically. Interestingly, at a subsequent passage of the cells sporadically scattered neuronal-like and beating cells were observed. The suitable temperature for growth of these cell cultures was 28–30°C. Chromosome analysis was performed from the actively proliferating cells, which revealed 5 pairs of macrochromosomes and 15 pairs of microchromosomes in Natrix piscator, and 15 pairs of only macrochromosomes in Crocodylus palustris. (GATA)_n repeats are well known to be associated with sex chromosomes. Fluorescence in situ hybridization performed with (GATA)₁₀ repeats delineated the W chromosome in the cells of Natrix piscator which has so far not been reported. This cell culture method has presently only been applied to water snakes and crocodile embryos in the current study, but it will be employed in other reptilian species and could go a long way to being a sustainable source of primary cells. This would eventually serve as an important tool for molecular studies in reptiles and other species in the future.

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Section: Discussionsupporting

confidence: 75%

Cytogenetic Characterization and Fluorescence in situ Hybridization of (GATA)10 Repeats on Established Primary Cell Cultures from Indian Water Snake (Natrix piscator) and Indian Mugger (Crocodylus palustris) Embryos

Rao

Raseswari

Patel

et al. 2009

Cytogenet Genome Res

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“…The 495-bp DMRT1 fragment sequenced here (accession numbers FJ791118 for C. picta and FJ791119 for A. mutica ) was similar to Crocodylus palustris isoforms a1, a2, c, d, and e [Anand et al, 2008] and included the 3 half of the DM domain box [Raymond et al, 1999]. Sequence identity between the Chrysemys fragment and that of other reptiles and chicken was high: 99% identical to Trachemys, 97% to Mauremys , 96% to Apalone, Pelodiscus, and Lepidochelys, 89% to Crocodylus and Alligator , 83% to Gallus , 82% to Elaphe and Trimeresurus , and 80% to Calotes and Gekko.…”

Section: Dmrt1 and Sox9 Cloning And Expressionmentioning

confidence: 82%

Multivariate Expression Analysis of the Gene Network Underlying Sexual Development in Turtle Embryos with Temperature-Dependent and Genotypic Sex Determination

Valenzuela

2010

Sex Dev

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Sexual development has long been the target of study and despite great advances in our understanding of the composition and regulation of the gene network underlying gonadogenesis, our knowledge remains incomplete. Of particular interest is the relative role that the environment and the genome play in directing gonadal formation, especially the effect of environmental temperature in directing this process in vertebrates. Comparative analyses in closely related taxa with contrasting sex-determining mechanisms should help fill this gap. Here I present a multivariate study of the regulation of the gene network underlying sexual development in turtles with temperature-dependent (TSD; Chrysemys picta) and genotypic sex determination (GSD; Apalone mutica). I combine novel data on SOX9 and DMRT1 from these species with contrasting sex-determining mechanisms for the first time with previously reported data on DAX1, SF-1 (NR5A1), WT1, and aromatase (CYP19A1) from these same taxa. Comparative expression analyses of SOX9 and DMRT1 from these and other species indicate additional elements whose expression has diverged among TSD taxa, further supporting the notion that significant evolutionary changes have accrued in the regulation of the TSD gene network in reptiles. A non-parametric MANOVA revealed that temperature had a significant effect in multivariate gene expression in C. picta that varied during embryonic development, whereas the covariation of gene expression in A. mutica was insensitive to temperature. A phenotypic trajectory analysis (PTA) of gene expression comparing both species directly indicated that the relative covariation in gene expression varied between temperatures in C. picta. Furthermore, the 25°C trajectory of C. picta differed from that of A. mutica in the magnitude of gene expression change. Additional analyses revealed a stronger covariation in gene expression and a more interconnected regulatory network in A. mutica, consistent with the hypothesis that sexual development is a more canalized process in A. mutica, as would be expected if GSD evolved in this lineage through directional selection from its TSD ancestor.

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“…For example, in both the redeared slider and the olive ridley turtles, the gonadal ex- pression of DMRT1 is dimorphic during the early stages of the TSP in embryos incubated at MPT or FPT [Torres Maldonado et al, 2002;Shoemaker et al, 2007]. In the Indian mugger, Crocodylus palustris , the expression of DMRT1 is dimorphic during the TSP and has also been shown to undergo alternative splicing [Anand et al, 2008]. However, each isoform appears to be expressed more highly in the gonad of embryos incubated at the MPT relative to those incubated at FPT and, thus, would suggest a more critical role for transcriptional regulation in establishing dimorphic activity.…”

Section: Mechanisms Of Temperature-dependent Sex Determination In Repmentioning

confidence: 99%

Gonadal Differentiation in Reptiles Exhibiting Environmental Sex Determination

Kohno¹,

Parrott²,

Yatsu³

et al. 2014

Sex Dev

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As temperature-dependent sex determination (TSD) and homozygote or heterozygote genetic sex determination (GSD) exist in multiple reptilian taxa, they represent sex determination systems that have emerged de novo. Current investigations have revealed that the genetic mechanisms used by various reptilian species are similar to those used by other vertebrates. However, the recent completion or near completion of various reptilian genome projects suggests that new studies examining related species with and without TSD could begin to provide additional insight into the evolution of TSD and GSD in vertebrate ancestors. Major questions still remain concerning germ cell migration and specification, the differentiation of gonadal accessory cells, such as the Sertoli cells and granulosa cells of the developing testis and ovary, respectively, and the mechanisms by which gene expression is regulated during TSD events. Further, reptilian sentinels and their mechanisms of gonadogenesis will likely remain important indicator species for environmental health. Thus, ongoing and new investigations need to tie molecular information to gonadal morphogenesis and function in reptiles. Such data will not only provide important information for an understanding of the evolution of these phenomena in vertebrates, but could also provide an important understanding of the health of the environment around us.

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Multiple alternative splicing of Dmrt1 during gonadogenesis in Indian mugger, a species exhibiting temperature-dependent sex determination

Cited by 33 publications

References 32 publications

Cytogenetic Characterization and Fluorescence in situ Hybridization of (GATA)<sub>10</sub> Repeats on Established Primary Cell Cultures from Indian Water Snake <i>(Natrix piscator)</i> and Indian Mugger <i>(Crocodylus palustris)</i> Embryos

Cytogenetic Characterization and Fluorescence in situ Hybridization of (GATA)<sub>10</sub> Repeats on Established Primary Cell Cultures from Indian Water Snake <i>(Natrix piscator)</i> and Indian Mugger <i>(Crocodylus palustris)</i> Embryos

Multivariate Expression Analysis of the Gene Network Underlying Sexual Development in Turtle Embryos with Temperature-Dependent and Genotypic Sex Determination

Gonadal Differentiation in Reptiles Exhibiting Environmental Sex Determination

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