An Immune-Related Gene Evolved into the Master Sex-Determining Gene in Rainbow Trout, Oncorhynchus mykiss

Since the discovery of Sry in mammals [1, 2], few other master sex-determining genes have been identified in vertebrates [3-7]. To date, all of these genes have been characterized as well-known factors in the sex differentiation pathway, suggesting that the same subset of genes have been repeatedly and independently selected throughout evolution as master sex determinants [8, 9]. Here, we characterized in rainbow trout an unknown gene expressed only in the testis, with a predominant expression during testicular differentiation. This gene is a male-specific genomic sequence that is colocalized along with the sex-determining locus. This gene, named sdY for sexually dimorphic on the Y?chromosome, encodes a protein that displays similarity to the C-terminal domain of interferon regulatory factor 9. The targeted inactivation of sdY in males using zinc-finger nuclease induces ovarian differentiation, and the overexpression of sdY in females using additive transgenesis induces testicular differentiation. Together, these results demonstrate that sdY is a novel vertebrate master sex-determining gene not related to any known sex-differentiating gene. These findings highlight an unexpected evolutionary plasticity in vertebrate sex determination through the demonstration that master sex determinants can arise from the de novo evolution of genes that have not been previously implicated in sex differentiation.     

TEMPERATURE‐DEPENDENT TURNOVERS IN SEX‐DETERMINATION MECHANISMS: A QUANTITATIVE MODEL

Sex determination is often seen as a dichotomous process: individual sex is assumed to be determined either by genetic (genotypic sex determination, GSD) or by environmental factors (environmental sex determination, ESD), most often temperature (temperature sex determination, TSD). We endorse an alternative view, which sees GSD and TSD as the ends of a continuum. Both effects interact a priori, because temperature can affect gene expression at any step along the sex‐determination cascade. We propose to define sex‐determination systems at the population‐ (rather than individual) level, via the proportion of variance in phenotypic sex stemming from genetic versus environmental factors, and we formalize this concept in a quantitative‐genetics framework. Sex is seen as a threshold trait underlain by a liability factor, and reaction norms allow modeling interactions between genotypic and temperature effects (seen as the necessary consequences of thermodynamic constraints on the underlying physiological processes). As this formalization shows, temperature changes (due to e.g., climatic changes or range expansions) are expected to provoke turnovers in sex‐ determination mechanisms, by inducing large‐scale sex reversal and thereby sex‐ratio selection for alternative sex‐determining genes. The frequency of turnovers and prevalence of homomorphic sex chromosomes in cold‐blooded vertebrates might thus directly relate to the temperature dependence in sex‐determination mechanisms.     

Dosage compensation in birds

The Z and W sex chromosomes of birds have evolved independently from the mammalian X and Y chromosomes [1]. Unlike mammals, female birds are heterogametic (ZW), while males are homogametic (ZZ). Therefore male birds, like female mammals, carry a double dose of sex-linked genes relative to the other sex. Other animals with nonhomologous sex chromosomes possess "dosage compensation" systems to equalize the expression of sex-linked genes. Dosage compensation occurs in animals as diverse as mammals, insects, and nematodes, although the mechanisms involved differ profoundly [2]. In birds, however, it is widely accepted that dosage compensation does not occur [3-5], and the differential expression of Z-linked genes has been suggested to underlie the avian sex-determination mechanism [6]. Here we show equivalent expression of at least six of nine Z chromosome genes in male and female chick embryos by using real-time quantitative PCR [7]. Only the Z-linked ScII gene, whose ortholog in Caenorhabditis elegans plays a crucial role in dosage compensation [8], escapes compensation by this assay. Our results imply that the majority of Z-linked genes in the chicken are dosage compensated.     

The sex-determination genes fruitless and doublesex specify a neural substrate required for courtship song

Courtship song is a critical component of male courtship behavior in Drosophila, making the female more receptive to copulation and communicating species-specific information [1-6]. Sex mosaic studies have shown that the sex of certain regions of the central nervous system (CNS) is critical to song production [7]. Our examination of one of these regions, the mesothoracic ganglion (Msg), revealed the coexpression of two sex-determination genes, fruitless (fru) and doublesex (dsx). Because both genes are involved in creating a sexually dimorphic CNS [8, 9] and are necessary for song production [10-13], we investigated the individual contributions of fru and dsx to the specification of a male CNS and song production. We show a novel requirement for dsx in specifying a sexually dimorphic population of fru-expressing neurons in the Msg. Moreover, by using females constitutively expressing the male-specific isoforms of fru (Fru(M)), we show a critical requirement for the male isoform of dsx (Dsx(M)), alongside Fru(M), in the specification of courtship song. Therefore, although Fru(M) expression is sufficient for the performance of many male-specific behaviors [14], we have shown that without Dsx(M), the determination of a male-specific CNS and thus a full complement of male behaviors are not realized.     

脊椎动物性别决定和分化的分子机制研究进展

哺乳类性别决定是多种转录因子和生长因子相继表达和相互调控的结果。SRY的表达启动雄性通路并诱导下游雄性特异基因SOX9、AMH等的表达。FOXL2在雌性未分化性腺表达,WNT-4和DAX1也在雌性性别决定或分化时期表达,表明雌性通路也是受特定基因调控的,而并非“默认通路”。鸟类的性别也是由遗传基因决定的,EFT1(雌性)和DMRT1(雄性)可能是性别决定候选基因。爬行类为温度性别决定的典型,温度可能通过调节雌激素水平和控制性别特异遗传基因表达决定性别。大部分两栖类性别受环境因素影响,但发现DMRT1和DAX1可能与其精巢发育有关。鱼类性别决定和分化方式差异很大,多种因素(遗传基因、环境因素、类固醇激素等)参与了这一过程。从青Q鳉Y染色体定位克隆的DMY,被认为是第一个非哺乳类脊椎动物雄性性别决定基因。所有这些表明脊椎动物性别决定和分化机制是多样化的。     

Phylogeny of sex‐determining mechanisms in squamate reptiles: are sex chromosomes an evolutionary trap?

Squamate reptiles possess two general modes of sex determination: (1) genotypic sex determination (GSD), where the sex of an individual is determined by sex chromosomes, i.e. by sex‐specific differences in genotype; and (2) temperature‐dependent sex determination (TSD), where sex chromosomes are absent and sex is determined by nongenetic factors. After gathering information about sex‐determining mechanisms for more than 400 species, we employed comparative phylogenetic analyses to reconstruct the evolution of sex determination in Squamata. Our results suggest relative uniformity in sex‐determining mechanisms in the majority of the squamate lineages. Well‐documented variability is found only in dragon lizards (Agamidae) and geckos (Gekkota). Polarity of the sex‐determining mechanisms in outgroups identified TSD as the ancestral mode for Squamata. After extensive review of the literature, we concluded that to date there is no known well‐documented transition from GSD to TSD in reptiles, although transitions in the opposite direction are plentiful and well corroborated by cytogenetic evidence. We postulate that the evolution of sex‐determining mechanisms in Squamata was probably restricted to the transitions from ancestral TSD to GSD. In other words, transitions were from the absence of sex chromosomes to the emergence of sex chromosomes, which have never disappeared and constitute an evolutionary trap. This evolutionary trap hypothesis could change the understanding of phylogenetic conservatism of sex‐determining systems in many large clades such as butterflies, snakes, birds, and mammals. © 2009 The Linnean Society of London, Zoological Journal of the Linnean Society, 2009, 156 , 168–183.     

Preservation of the Y transcriptome in a 10-million-year-old plant sex chromosome system

Classical genetic studies discovered loss of genes from the ancient sex chromosome systems of several animals (genetic degeneration), and complete genome sequencing confirms that the heterogametic sex is hemizygous for most sex-linked genes. Genetic degeneration is thought to result from the absence of recombination between the sex chromosome pair (reviewed by [1]) and is very rapid after sex chromosome-autosome fusions in Drosophila [2-4]. Plant sex chromosome systems allow study of the time course of degeneration, because they evolved from a state wholly without sex chromosomes (rather than after a large genome region fused to a preexisting sex chromosome), and, in several taxa, recombination stopped very recently. However, despite increasing genetic and physical mapping of plant nonrecombining sex-determining regions [5-8], it remains very difficult to discover sex-linked genes, and it is unclear whether Y-linked genes are losing full function. We therefore developed a high-throughput method using RNA-Seq to identify sex linkage in Silene latifolia. Recombination suppression between this plant's XY sex chromosome pair started only about 10 million years ago [9]. Our approach identifies several hundred new sex-linked genes, and we show that this young Y chromosome retains many genes, yet these already have slightly reduced gene expression and are accumulating changes likely to reduce protein functions.     

Sexual maldevelopment and sex reversal, chromosomal causes

The SRY gene on the Y chromosome is the testis determining factor (TDF). It is therefore the initial male determining factor. However, phenotypic sex determination includes a cascade of genes located on autosomes as well as sex chromosomes. Aberrations of these genes may cause sexual maldevelopment or sex reversal. Abnormalities may include single gene mutations and gene loss or gain-changes may involve only sex organs or may be part of syndromes. These changes may also arise as chromosome abnormalities involving contiguous genes. Eight cases with chromosomal abnormalities involving different causative mechanisms are described herein. The most common cause is nondisjunction, including loss or gain of sex chromosomes. Less common causes are mispairing and crossing over in meiosis, chromosome breaks with repair, nonhomologous pairing due to low copy repeats and crossing over, and translocation (familial or de novo) with segregation. Cases include: [see: text].     

Regulation of ARNO nucleotide exchange by a PH domain electrostatic switch.

ARNO is a member of a family of guanine nucleotide exchange factors that activate small GTPases called ADP-ribosylation factors (ARFs) [1] [2] [3], which regulate vesicular trafficking and, in one case (ARF6), also regulate cortical actin structure [4]. ARNO is located at the plasma membrane, and in the presence of activated protein kinase C (PKC) can induce cortical actin rearrangements reminiscent of those produced by active ARF6 [5] [6] [7] [8]. High-affinity binding of ARNO to membranes, which is required for exchange activity, is mediated cooperatively by a pleckstrin homology (PH) domain and an adjacent carboxy-terminal polybasic domain [3] [9]. ARNO is phosphorylated in vivo by PKC on a single serine residue, S392, located within the carboxy-terminal polybasic domain. Mutation of S392 to alanine does not prevent ARNO-mediated actin rearrangements, suggesting that phosphorylation does not lead to ARNO activation [6]. Here, we report that phosphorylation negatively regulates ARNO exchange activity through a 'PH domain electrostatic switch'. Introduction of a negatively charged phosphate into the polybasic domain reduced interaction of ARNO with membranes both in vitro and in vivo, and inhibited exchange in vitro. This regulated membrane association is similar to the myristoyl electrostatic switch that controls membrane binding of the myristoylated alanine-rich C kinase substrate (MARCKS) [10], but to our knowledge is the first demonstration of an electrostatic switch regulating the membrane interaction of a protein containing a PH domain. This mechanism allows regulation of ARNO lipid binding and exchange activity at two levels, phosphoinositide-dependent recruitment and PKC-dependent displacement from the membrane.     

A G protein-coupled receptor with a lipid kinase domain is involved in cell-density sensing

One mechanism multicellular structures use for controlling cell number [1, 2] involves the secretion and sensing of a factor, such as leptin [3] or myostatin [4], in mammals. Dictyostelium cells secrete autocrine factors for sensing cell density prior to aggregation and multicellular development [5, 6] such as CMF (conditioned-medium factor), which enables starving cells to respond to cAMP pulses [7-9]. Its actions are mediated by two receptors. CMFR1 activates a G protein-independent signaling pathway regulating gene expression [10]. An unknown Galpha1-dependent receptor activates phospholipase C (PLC), which regulates the lifetime of Galpha2-GTP [11-13]. Here, we describe RpkA, an unusual seven-transmembrane receptor that is fused to a C-terminal PIP5 kinase domain and that localizes in membranes of a late endosomal compartment. Loss of RpkA resulted in formation of persistent loose aggregates and altered expression of cAMP-regulated genes. The developmental defect can be rescued by full-length RpkA and the transmembrane domain only. The PIP5 kinase domain is dispensable for the developmental role of RpkA. rpkA- cells secrete and bind CMF but are unable to induce downstream responses. Inactivation of Galpha1, a negative regulator of CMF signaling, rescued the developmental defect of the rpkA- cells, suggesting that RpkA actions are mediated by Galpha1.     

Role of acylCoA binding protein in acylCoA transport,metabolism and cell signaling

Long chain acylCoA esters (LCAs) act both as substrates and intermediates in intermediary metabolism and as regulators in various intracellular functions. AcylCoA binding protein (ACBP) binds LCAs with high affinity and is believed to play an important role in intracellular acylCoA transport and pool formation and therefore also for the function of LCAs as metabolites and regulators of cellular functions [1]. The major factors controlling the free concentration of cytosol long chain acylCoA ester (LCA) include ACBP [2], sterol carrier protein 2 (SCP2) [3] and fatty acid binding protein (FABP) [4]. Additional factors affecting the concentration of free LCA include feed back inhibition of the acylCoA synthetase [5], binding to acylCoA receptors (LCA-regulated molecules and enzymes), binding to membranes and the activity of acylCoA hydrolases [6].     

Sex determination in the germ line of Drosophila depends on genetic signals and inductive somatic factors

We have analyzed the mechanism of sex determination in the germ line of Drosophila by manipulating three parameters: (1) the ratio of X-chromosomes to sets of autosomes (X:A); (2) the state of activity of the gene Sex-lethal (Sxl), and (3) the sex of the gonadal soma. To this end, animals with a ratio of 2X:2A and 2X:3A were sexually transformed into pseudomales by mutations at the sex-determining genes Sxl (Sex-lethal), tra (transformer), tra-2 (transformer-2), or dsx (double-sex). Animals with the karyotype 2X;3A were also transformed into pseudofemales by the constitutive mutation SxlM1. The sexual phenotype of the gonads and of the germ cells was assessed by phase-contrast microscopy. Confirming the conclusions of Steinmann-Zwicky et al. (Cell 57, 157, 1989), we found that all three parameters affect sex determination in germ cells. In contrast to the soma in which sex determination is completely cell-autonomous, sex determination in the germ line has a non-autonomous component inasmuch as the sex of the soma can influence the sexual pathway of the germ cells. Somatic induction has a clear effect on 2X;2A germ cells that carry a Sxl+ allele. These cells, which form eggs in an ovary, can enter spermatogenesis in testes. Mutations that cause partial loss of function or gain of function of Sxl thwart somatic induction and, independently of the sex of the soma, dictate spermatogenesis or oogenesis, respectively. Somatic induction has a much weaker effect on 2X;3A germ cells. This ratio is essentially a male signal for germ cells which consistently enter spermatogenesis in testes, even when they carry SxlM1. In a female soma, however, SxlM1 enables the 2X;3A germ cells to form almost normal eggs. Our results show that sex determination in the germ line is more complex than in the soma. They provide further evidence that the state of Sxl, the key gene for sex determination and dosage compensation in the soma, also determines the sex of the germ cells, and that, in the germ line, the state of activity of Sxl is regulated not only by the X:A ratio, but also by somatic inductive stimuli.     

Association of a phosphatidylinositol-specific 3-kinase with a human trans-Golgi network resident protein

The eukaryotic trans-Golgi network (TGN) is a key site for the formation of transport vesicles destined for different intracellular compartments [1]. A key marker for the mammalian TGN is TGN38/46 [2]. This integral membrane glycoprotein cycles between the TGN and the cell surface and is implicated in recruitment of cytosolic factors and regulation of at least one type of vesicle formation at the mammalian TGN [2] and [3]. In this study, we have identified a phosphatidylinositol (PtdIns)-specific 3-kinase activity associated with the human orthologue (TGN46), which is sensitive to lipid kinase inhibitors. Treatment of HeLa cells with low levels of these inhibitors reveals subtle morphological changes in TGN46-positive compartments. Our findings suggest a role for PtdIns 3-kinases and presumably for the product, PtdIns 3-phosphate (PtdIns3P), in the formation of secretory transport vesicles by mechanisms conserved in yeast and mammals.     

Familial Clustering of Abdominal Visceral Fat and Total Fat Mass: The Québec Family Study

The evidence for common familial factors underlying total fat mass (estimated from underwater weighing) and abdominal visceral fat (assessed from CT scan) was examined in families participating in phase 2 of the Québec Family Study (QFS) using a bivariate familial correlation model. Previous QFS investigations suggest that both genetic (major and polygenic) and familial environmental factors influence each phenotype, accounting for between 55% to 71% of the phenotypic variance in fat mass, and between 55% to 72% for abdominal visceral fat The current study suggests that the bivariate familial effect ranges from 29% to 50%. This pattern suggests that there may be common familial determinants for abdominal visceral fat and total fat mass, as well as additional familial factors which are specific to each. The relatively high spouse cross-trait correlations usually suggest that a large percent of the bivariate familial effect may be environmental in origin. However, if mating is not random, then the spouse resemblance may reflect either genetic or environmental causes, depending on the source [i.e., through similar genes or cohabitation (environmental) effects]. Finally, there are significant sex differences in the magnitude of the familial cross-trait correlations involving parents, but not offspring, suggesting complex generation (i.e., age) and sex effects. For example, genes may turn on or off as a function of age and sex, and/or there may be an accumulation over time of effects due to the environment which may vary by sex. Whether the common familial factors are genetic (major and/or polygenic), environmental, or some combination of both, and whether the familial expression depends on sex and/or age warrants further investigation using more complex models.     

Adaptive secondary sex ratio adjustments via sex-specific infanticide in a bird

Infanticide is easiest to understand when it involves killing the offspring of others [1], but a parent may also kill its own offspring if the sacrifice of currently dependent young leads to higher survival of brood mates [2] or an improvement in the parent's likely future reproduction [3]. However, sex-specific infanticide by parents of their own offspring, although occurring in some human societies [4], is rare across species. Its rarity may be because killing one sex combines wasted parental effort with consequent biases in population sex ratios that are detrimental for the fitness of the overproduced sex [5-7]. We show that killing male offspring can be advantageous to Eclectus parrot (Eclectus roratus) mothers even though frequency-dependent selection then elevates the reproductive value of sons above that of daughters. In poorer-quality nest hollows, broods with a single female nestling had higher reproductive value than broods in which the female had a younger brother. Our data demonstrate frequent targeted removal of male nestlings within 3 days of hatching in these specific brood types and nesting conditions. The ability of Eclectus parrots to perceive the sex of their offspring relatively early may favor decisions to kill one sex before further investment in parental care.