The genetic basis of sex determination in Populus provides molecular markers across the genus and indicates convergent evolution

Abstract Many dioecious angiosperms are trees, which only flower after years of vegetative development and do not usually exhibit marked secondary sexual dimorphism. Nevertheless, if the genetic basis of sex determination is known, the sex of an individual can be determined using molecular markers. Here, we report that in the genus Populus sect. Populus an XY system of sex determination, which is found in P. tremula and P. tremuloides, likely re-evolved from a ZW system present in P. alba, P. adenopoda and P. qiongdaoensis. Strikingly, this new XY system is mechanistically identical to the older system found in several species of the Populus sections Tacamahaca, Aigeiros and Turanga demonstrating a remarkable example of convergent evolution. In both XY systems, male-specific inversely repeated sequences appear to silence the ARR17 gene, which functions as a sex switch, via small interfering RNAs and DNA methylation. In the ZW system, female-specific copies of ARR17 appear to regulate dioecy. With this detailed information on the genetic basis of sex determination it was possible to develop molecular markers that can be utilized to determine the sex in seedlings and non-flowering trees of different poplar species. We used the female-specific ARR17 gene to develop a sex marker for P. alba and P. adenopoda. For P. grandidentata, we employed the male-specific ARR17 inverted repeat. Finally, we summarize previously described markers for P. tremula, P. tremuloides, P. trichocarpa, P. deltoides and P. nigra. These markers can be useful for poplar ecologists, geneticists and breeders.

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Targeting the autosomal Ceratitis capitata transformer gene using Cas9 or dCas9 to masculinize XX individuals without inducing mutations

BMC Genetics ◽

10.1186/s12863-020-00941-4 ◽

2020 ◽

Vol 21 (S2) ◽

Author(s):

Pasquale Primo ◽

Angela Meccariello ◽

Maria Grazia Inghilterra ◽

Andrea Gravina ◽

Giuseppe Del Corsano ◽

...

Keyword(s):

Sex Determination ◽

Ceratitis Capitata ◽

Mutagenic Activity ◽

Fruit Fly ◽

Agricultural Pests ◽

Guide Rna ◽

Xx Males ◽

Biallelic Mutations ◽

Female Specific ◽

Male Specific

Abstract Background Females of the Mediterranean fruit fly Ceratitis capitata (Medfly) are major agricultural pests, as they lay eggs into the fruit crops of hundreds of plant species. In Medfly, female sex determination is based on the activation of Cctransformer (Cctra). A maternal contribution of Cctra is required to activate Cctra itself in the XX embryos and to start and epigenetically maintain a Cctra positive feedback loop, by female-specific alternative splicing, leading to female development. In XY embryos, the male determining Maleness-on-the-Y gene (MoY) blocks this activation and Cctra produces male-specific transcripts encoding truncated CcTRA isoforms and male differentiation occurs. Results With the aim of inducing frameshift mutations in the first coding exon to disrupt both female-specific and shorter male-specific CcTRA open reading frames (ORF), we injected Cas9 ribonucleoproteins (Cas9 and single guide RNA, sgRNA) in embryos. As this approach leads to mostly monoallelic mutations, masculinization was expected only in G1 XX individuals carrying biallelic mutations, following crosses of G0 injected individuals. Surprisingly, these injections into XX-only embryos led to G0 adults that included not only XX females but also 50% of reverted fertile XX males. The G0 XX males expressed male-specific Cctra transcripts, suggesting full masculinization. Interestingly, out of six G0 XX males, four displayed the Cctra wild type sequence. This finding suggests that masculinization by Cas9-sgRNA injections was independent from its mutagenic activity. In line with this observation, embryonic targeting of Cctra in XX embryos by a dead Cas9 (enzymatically inactive, dCas9) also favoured a male-specific splicing of Cctra, in both embryos and adults. Conclusions Our data suggest that the establishment of Cctra female-specific autoregulation during the early embryogenesis has been repressed in XX embryos by the transient binding of the Cas9-sgRNA on the first exon of the Cctra gene. This hypothesis is supported by the observation that the shift of Cctra splicing from female to male mode is induced also by dCas9. Collectively, the present findings corroborate the idea that a transient embryonic inactivation of Cctra is sufficient for male sex determination.

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Both Loss-of-Function and Gain-of-Function Mutations in snf Define a Role for snRNP Proteins in Regulating Sex-lethal Pre-mRNA Splicing in Drosophila Development

Genetics ◽

10.1093/genetics/144.1.95 ◽

1996 ◽

Vol 144 (1) ◽

pp. 95-108

Author(s):

Helen K Salz ◽

Thomas W Flickinger

Keyword(s):

Sex Determination ◽

Rna Splicing ◽

Loss Of Function ◽

Functional Homology ◽

Snrnp Protein ◽

A Cell ◽

Sex Lethal ◽

Mutant Phenotypes ◽

Female Specific ◽

Male Specific

Abstract The Drosophila snf gene encodes a protein with functional homology to the mammalian UlA and U2B″ snRNP proteins. Studies, based on the analysis of three viable alleles, have suggested a role for snf in establishing the female-specific splicing pattern of the sex determination switch gene, Sex-lethal. Here, we show that the non-sex-specific lethal null allele is required for female sex determination, arguing against the formal possibility that the viable alleles disrupt a function unrelated to snf's wild-type function. Moreover, we find snf is required for normal cell growth and/or survival, as expected for a protein involved in a cell-vital process such as RNA splicing. We also show that of the three viable alleles only one, snfJA2, is a partial loss-of-function mutation. The other two viable alleles, snf1621 and snfe8H, encode antimorphic proteins. We find the antimorphic proteins are mislocalized and correlate their mislocalization with their molecular lesions and mutant phenotypes. Finally, we provide genetic evidence that the antimorphic alleles interfere with the autoregulatory splicing function of the Sex-lethal protein. Based on these studies we suggest a model in which the snRNP protein, Snf, functions with Sex-lethal to block recognition of the regulated male-specific exon.

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Assessment of seed quality and sex determination in hop seedlings using molecular markers

Ciência e Natura ◽

10.5902/2179460x43135 ◽

2020 ◽

Vol 42 ◽

pp. e45

Author(s):

Marília Pereira Machado ◽

Andreza Cerioni Belniaki ◽

André Felipe Bernert ◽

Erik Nunes Gomes ◽

João Carlos Bespalhok Filho ◽

...

Keyword(s):

Molecular Markers ◽

Sex Determination ◽

Seed Quality ◽

Germination Percentage ◽

Brewing Industry ◽

Breeding Programs ◽

Normal Seedling ◽

Physiological Quality ◽

Male Specific

Brazil is the world's third largest beer consumer and currently imports all of its hops for the brewing industry. Such a fact justifies the selection of hop genotypes adapted for cultivation locally, which requires high quality seeds and efficient sex determination of the seedlings. The objectives of this study were to develop a methodology to assess hop seed quality and to efficiently determine hop seedling sex through the use of male-specific molecular markers. Freshly harvested hop seeds were germinated with and without pre-chilling (3-5 ° C) for 3, 6 and 12 weeks and then germinated at 20 or 25 ° C in the presence or absence of light, evaluating germination percentage and germination speed index. F1 progenies were obtained from after seed germination in a greenhouse and seedlings sex was determined using male-specific molecular markers. The best conditions for physiological quality assessment of hop seeds used in the present study were pre-chilling for 12 weeks, followed by germination at 25 ° C, and normal seedling counts at 7 and 15 days. The progeny submitted to molecular marker sexing was composed of 61.3% female plants. The established methodologies presented here can be considered efficient and may contribute to expedite hops breeding programs.

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Sex-Biased Gene Expression and Isoform Profile of Brine Shrimp Artemia franciscana by Transcriptome Analysis

Animals ◽

10.3390/ani11092630 ◽

2021 ◽

Vol 11 (9) ◽

pp. 2630

Author(s):

Euna Jo ◽

Seung-Jae Lee ◽

Eunkyung Choi ◽

Jinmu Kim ◽

Jun-Hyuck Lee ◽

...

Keyword(s):

Sex Determination ◽

Brine Shrimp ◽

Model Organism ◽

Enrichment Analysis ◽

Go Enrichment ◽

Determination System ◽

Sex Determination System ◽

Female Specific ◽

Male Specific ◽

Go Enrichment Analysis

The brine shrimp Artemia has a ZW sex determination system with ZW chromosomes in females and ZZ chromosomes in males. Artemia has been considered a promising model organism for ZW sex-determining systems, but the genes involved in sex determination and differentiation of Artemia have not yet been identified. Here, we conducted transcriptome sequencing of female and male A. franciscana using PacBio Iso-Seq and Illumina RNA-Seq techniques to identify candidate sex determination genes. Among the 42,566 transcripts obtained from Iso-Seq, 23,514 were analyzed. Of these, 2065 (8.8%) were female specific, 2513 (10.7%) were male specific, and 18,936 (80.5%) were co-expressed in females and males. Based on GO enrichment analysis and expression values, we found 10 female-biased and 29 male-biased expressed genes, including DMRT1 and Sad genes showing male-biased expression. Our results showed that DMRT1 has three isoforms with five exons, while Sad has seven isoforms with 2–11 exons. The Sad gene is involved in ecdysteroid signaling related to molting and metamorphosis in arthropods. Further studies on ecdysteroid biosynthetic genes are needed to improve our understanding of Artemia sex determination. This study will provide a valuable resource for sex determination and differentiation studies on Artemia and other crustaceans with ZW systems.

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Sex Determination in the Drosophila Germline Is Dictated by the Sexual Identity of the Surrounding Soma

Genetics ◽

10.1093/genetics/155.4.1741 ◽

2000 ◽

Vol 155 (4) ◽

pp. 1741-1756 ◽

Cited By ~ 1

Author(s):

J A Waterbury ◽

J I Horabin ◽

D Bopp ◽

P Schedl

Keyword(s):

Sex Determination ◽

Sexual Identity ◽

Signaling Pathway ◽

X Chromosome ◽

Germ Cells ◽

Chromosome Counting ◽

Counting System ◽

Germline Expression ◽

Female Specific ◽

Male Specific

Abstract It has been suggested that sexual identity in the germline depends upon the combination of a nonautonomous somatic signaling pathway and an autonomous X chromosome counting system. In the studies reported here, we have examined the role of the sexual differentiation genes transformer (tra) and doublesex (dsx) in regulating the activity of the somatic signaling pathway. We asked whether ectopic somatic expression of the female products of the tra and dsx genes could feminize the germline of XY animals. We find that TraF is sufficient to feminize XY germ cells, shutting off the expression of male-specific markers and activating the expression of female-specific markers. Feminization of the germline depends upon the constitutively expressed transformer-2 (tra-2) gene, but does not seem to require a functional dsx gene. However, feminization of XY germ cells by TraF can be blocked by the male form of the Dsx protein (DsxM). Expression of the female form of dsx, DsxF, in XY animals also induced germline expression of female markers. Taken together with a previous analysis of the effects of mutations in tra, tra-2, and dsx on the feminization of XX germ cells in XX animals, our findings indicate that the somatic signaling pathway is redundant at the level tra and dsx. Finally, our studies call into question the idea that a cell-autonomous X chromosome counting system plays a central role in germline sex determination.

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AUTOREGULATORY FUNCTIONING OF A DROSOPHILA GENE PRODUCT THAT ESTABLISHES AND MAINTAINS THE SEXUALLY DETERMINED STATE

Genetics ◽

10.1093/genetics/107.2.231 ◽

1984 ◽

Vol 107 (2) ◽

pp. 231-277

Author(s):

Thomas W Cline

Keyword(s):

Sex Determination ◽

Dosage Compensation ◽

Cellular Response ◽

Regulatory Gene ◽

Activity Level ◽

Loss Of Function ◽

Lethal Effects ◽

Epigenetic Effect ◽

Female Specific ◽

Male Specific

ABSTRACT Sxl appears to head a regulatory gene hierarchy that controls Drosophila sexual dimorphism in response to the X chromosome/autosome balance. Only XXAA cells normally have Sxl + activity. It maintains both the female morphogenetic sequence and a level of X-linked dosage-compensated gene expression compatible with diplo-X cell survival. In the absence of this activity, male sexual development and dosage-compensated gene hyperactivation ensure. Loss-of-function Sxl mutations generally have female-specific lethal effects caused by upsets in dosage compensation. New female-viable Sxl mutant alleles and combinations which lack Sxl's female sex determination function, yet still provide sufficient dosage compensation function for diplo-X survival, are described here. Consequently, such mutants cause genotypic females to develop as phenotypic males. Some of these sex-transforming Sxl mutants do not require the maternally produced da + activity that is normally essential for the functioning of zygotic Sxl alleles. In this paper, products of these unusual alleles are shown to act in trans to induce the expression of zygotic Sxl + alleles that would otherwise be unable to function due to a lack of maternal da + activity. This result indicates a third function for Sxl + product: a positive autoregulatory role. Controls for the autoregulation experiments demonstrated the sex-trans-forming epigenetic effect of the da mutation for the first time in diploids. In these experiments the female-specific zygotic lethal effects that normally would have accompanied loss of maternal da + activity were suppressed by mutations known to block dosage-compensation gene hyperactivation-the autosomal, male-specific lethals. Three types of abnormal sexual phenotypes were produced in the experiments described here, each with important implications for the mechanism of sex determination: (1) a true intersex phenotype produced by one particular Sxl allele shows that Sxl + must be involved in the cellular response to the X/A balance rather than in its establishment; (2) a maternally induced, female-sterile phenotype indicates that either the process of autoregulation or the mutants used to demonstrate it are tissue specific and (3) a mosaic intersexual phenotype whose character implies that the Sxl + activity level is set early in development, both by the da +-mediated X/A balance signal and by autoregulation, and is maintained subsequently in a cell autonomous fashion, independent of the initiating X/A balance signal. Thus, this study supports the view that sex determination is truly determinative in the standard developmental sense, and that Sxl is the carrier of the sexually determined state.

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The gene virilizer is required for female-specific splicing controlled by Sxl, the master gene for sexual development in Drosophila

Development ◽

10.1242/dev.121.12.4017 ◽

1995 ◽

Vol 121 (12) ◽

pp. 4017-4026 ◽

Cited By ~ 12

Author(s):

A. Hilfiker ◽

H. Amrein ◽

A. Dubendorfer ◽

R. Schneiter ◽

R. Nothiger

Keyword(s):

Sex Determination ◽

Dosage Compensation ◽

Sexual Development ◽

Constitutive Expression ◽

Vital Function ◽

Genetic Mosaics ◽

Male Specific Lethal ◽

Female Specific ◽

Male Specific ◽

Lethal Genes

The gene virilizer (vir) is needed for dosage compensation and sex determination in females and for an unknown vital function in both sexes. In genetic mosaics, XX somatic cells mutant for vir differentiate male structures. One allele, vir2f, is lethal for XX, but not for XY animals. This female-specific lethality can be rescued by constitutive expression of Sxl or by mutations in msl (male-specific lethal) genes. Rescued animals develop as strongly masculinized intersexes or pseudomales. They have male-specifically spliced mRNA of tra, and when rescued by msl, also of Sxl. Our data indicate that vir is a positive regulator of female-specific splicing of Sxl and of tra pre-mRNA.

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Meta-QTL analysis and identification of candidate genes for quality, abiotic and biotic stress in durum wheat

Scientific Reports ◽

10.1038/s41598-021-91446-2 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Jose Miguel Soriano ◽

Pasqualina Colasuonno ◽

Ilaria Marcotuli ◽

Agata Gadaleta

Keyword(s):

Molecular Markers ◽

Abiotic Stress ◽

Durum Wheat ◽

Candidate Genes ◽

Biotic Stress ◽

Qtl Analysis ◽

Complex Traits ◽

Genetic Basis ◽

Agronomic Traits ◽

Plant Performance

AbstractThe genetic improvement of durum wheat and enhancement of plant performance often depend on the identification of stable quantitative trait loci (QTL) and closely linked molecular markers. This is essential for better understanding the genetic basis of important agronomic traits and identifying an effective method for improving selection efficiency in breeding programmes. Meta-QTL analysis is a useful approach for dissecting the genetic basis of complex traits, providing broader allelic coverage and higher mapping resolution for the identification of putative molecular markers to be used in marker-assisted selection. In the present study, extensive QTL meta-analysis was conducted on 45 traits of durum wheat, including quality and biotic and abiotic stress-related traits. A total of 368 QTL distributed on all 14 chromosomes of genomes A and B were projected: 171 corresponded to quality-related traits, 127 to abiotic stress and 71 to biotic stress, of which 318 were grouped in 85 meta-QTL (MQTL), 24 remained as single QTL and 26 were not assigned to any MQTL. The number of MQTL per chromosome ranged from 4 in chromosomes 1A and 6A to 9 in chromosome 7B; chromosomes 3A and 7A showed the highest number of individual QTL (4), and chromosome 7B the highest number of undefined QTL (4). The recently published genome sequence of durum wheat was used to search for candidate genes within the MQTL peaks. This work will facilitate cloning and pyramiding of QTL to develop new cultivars with specific quantitative traits and speed up breeding programs.

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The Sex Determination Cascade in the Silkworm

Genes ◽

10.3390/genes12020315 ◽

2021 ◽

Vol 12 (2) ◽

pp. 315

Author(s):

Xu Yang ◽

Kai Chen ◽

Yaohui Wang ◽

Dehong Yang ◽

Yongping Huang

Keyword(s):

Sex Determination ◽

Molecular Mechanisms ◽

Future Research ◽

Model Species ◽

Male And Female ◽

Master Regulators ◽

Primary Signal ◽

Basic Understanding ◽

Female Specific ◽

Working Mechanisms

In insects, sex determination pathways involve three levels of master regulators: primary signals, which determine the sex; executors, which control sex-specific differentiation of tissues and organs; and transducers, which link the primary signals to the executors. The primary signals differ widely among insect species. In Diptera alone, several unrelated primary sex determiners have been identified. However, the doublesex (dsx) gene is highly conserved as the executor component across multiple insect orders. The transducer level shows an intermediate level of conservation. In many, but not all examined insects, a key transducer role is performed by transformer (tra), which controls sex-specific splicing of dsx. In Lepidoptera, studies of sex determination have focused on the lepidopteran model species Bombyx mori (the silkworm). In B. mori, the primary signal of sex determination cascade starts from Fem, a female-specific PIWI-interacting RNA, and its targeting gene Masc, which is apparently specific to and conserved among Lepidoptera. Tra has not been found in Lepidoptera. Instead, the B. mori PSI protein binds directly to dsx pre-mRNA and regulates its alternative splicing to produce male- and female-specific transcripts. Despite this basic understanding of the molecular mechanisms underlying sex determination, the links among the primary signals, transducers and executors remain largely unknown in Lepidoptera. In this review, we focus on the latest findings regarding the functions and working mechanisms of genes involved in feminization and masculinization in Lepidoptera and discuss directions for future research of sex determination in the silkworm.

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