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Sex determination in the Squalius alburnoides complex: an initial characterization of sex cascade elements in the context of a hybrid polyploid genome.

Pala I, Schartl M, Thorsteinsdóttir S, Coelho MM - PLoS ONE (2009)

Bottom Line: Dmrt1 and wt1 transcripts were found at early stages of male development in S. alburnoides and are most likely implicated in the process of gonad development.For the first time in the study of this hybrid complex, it was possible to directly compare the gene expression patterns between the bisexual parental species and the various hybrid forms, for an extended set of genes.The contribution of these genes to gonad integrity maintenance and functionality is apparently unaltered in the hybrids, suggesting that no abrupt shifts in gene expression occurred as a result of hybridisation.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biologia Ambiental, Departamento de Biologia Animal, Faculdade de Ciências da Universidade de Lisboa, Lisbon, Portugal. iapala@fc.ul.pt

ABSTRACT

Background: Sex determination processes vary widely among different vertebrate taxa, but no group offers as much diversity for the study of the evolution of sex determination as teleost fish. However, the knowledge about sex determination gene cascades is scarce in this species-rich group and further difficulties arise when considering hybrid fish taxa, in which mechanisms exhibited by parental species are often disrupted. Even though hybridisation is frequent among teleosts, gene based approaches on sex determination have seldom been conducted in hybrid fish. The hybrid polyploid complex of Squalius alburnoides was used as a model to address this question.

Methodology/principal findings: We have initiated the isolation and characterization of regulatory elements (dmrt1, wt1, dax1 and figla) potentially involved in sex determination in S. alburnoides and in the parental species S. pyrenaicus and analysed their expression patterns by in situ hybridisation. In adults, an overall conservation in the cellular localization of the gene transcripts was observed between the hybrids and parental species. Some novel features emerged, such as dmrt1 expression in adult ovaries, and the non-dimorphic expression of figla, an ovarian marker in other species, in gonads of both sexes in S. alburnoides and S. pyrenaicus. The potential contribution of each gene to the sex determination process was assessed based on the timing and location of expression. Dmrt1 and wt1 transcripts were found at early stages of male development in S. alburnoides and are most likely implicated in the process of gonad development.

Conclusions/significance: For the first time in the study of this hybrid complex, it was possible to directly compare the gene expression patterns between the bisexual parental species and the various hybrid forms, for an extended set of genes. The contribution of these genes to gonad integrity maintenance and functionality is apparently unaltered in the hybrids, suggesting that no abrupt shifts in gene expression occurred as a result of hybridisation.

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Expression patterns of candidate genes in the adult gonads S. pyrenaicus and S. alburnoides.Dmrt1: (a) S. pyrenaicus (PP genotype); (b) S. alburnoides (AA genotype) and (c) S. alburnoides (PA genotype) testis; (d) S. alburnoides (PA genotype) and S. alburnoides (PAA genotype) ovary with (e) antisense and (f) sense probes. Wt1: (g) S. pyrenaicus (PP genotype); (h) S. alburnoides (AA genotype) and (i) S. alburnoides (PA genotype) testis; (j) S. alburnoides (PA genotype) and (k) S. alburnoides (PAA genotype) ovary. Dax1: (l) S. alburnoides (AA genotype), (m) S. pyrenaicus (PP genotype) and (n) S. alburnoides (PAA genotype) ovary. Examples of areas with positive signal are indicated by black arrows; examples of negative cells are highlighted with grey arrowheads. Germ cells (GS), early perinuclear (P), cortical alveolar (CA) and vitellogenic (V) oocytes. Scale bar = 100 µm (a, b, e, f, g, h, k, l, m); scale bar = 200 µm (c, d, i, j, n).
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pone-0006401-g005: Expression patterns of candidate genes in the adult gonads S. pyrenaicus and S. alburnoides.Dmrt1: (a) S. pyrenaicus (PP genotype); (b) S. alburnoides (AA genotype) and (c) S. alburnoides (PA genotype) testis; (d) S. alburnoides (PA genotype) and S. alburnoides (PAA genotype) ovary with (e) antisense and (f) sense probes. Wt1: (g) S. pyrenaicus (PP genotype); (h) S. alburnoides (AA genotype) and (i) S. alburnoides (PA genotype) testis; (j) S. alburnoides (PA genotype) and (k) S. alburnoides (PAA genotype) ovary. Dax1: (l) S. alburnoides (AA genotype), (m) S. pyrenaicus (PP genotype) and (n) S. alburnoides (PAA genotype) ovary. Examples of areas with positive signal are indicated by black arrows; examples of negative cells are highlighted with grey arrowheads. Germ cells (GS), early perinuclear (P), cortical alveolar (CA) and vitellogenic (V) oocytes. Scale bar = 100 µm (a, b, e, f, g, h, k, l, m); scale bar = 200 µm (c, d, i, j, n).

Mentions: Expression of dmrt1 was observed in specific cellular locations in males of the bisexual ancestor S. pyrenaicus (PP) and both in nuclear non-hybrid males (AA) and diploid hybrids (PA) of S. alburnoides (Figure 5a, b and c). The location of dmrt1 signal is observed in positions expected for Sertoli cells, as predicted previously from morphological comparison and gonad structural analysis [15]. Since the signal is more diffuse in AA gonads, we cannot exclude that dmrt1 may also be expressed in some peripheral spermatogonia (Figure 5b). Otherwise, no differences were observed between hybrids and parental species in terms of the location of positive sites of dmrt1 expression, although a comparatively stronger signal was usually obtained in PP and AA gonads (Figure 5a and b) compared to PA male hybrids (Figure 5c). Dmrt1 expression was observed in adult ovaries of diploid (PA) and triploid (PAA) hybrid females of S. alburnoides. Expression was confined to the more developed cortical alveolar and yolk vesicle oocytes and was apparently absent from earlier stage oocytes (Figure 5d and e).


Sex determination in the Squalius alburnoides complex: an initial characterization of sex cascade elements in the context of a hybrid polyploid genome.

Pala I, Schartl M, Thorsteinsdóttir S, Coelho MM - PLoS ONE (2009)

Expression patterns of candidate genes in the adult gonads S. pyrenaicus and S. alburnoides.Dmrt1: (a) S. pyrenaicus (PP genotype); (b) S. alburnoides (AA genotype) and (c) S. alburnoides (PA genotype) testis; (d) S. alburnoides (PA genotype) and S. alburnoides (PAA genotype) ovary with (e) antisense and (f) sense probes. Wt1: (g) S. pyrenaicus (PP genotype); (h) S. alburnoides (AA genotype) and (i) S. alburnoides (PA genotype) testis; (j) S. alburnoides (PA genotype) and (k) S. alburnoides (PAA genotype) ovary. Dax1: (l) S. alburnoides (AA genotype), (m) S. pyrenaicus (PP genotype) and (n) S. alburnoides (PAA genotype) ovary. Examples of areas with positive signal are indicated by black arrows; examples of negative cells are highlighted with grey arrowheads. Germ cells (GS), early perinuclear (P), cortical alveolar (CA) and vitellogenic (V) oocytes. Scale bar = 100 µm (a, b, e, f, g, h, k, l, m); scale bar = 200 µm (c, d, i, j, n).
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pone-0006401-g005: Expression patterns of candidate genes in the adult gonads S. pyrenaicus and S. alburnoides.Dmrt1: (a) S. pyrenaicus (PP genotype); (b) S. alburnoides (AA genotype) and (c) S. alburnoides (PA genotype) testis; (d) S. alburnoides (PA genotype) and S. alburnoides (PAA genotype) ovary with (e) antisense and (f) sense probes. Wt1: (g) S. pyrenaicus (PP genotype); (h) S. alburnoides (AA genotype) and (i) S. alburnoides (PA genotype) testis; (j) S. alburnoides (PA genotype) and (k) S. alburnoides (PAA genotype) ovary. Dax1: (l) S. alburnoides (AA genotype), (m) S. pyrenaicus (PP genotype) and (n) S. alburnoides (PAA genotype) ovary. Examples of areas with positive signal are indicated by black arrows; examples of negative cells are highlighted with grey arrowheads. Germ cells (GS), early perinuclear (P), cortical alveolar (CA) and vitellogenic (V) oocytes. Scale bar = 100 µm (a, b, e, f, g, h, k, l, m); scale bar = 200 µm (c, d, i, j, n).
Mentions: Expression of dmrt1 was observed in specific cellular locations in males of the bisexual ancestor S. pyrenaicus (PP) and both in nuclear non-hybrid males (AA) and diploid hybrids (PA) of S. alburnoides (Figure 5a, b and c). The location of dmrt1 signal is observed in positions expected for Sertoli cells, as predicted previously from morphological comparison and gonad structural analysis [15]. Since the signal is more diffuse in AA gonads, we cannot exclude that dmrt1 may also be expressed in some peripheral spermatogonia (Figure 5b). Otherwise, no differences were observed between hybrids and parental species in terms of the location of positive sites of dmrt1 expression, although a comparatively stronger signal was usually obtained in PP and AA gonads (Figure 5a and b) compared to PA male hybrids (Figure 5c). Dmrt1 expression was observed in adult ovaries of diploid (PA) and triploid (PAA) hybrid females of S. alburnoides. Expression was confined to the more developed cortical alveolar and yolk vesicle oocytes and was apparently absent from earlier stage oocytes (Figure 5d and e).

Bottom Line: Dmrt1 and wt1 transcripts were found at early stages of male development in S. alburnoides and are most likely implicated in the process of gonad development.For the first time in the study of this hybrid complex, it was possible to directly compare the gene expression patterns between the bisexual parental species and the various hybrid forms, for an extended set of genes.The contribution of these genes to gonad integrity maintenance and functionality is apparently unaltered in the hybrids, suggesting that no abrupt shifts in gene expression occurred as a result of hybridisation.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biologia Ambiental, Departamento de Biologia Animal, Faculdade de Ciências da Universidade de Lisboa, Lisbon, Portugal. iapala@fc.ul.pt

ABSTRACT

Background: Sex determination processes vary widely among different vertebrate taxa, but no group offers as much diversity for the study of the evolution of sex determination as teleost fish. However, the knowledge about sex determination gene cascades is scarce in this species-rich group and further difficulties arise when considering hybrid fish taxa, in which mechanisms exhibited by parental species are often disrupted. Even though hybridisation is frequent among teleosts, gene based approaches on sex determination have seldom been conducted in hybrid fish. The hybrid polyploid complex of Squalius alburnoides was used as a model to address this question.

Methodology/principal findings: We have initiated the isolation and characterization of regulatory elements (dmrt1, wt1, dax1 and figla) potentially involved in sex determination in S. alburnoides and in the parental species S. pyrenaicus and analysed their expression patterns by in situ hybridisation. In adults, an overall conservation in the cellular localization of the gene transcripts was observed between the hybrids and parental species. Some novel features emerged, such as dmrt1 expression in adult ovaries, and the non-dimorphic expression of figla, an ovarian marker in other species, in gonads of both sexes in S. alburnoides and S. pyrenaicus. The potential contribution of each gene to the sex determination process was assessed based on the timing and location of expression. Dmrt1 and wt1 transcripts were found at early stages of male development in S. alburnoides and are most likely implicated in the process of gonad development.

Conclusions/significance: For the first time in the study of this hybrid complex, it was possible to directly compare the gene expression patterns between the bisexual parental species and the various hybrid forms, for an extended set of genes. The contribution of these genes to gonad integrity maintenance and functionality is apparently unaltered in the hybrids, suggesting that no abrupt shifts in gene expression occurred as a result of hybridisation.

Show MeSH
Related in: MedlinePlus