Errors in chromosome segregation during oogenesis and early embryogenesis

Errors in chromosome segregation occurring during human oogenesis and early embryogenesis are very common. Meiotic chromosome development during oogenesis is subdivided into three distinct phases. The crucial events, including meiotic chromosome pairing and recombination, take place from around 11 weeks until birth. Oogenesis is then arrested until ovulation, when the first meiotic division takes place, with the second meiotic division not completed until after fertilization. It is generally accepted that most aneuploid fetal conditions, such as trisomy 21 Down syndrome, are due to maternal chromosome segregation errors. The underlying reasons are not yet fully understood. It is also clear that superimposed on the maternal meiotic chromosome segregation errors, there are a large number of mitotic errors taking place post-zygotically during the first few cell divisions in the embryo. In this chapter, we summarise current knowledge of errors in chromosome segregation during oogenesis and early embryogenesis, with special reference to the clinical implications for successful assisted reproduction

Systems developmental biology: the use of ontologies in annotating models and in identifying gene function within and across species

Systems developmental biology is an approach to the study of embryogenesis that attempts to analyze complex developmental processes through integrating the roles of their molecular, cellular, and tissue participants within a computational framework. This article discusses ways of annotating these participants using standard terms and IDs now available in public ontologies (these are areas of hierarchical knowledge formalized to be computationally accessible) for tissues, cells, and processes. Such annotations bring two types of benefit. The first comes from using standard terms: This allows linkage to other resources that use them (e.g., GXD, the gene-expression [G-E] database for mouse development). The second comes from the annotation procedure itself: This can lead to the identification of common processes that are used in very different and apparently unrelated events, even in other organisms. One implication of this is the potential for identifying the genes underpinning common developmental processes in different tissues through Boolean analysis of their G-E profiles. While it is easiest to do this for single organisms, the approach is extendable to analyzing similar processes in different organisms. Although the full computational infrastructure for such an analysis has yet to be put in place, two examples are briefly considered as illustration. First, the early development of the mouse urogenital system shows how a line of development can be graphically formalized using ontologies. Second, Boolean analysis of the G-E profiles of the mesenchyme-to-epithelium transitions that take place during mouse development suggest Lhx1, Foxc1, and Meox1 as candidate transcription factors for mediating this process

The MetaCyc database of metabolic pathways and enzymes and the BioCyc collection of pathway/genome databases

The MetaCyc database (http://metacyc.org/) provides a comprehensive and freely accessible resource for metabolic pathways and enzymes from all domains of life. The pathways in MetaCyc are experimentally determined, small-molecule metabolic pathways and are curated from the primary scientific literature. MetaCyc contains more than 1800 pathways derived from more than 30 000 publications, and is the largest curated collection of metabolic pathways currently available. Most reactions in MetaCyc pathways are linked to one or more well-characterized enzymes, and both pathways and enzymes are annotated with reviews, evidence codes and literature citations. BioCyc (http://biocyc.org/) is a collection of more than 1700 organism-specific Pathway/Genome Databases (PGDBs). Each BioCyc PGDB contains the full genome and predicted metabolic network of one organism. The network, which is predicted by the Pathway Tools software using MetaCyc as a reference database, consists of metabolites, enzymes, reactions and metabolic pathways. BioCyc PGDBs contain additional features, including predicted operons, transport systems and pathway-hole fillers. The BioCyc website and Pathway Tools software offer many tools for querying and analysis of PGDBs, including Omics Viewers and comparative analysis. New developments include a zoomable web interface for diagrams; flux-balance analysis model generation from PGDBs; web services; and a new tool called Web Groups

Subcellular Distribution of Mitochondrial Ribosomal RNA in the Mouse Oocyte and Zygote

Mitochondrial ribosomal RNAs (mtrRNAs) have been reported to translocate extra-mitochondrially and localize to the germ cell determinant of oocytes and zygotes in some metazoa except mammals. To address whether the mtrRNAs also localize in the mammals, expression and distribution of mitochondrion-encoded RNAs in the mouse oocytes and zygotes was examined by whole-mount in situ hybridization (ISH). Both 12S and 16S rRNAs were predominantly distributed in the animal hemisphere of the mature oocyte. This distribution pattern was rearranged toward the second polar body in zygotes after fertilization. The amount of mtrRNAs decreased around first cleavage, remained low during second cleavage and increased after third cleavage. Staining intensity of the 12S rRNA was weaker than that of the 16S rRNA throughout the examined stages. Similar distribution dynamics of the 16S rRNA was observed in strontium-activated haploid parthenotes, suggesting the distribution rearrangement does not require a component from sperm. The distribution of 16S rRNAs did not coincide with that of mitochondrion-specific heat shock protein 70, suggesting that the mtrRNA is translocated from mitochondria. The ISH-scanning electron microscopy confirms the extra-mitochondrial mtrRNA in the mouse oocyte. Chloramphenicol (CP) treatment of late pronuclear stage zygotes perturbed first cleavage as judged by the greater than normal disparity in size of blastomeres of 2-cell conceptuses. Two-third of the CP-treated zygotes arrested at either 2-cell or 3-cell stage even after the CP was washed out. These findings indicate that the extra-mitochondrial mtrRNAs are localized in the mouse oocyte and implicated in correct cytoplasmic segregation into blastomeres through cleavages of the zygote

Does Selection against Transcriptional Interference Shape Retroelement-Free Regions in Mammalian Genomes?

BACKGROUND: Eukaryotic genomes are scattered with retroelements that proliferate through retrotransposition. Although retroelements make up around 40 percent of the human genome, large regions are found to be completely devoid of retroelements. This has been hypothesised to be a result of genomic regions being intolerant to insertions of retroelements. The inadvertent transcriptional activity of retroelements may affect neighbouring genes, which in turn could be detrimental to an organism. We speculate that such retroelement transcription, or transcriptional interference, is a contributing factor in generating and maintaining retroelement-free regions in the human genome. METHODOLOGY/PRINCIPAL FINDINGS: Based on the known transcriptional properties of retroelements, we expect long interspersed elements (LINEs) to be able to display a high degree of transcriptional interference. In contrast, we expect short interspersed elements (SINEs) to display very low levels of transcriptional interference. We find that genomic regions devoid of long interspersed elements (LINEs) are enriched for protein-coding genes, but that this is not the case for regions devoid of short interspersed elements (SINEs). This is expected if genes are subject to selection against transcriptional interference. We do not find microRNAs to be associated with genomic regions devoid of either SINEs or LINEs. We further observe an increased relative activity of genes overlapping LINE-free regions during early embryogenesis, where activity of LINEs has been identified previously. CONCLUSIONS/SIGNIFICANCE: Our observations are consistent with the notion that selection against transcriptional interference has contributed to the maintenance and/or generation of retroelement-free regions in the human genome

The MetaCyc database of metabolic pathways and enzymes and the BioCyc collection of pathway/genome databases

The MetaCyc database (MetaCyc.org) is a comprehensive and freely accessible resource for metabolic pathways and enzymes from all domains of life. The pathways in MetaCyc are experimentally determined, small-molecule metabolic pathways and are curated from the primary scientific literature. With more than 1400 pathways, MetaCyc is the largest collection of metabolic pathways currently available. Pathways reactions are linked to one or more well-characterized enzymes, and both pathways and enzymes are annotated with reviews, evidence codes, and literature citations. BioCyc (BioCyc.org) is a collection of more than 500 organism-specific Pathway/Genome Databases (PGDBs). Each BioCyc PGDB contains the full genome and predicted metabolic network of one organism. The network, which is predicted by the Pathway Tools software using MetaCyc as a reference, consists of metabolites, enzymes, reactions and metabolic pathways. BioCyc PGDBs also contain additional features, such as predicted operons, transport systems, and pathway hole-fillers. The BioCyc Web site offers several tools for the analysis of the PGDBs, including Omics Viewers that enable visualization of omics datasets on two different genome-scale diagrams and tools for comparative analysis. The BioCyc PGDBs generated by SRI are offered for adoption by any party interested in curation of metabolic, regulatory, and genome-related information about an organism

Nlrp2, a Maternal Effect Gene Required for Early Embryonic Development in the Mouse

Maternal effect genes encode proteins that are produced during oogenesis and play an essential role during early embryogenesis. Genetic ablation of such genes in oocytes can result in female subfertility or infertility. Here we report a newly identified maternal effect gene, Nlrp2, which plays a role in early embryogenesis in the mouse. Nlrp2 mRNAs and their proteins (∼118 KDa) are expressed in oocytes and granulosa cells during folliculogenesis. The transcripts show a striking decline in early preimplantation embryos before zygotic genome activation, but the proteins remain present through to the blastocyst stage. Immunogold electron microscopy revealed that the NLRP2 protein is located in the cytoplasm, nucleus and close to nuclear pores in the oocytes, as well as in the surrounding granulosa cells. Using RNA interference, we knocked down Nlrp2 transcription specifically in mouse germinal vesicle oocytes. The knockdown oocytes could progress through the metaphase of meiosis I and emit the first polar body. However, the development of parthenogenetic embryos derived from Nlrp2 knockdown oocytes mainly blocked at the 2-cell stage. The maternal depletion of Nlrp2 in zygotes led to early embryonic arrest. In addition, overexpression of Nlrp2 in zygotes appears to lead to normal development, but increases blastomere apoptosis in blastocysts. These results provide the first evidence that Nlrp2 is a member of the mammalian maternal effect genes and required for early embryonic development in the mouse

miR-135A Regulates Preimplantation Embryo Development through Down-Regulation of E3 Ubiquitin Ligase Seven in Absentia Homolog 1A (SIAH1A) Expression

Background: MicroRNAs (miRNAs) are small non-coding RNA molecules capable of regulating transcription and translation. Previously, a cluster of miRNAs that are specifically expressed in mouse zygotes but not in oocytes or other preimplantation stages embryos are identified by multiplex real-time polymerase chain reaction-based miRNA profiling. The functional role of one of these zygote-specific miRNAs, miR-135a, in preimplantation embryo development was investigated. Methodology/Principal Findings: Microinjection of miR-135a inhibitor suppressed first cell cleavage in more than 30% of the zygotes. Bioinformatics analysis identified E3 Ubiquitin Ligase Seven In Absentia Homolog 1A (Siah1a) as a predicted target of miR-135a. Western blotting and 3′UTR luciferase functional assays demonstrated that miR-135a down-regulated the expression of Siah1 in HeLa cells and in mouse zygotes. Siah1a was expressed in preimplantation embryos and its expression pattern negatively correlated with that of miR-135a. Co-injection of Siah1a-specific antibody with miR-135a inhibitor partially nullified the effect of miR-135a inhibition. Proteasome inhibition by MG-132 revealed that miR-135a regulated proteasomal degradation and potentially controlled the expression of chemokinesin DNA binding protein (Kid). Conclusions/Significance: The present study demonstrated for the first time that zygotic specific miRNA modulates the first cell cleavage through regulating expression of Siah1a. © 2011 Pang et al.published_or_final_versio

Cortical Mechanics and Meiosis II Completion in Mammalian Oocytes Are Mediated by Myosin-II and Ezrin-Radixin-Moesin (ERM) Proteins

Analysis of mouse oocyte mechanics shows that effective tension drops 6-fold from prophase I to metaphase II; the metaphase II egg has a 2.5-fold tension differential between the cortex over the spindle and the opposite cortex. Manipulation of actin, myosin-II, or ERMs alters tension levels and induces spindle abnormalities during meiosis II

ADAM2 Interactions with Mouse Eggs and Cell Lines Expressing α4/α9 (ITGA4/ITGA9) Integrins: Implications for Integrin-Based Adhesion and Fertilization

Integrins are heterodimeric cell adhesion molecules, with 18 α (ITGA) and eight β (ITGB) subunits forming 24 heterodimers classified into five families. Certain integrins, especially the α(4)/α(9) (ITGA4/ITGA9) family, interact with members of the ADAM (a disintegrin and metalloprotease) family. ADAM2 is among the better characterized and also of interest because of its role in sperm function. Having shown that ITGA9 on mouse eggs participates in mouse sperm-egg interactions, we sought to characterize ITGA4/ITGA9-ADAM2 interactions.An anti-β(1)/ITGB1 function-blocking antibody that reduces sperm-egg binding significantly inhibited ADAM2 binding to mouse eggs. Analysis of integrin subunit expression indicates that mouse eggs could express at least ten different integrins, five in the RGD-binding family, two in the laminin-binding family, two in the collagen-binding family, and ITGA9-ITGB1. Adhesion assays to characterize ADAM2 interactions with ITGA4/ITGA9 family members produced the surprising result that RPMI 8866 cell adhesion to ADAM2 was inhibited by an anti-ITGA9 antibody, noteworthy because ITGA9 has only been reported to dimerize with ITGB1, and RPMI 8866 cells lack detectable ITGB1. Antibody and siRNA studies demonstrate that ITGB7 is the β subunit contributing to RPMI 8866 adhesion to ADAM2.These data indicate that a novel integrin α-β combination, ITGA9-ITGB7 (α(9)β(7)), in RPMI 8866 cells functions as a binding partner for ADAM2. ITGA9 had previously only been reported to dimerize with ITGB1. Although ITGA9-ITGB7 is unlikely to be a widely expressed integrin and appears to be the result of "compensatory dimerization" occurring in the context of little/no ITGB1 expression, the data indicate that ITGA9-ITGB7 functions as an ADAM binding partner in certain cellular contexts, with implications for mammalian fertilization and integrin function