Project description:The Ca2+ oscillations initiated by the fertilizing sperm (but terminating concomitant with pronucleus formation) apparently ensure that the events constituting egg activation occur in the correct temporal order; early events (e.g., cortical granule exocytosis) require fewer oscillations than later events (e.g., recruitment of maternal mRNA). Whether the Ca2+ signaling events impact long-term development, in particular development to term, is unknown. Using fertilized eggs that have undergone the first few Ca2+ oscillations, we developed procedures that result either in inhibiting or stimulating the natural pattern of Ca2+ signaling of inseminated eggs. Although the incidence of development to the blastocyst stage is unaltered by these procedures, fewer offspring are born following embryo transfer, indicating that developmental competence of the blastocysts is reduced. Interestingly, embryo transfer experiments reveal that when the natural regime of Ca2+ oscillations is precociously interrupted, the incidence of implantation is compromised whereas hyper-stimulation of Ca2+ signaling events compromises post-implantation development. Moreover, although there was no major difference in the overall growth rates of the offspring, those obtained following hyper-stimulation exhibited a far greater variability in their weight. Analysis of global patterns of gene expression by microarray analysis revealed that approximately 20% of the transcripts are mis-regulated when too few oscillations are experienced by the embryo and EASE analysis indicates that genes preferentially involved in RNA processing and polymerase II transcription are differentially affected. In addition, a set of genes involved in cell adhesion is also mis-expressed and could thus be mechanistically linked to the observed reduced implantation. Only about 3% of the transcripts were mis-regulated following hyper-stimulation, and EASE analysis indicates that genes preferentially involved in metabolism are differentially affected. In toto, these results indicate that a range Ca2+ signaling events following fertilization (an excess or reduction) has long-term effects on both gene expression and development to term. Experiment Overall Design: We profiled the global gene expression in the blastocysts by treatment of Ca2+ oscillations, and identified the genes differentially expressed.
Project description:The Ca2+ oscillations initiated by the fertilizing sperm (but terminating concomitant with pronucleus formation) apparently ensure that the events constituting egg activation occur in the correct temporal order; early events (e.g., cortical granule exocytosis) require fewer oscillations than later events (e.g., recruitment of maternal mRNA). Whether the Ca2+ signaling events impact long-term development, in particular development to term, is unknown. Using fertilized eggs that have undergone the first few Ca2+ oscillations, we developed procedures that result either in inhibiting or stimulating the natural pattern of Ca2+ signaling of inseminated eggs. Although the incidence of development to the blastocyst stage is unaltered by these procedures, fewer offspring are born following embryo transfer, indicating that developmental competence of the blastocysts is reduced. Interestingly, embryo transfer experiments reveal that when the natural regime of Ca2+ oscillations is precociously interrupted, the incidence of implantation is compromised whereas hyper-stimulation of Ca2+ signaling events compromises post-implantation development. Moreover, although there was no major difference in the overall growth rates of the offspring, those obtained following hyper-stimulation exhibited a far greater variability in their weight. Analysis of global patterns of gene expression by microarray analysis revealed that approximately 20% of the transcripts are mis-regulated when too few oscillations are experienced by the embryo and EASE analysis indicates that genes preferentially involved in RNA processing and polymerase II transcription are differentially affected. In addition, a set of genes involved in cell adhesion is also mis-expressed and could thus be mechanistically linked to the observed reduced implantation. Only about 3% of the transcripts were mis-regulated following hyper-stimulation, and EASE analysis indicates that genes preferentially involved in metabolism are differentially affected. In toto, these results indicate that a range Ca2+ signaling events following fertilization (an excess or reduction) has long-term effects on both gene expression and development to term.
Project description:In our study, differential male nucleus events and development behaviors were revealed from the fertilized eggs in response to the sperm from males of genotypic sex determination (GSD) and temperature-dependent sex determination (TSD) in gibel carp. When the eggs of maternal fish were fertilized by the sperm from males of GSD, the fertilized egg encountered similar sexual reproduction events and behaviors. However, when the eggs of maternal fish were fertilized by the sperm from males of TSD, a typical process of gynogenesis was observed. To reveal the underlying molecular mechanism of differential sperm nucleus development behaviors in the fertilized eggs, iTRAQ-based quantitative semen proteomics were performed on three semen samples from three males of GSD and three semen samples from three males of TSD respectively.
Project description:A stable supply of viable eggs and embryos is crucial for successful farming of Atlantic cod. Broodstock stress can have negative effects, but little is known about the molecular mechanisms that cause abnormal development. Maternally transferred mRNAs have been shown to be essential for normal development, and stress may therefore influence their expression and the subsequent embryonic development. We investigated if mimicked stress in prespawning Atlantic cod females affects mRNA concentrations in eggs and embryos, and if this can be linked to egg/embryo viability, and if we could identify potential molecular markers for stress and egg/embryo viability. Consequently, 3 weeks prior to expected peak spawning, 20 females were intraperitoneally implanted with either cortisol-containing or cortisol-free (sham) osmotic pumps. At peak spawning all individuals were stripped and eggs were fertilized and incubated until hatching. Samples were collected from unfertilized eggs and embryos for gene expression profiling.
Project description:In vitro culture of preimplantation mouse embryos is associated with changes in gene expression. It is however not known if the method of fertilization affects the global pattern of gene expression. We compared gene expression and development of mouse blastocysts produced in vitro (IVF), vs. blastocysts fertilized in vivo and cultured in vitro from the zygote stage (IVC), vs. control blastocysts flushed out of the uterus on post coital day 3.5. The global pattern of gene expression was assessed using the Affymetrix 430 2.0 chip.
Project description:Calcium is an essential macronutrient and plant requires it in large amounts for normal growth and development. This ion participated in innumerable processes and affects nearly all aspects of plant growth and development such as signal transduction, metabolism of lipids, proteins, and carbohydrates, cell growth, cell wall and membrane stabilization. We used whole genome microarrays to determine the transcriptomic profile of rice seedlings exposed to short-term and long term Ca2+ deficiency followed by Ca2+ resupply.
Project description:Calcium is an essential macronutrient and plant requires it in large amounts for normal growth and development. This ion participated in innumerable processes and affects nearly all aspects of plant growth and development such as signal transduction, metabolism of lipids, proteins, and carbohydrates, cell growth, cell wall and membrane stabilization. We used whole genome microarrays to determine the transcriptomic profile of rice seedlings exposed to short-term and long term Ca2+ deficiency followed by Ca2+ resupply. Calcium treated seedlings were used for for RNA extraction and hybridization on Agilent microarray platform. Three biological replicates of each sample were used for microarray analysis. We wanted to know the altered expression patterns of calcium-responsive genes majorly involved in metabolic processes, signal transdction pathways, transcriptional regulation, and transport of multiple molecules including Ca2+. Seeds of indica rice were surface sterilized and grown hydroponically in rice growing medium. Plants were grown in the rice growth room under the condition of 16 h light/8 h dark (28M-BM-0C) photoperiod with 70 % humidity. After 5 days of normal growth, some of the seedlings were transferred to nutrient solution lacking CaCl2 (-Ca2+). After 5 and 14 days, calcium (0.798 mM CaCl2) was resupplied to the plants grown in -Ca2+ medium for 6 h.
Project description:Using a tiled whole-genome microarray, we found that 58.2% of Tribolium castaneum genes are maternally loaded into eggs. Comparison of known Drosophila melanogaster maternal genes to our results showed widespread conservation of maternal function with T. castaneum. We also found many T. castaneum genes with previously identified gender or tissue specific expression were also maternally loaded into eggs. The microarray design also allowed the detection of 2315 and 4060 novel transcriptionally active regions greater in length than 100 bp in unfertilized and fertilized T. castaneum eggs, respectively. The primary objective of this study was to identify expressed regions of the Tribolium castaneum genome in unfertilized and fertilized eggs using a whole-genome tiled microarray.
Project description:Using a tiled whole-genome microarray, we found that 58.2% of Tribolium castaneum genes are maternally loaded into eggs. Comparison of known Drosophila melanogaster maternal genes to our results showed widespread conservation of maternal function with T. castaneum. We also found many T. castaneum genes with previously identified gender or tissue specific expression were also maternally loaded into eggs. The microarray design also allowed the detection of 2315 and 4060 novel transcriptionally active regions greater in length than 100 bp in unfertilized and fertilized T. castaneum eggs, respectively. The primary objective of this study was to identify expressed regions of the Tribolium castaneum genome in unfertilized and fertilized eggs using a whole-genome tiled microarray. The whole RNA of 3 samples of virgin laid eggs and 3 samples of fertilized eggs were compaired.