Ectopically expressed MSL2 females compared with normal females
ABSTRACT: We used H83M2 P element to ectopically expressed MSL2 protein in females to test if the novel formed MSL complex is the directly reason of dosage compensation Compare the MSL2 females with normal females and normal males; MSL2 males also included.
Project description:Linking the evolution of the phenotype to the underlying genotype is a key aim of evolutionary genetics and is crucial to our understanding of how natural selection shapes a trait. Here we consider the genetic basis of sex allocation behaviour in the parasitoid wasp Nasonia vitripennis using a transcriptomics approach. Females allocate offspring sex in line with Local Mate Competition (LMC) theory. Female-biased sex ratios are produced when one or few females lay eggs on a patch. As the number of females contributing offspring to a patch increases, less female-biased sex ratios are favoured. We contrasted the transcriptomic responses of females as they oviposit under conditions known to influence sex allocation: foundress number (a social cue) and the state of the host (parasitised or not). We found, that when females encounter other females on a patch, or assess host quality with their ovipositors, the resulting changes in sex allocation is not associated with significant changes in whole-body gene expression. We also found that the gene expression changes produced by females, as they facultatively allocate sex in response to a host cue and a social cue, are very closely correlated. We expanded the list of candidate genes associated with oviposition behaviour in Nasonia, some of which may be involved in fundamental processes underlying the ability to facultatively allocate sex, including sperm storage and utilisation. 2 x 3 factorial design. Females were placed into 1 of 2 "foundress number" groups: 1) alone or 2) in the presence of 9 other females (co-foundresses). Females were further subdivided into host treatment groups: i) given no host, ii) given a fresh host and iii) given a pre-parasitised host. This gives a total of 6 possible treatment combinations. For each of these 6 groups, 7 pools of 10 females were sequenced giving 42 libraries altogether.
Project description:Dosage compensation was referred as an equalized X chromosome gene expression between males and females in Drosophila. And inverse dosage effects, produced by genomic imbalance, are believed to account for this modulated expression. Here we made a global expression comparison of trisomy 2L with on extra copy of chromosome 2 long arm to normal diploid with two copies of 2L with high throughput RNA-sequencing. We want to test how about the gene expression pattern changes in those comparisons, including the genes on varied chromosome 2 long arm, some other autosomal genes except chromosome 2L and X chromosome genes. Dosage compensation with an expression level similar to normal diploid and inverse dosage effects should be detected. Comapare the global expression of trisomy 2L samples with the normal diploids Collected the females and males from trisomy 2L and Cantons and performed RNA-seq
Project description:Upstream of N-ras (UNR) is a conserved RNA-binding protein that regulates mRNA translation and stability by binding to sites generally located in untranslated regions (UTRs). In Drosophila, sex-specific binding of UNR to msl2 mRNA and the non-coding RNA roX plays key roles in the control of X-chromosome dosage compensation in both sexes. In order to investigate broader sex-specific functions of UNR, we have identified its RNA targets in adult male and female flies by high-throughput RNA binding and transcriptome analysis. Here we show that UNR binds to a large set of protein-coding transcripts and to a smaller set of non-coding RNAs in a sex-specific fashion. Two replicates of UNR IP were performed in D.melanogaster adult males and females, and enrichment in either sex was compared with IgG IP as control. To correlate sex-specific UNR binding with sex-specific transcription and splicing we performed RNA-Seq experiments in males and females.
Project description:The effect of ectopic expression of male specific lethal 2 (msl2) on chromatin modification and gene expression was studied in Drosophila diploid females and metafemales (3X;2A). Results show that ectopic expression of MSL2 in transgenic msl2 females and metafemales sequesters the MOF histone acetylase to the X, which occurs concordantly with an increase of histone acetylation. Gene expression studies indicate that the X-linked genes are not affected by direct targeting of the MSL complex and the resulting increased H4Lys16 acetylation on the X chromosomes, suggesting one function of the MSL complex is to nullify the effect of a high level of histone acetylation. These results are not consistent with the hypothesis that the presence of the MSL complex conditions a two-fold upregulation. Autosomal gene expression is generally decreased in ectopically expressed MSL2 females, which correlates with the reduced autosomal histone acetylation. Metafemales show dosage compensation of X-linked genes with some autosomal reductions in expression. Interestingly, in metafemales with ectopically expressed MSL2, the autosomal expression is returned to a more normal level. There is a lower autosomal level of histone acetylation compared to the normal metafemales, suggesting a nullifying effect on the negative dosage effect of the X chromosome as previously hypothesized to occur in normal males.
Project description:Sperm samples were extracted from adult A. aegypti male seminal vesicles. Semen samples were extracted from bursa of recently mated bursa of N15 labeled females. The goal was to differentiate between the contribution of seminal fluid proteins and sperm proteins in the semen transfered to the female. Our results yield insights into the molecular function, genome organization, regulation, and evolution of sperm proteins and SFPs in this important disease vector.
Project description:In previous studies we have shown that the two adult females morphs of S. ratti have very different lifespans. This experiment was designed to try to identify differentially expressed genes in these two adult morphs that may account for these differing lifespans. The genes expressed by S. ratti parasitic females at day 6 p.i. were compared to the genes expressed by S. ratti free living females at 3 days 19 degrees C. This comparison was done using a microarray chip that is spotted with PCR fragments from the libraries that were generated from parasitic females extracted at day 6 and day 15 p.i., and a microarray chip that is spotted with PCR fragments from the libraries that were generated from free-living larval stages L1, L2 and infective L3s and from free-living males and females.
Project description:The male-specific lethal (MSL) protein-RNA complex is required for X chromosome dosage compensation in Drosophila melanogaster. The MSL2 and MSL1 proteins form a complex and are essential for X chromosome binding. In addition, the MSL complex must integrate at least one of the noncoding roX RNAs for normal X chromosome binding. Here we find the amino-terminal RING finger domain of MSL2 binds as a complex with MSL1 to the heterochromatic chromocenter and a few sites on the chromosome arms. This binding required the same amino-terminal basic motif of MSL1 previously shown to be essential for binding to high-affinity sites on the X chromosome. While the RING finger domain of MSL2 is sufficient to increase the expression of roX1 in females, activation of roX2 requires motifs in the carboxyl-terminal domain. Binding to hundreds of sites on the X chromosome and efficient incorporation of the roX RNAs into the MSL complex require proline-rich and basic motifs in the carboxyl-terminal domain of MSL2. We suggest that incorporation of the roX RNAs into the MSL complex alters the binding specificity of the chromatin-binding module formed by the amino-terminal domains of MSL1 and MSL2.
Project description:Dosage compensation is achieved in male Drosophila by a twofold up-regulation of the single X chromosome to reach the level of the two X chromosomes in females. A popular hypothesis to explain this phenomenon is that the male-specific lethal (MSL) complex, which is present at high levels on the male X, mediates this modulation of gene expression. One member of the complex, MOF, a histone acetyltransferase, acetylates lysine 16 of histone H4 and another, MSL2, which is only expressed in males, triggers its assembly. Here, we find that when a GAL4-MOF fusion protein is targeted to an upstream-activating sequence linked to a miniwhite reporter, up-regulation occurs in females but down-regulation in males, even though in the latter the whole MSL complex is recruited to the reporter genes and produces an increased histone acetylation. The expression of a GAL4-MSL2 fusion protein does not cause dosage compensation of X and autosomal reporters in females, although its expression causes the organization of the MSL complex on the reporter genes, leading to increased histone acetylation. RNAseq analysis of global endogenous gene expression in females with ectopic expression of MSL2 to coat the X chromosomes shows no evidence of increased expression compared with normal females. These data from multiple approaches indicate that the MSL complex does not mediate dosage compensation directly, but rather its activity overrides the high level of histone acetylation and counteracts the potential overexpression of X-linked genes to achieve the proper twofold up-regulation in males.
Project description:Mating triggers physiological and behavioral changes in females. To understand how females effect these changes, we used microarray,; to characterize gene expression in oviducts of mated and unmated Drosophila females. The transition from nonegg laying to egg laying elicits a distinct molecular profile in the oviduct. Experiment Overall Design: 3 microarrays (3 biologicl repeats) of RNA from oviducts of unmated females (control) and 3 microarrays (3 biological repeats) of RNA from oviducts of mated females