Teruko Taketo
Professor, Department of Surgey, McGill University
Associate Member, Department of Biology, McGill University
Associate Member, Department of Obstetrics and Gynecology, McGill University
   

Contact Information:
RI - MUHC
Room EM03220
1001 Decarie Blvd
687 Pine Avenue, West
Montreal, Quebec
Canada H4A 3J1

Tel.: (514)-934-1934 ext. -34197
e-mail: teruko.taketo@mcgill.ca

RESEARCH


1. Surveillance mechanism for the elimination of oocytes during the meiotic prophase progression in the mouse ovary

Primordial germ cells are specified at an extra-embryonic site and migrate into a gonadal primordium that is undergoing early-phase sexual differentiation. Subsequent sexual differentiation of germ cells is regulated by their gonadal environment in testis or ovary. In the fetal ovary, most, if not all, germ cells exit the proliferation phase and enter the meiotic prophase to become primary oocytes. Thus, the oocyte population becomes finite and eventually limits female reproductive life. In addition, more than half of the initial oocyte population is eliminated during the meiotic prophase progression, further restricting the oocyte reserve. The cause or mechanism of this major oocyte loss in normal ovaries is poorly understood. During the meiotic prophase paternal and maternal homologous chromosomes pair, synapse, and recombine. A failure in meiotic chromosome synapsis (asynapsis) or recombination causes aberrant chromosome segregation during meiotic divisions, leading to aneuploidy in embryos. Aneuploidy is the major cause of miscarriage, stillbirth, and the birth of children with chromosomal anomalies. We hypothesize that persistent meiotic chromosome asynapsis initiates a cascade of cellular events that result in elimination of the oocytes carrying unsynapsed chromosomes, thus minimizing the risk of aneuploidy in embryos. We have established a new method for analyzing the number and chromosomal structures in individual oocytes dissociated from mouse ovaries (named microspread ovarian cells). By using this method, we have shown that the mitochondrial apoptotic pathway mediated by caspase 9 is essential for oocyte elimination; however, caspase 9 is constitutively activated in most oocytes, suggesting that additional mechanisms are involved in the regulation of oocyte elimination. Our current specific aims are; (1) to clarify the regulatory mechanism of the caspase 9-mediated apoptotic pathway responsible for oocyte demise; (2) to examine whether persistent chromosome asynapsis leads to silencing of the unsynapsed chromatin region, which in turn causes the caspase 9-dependent oocyte demise; and (3) to determine whether oocytes with synaptic errors are selectively eliminated during the meiotic prophase progression. The results of these studies will clarify the role of oocyte elimination during normal ovarian development and may offer insight into the genetic mechanisms that underlie an increased risk of aneuploidy. (Supported by NSERC and CIHR grants)

Fig. 1. Pairing between homologous chromosomes at the pachytene stage of meiotic prophase is shown in microspread ovarian cells. (a) XX ovary. Synaptonemal complex (SC) (red) is formed between 20 sets of homologous chromosomes including the X-X pair (not identified). (b) XY ovary. The single Y-chromosome (arrow) is left alone while the single X-chromosome with fainter SC staining (arrowhead) is tangled with autosomal pairs. (c) XO ovary. The single X-chromosome (arrowhead) is left alone while 19 sets of autosomes are completely paired. The partial SC staining on the X-chromosome may indicate self-pairing. (Alton et al., 2008)

 

Selected Publications (Taketo’s laboratory members in bold face)

  1. McClellan, K.A., Gosden, R., Taketo, T. (2003) Continuous loss of oocytes throughout meiotic prophase in the normal mouse ovary. Developmental Biology 258: 334-348.
  2. Alton, M., Taketo, T. (2007) Switch from BAX-dependent to BAX-independent germ cell loss during the development of fetal mouse ovaries. Journal of Cell Science 120: 417-424.
  3. Alton, M., Lau, M. P., Villemure, M., Taketo, T. (2008) The behavior of the X- and Y-chromosomes in the oocyte during meiotic prophase in the B6.YTIR sex-reversed mouse ovary. Reproduction 135: 241-252.
  4. Taketo, T. (2012) Microspread oocyte preparations for the analysis of meiotic prophase progression with improved recovery by cytospin centrifugation. Methods in Molecular Biology 825: 173-181.
  5. Ene, A.C., Park, S., Edelmann, W., Taketo, T. (2013) Caspase 9 is constitutively activated in mouse oocytes and plays a key role in oocytes elimination during meiotic prophase progression. Developmental Biology 377: 213-223.
  6. Taketo, T., Naumova, A.K. (2013) Oocyte heterogeneity with respect to the meiotic silencing of unsynapsed X chromosomes in the XY female mouse. Chromosoma 122: 337-349.


2. Sex reversal and infertility in the B6.YTIR female mouse – a model for studying the regulation of sister chromatid separation in fertilized eggs

In mammalian development, gonadal sex, either testis or ovary, is determined by the presence or absence of SRY expression, which is encoded on the Y chromosome. Subsequently, germ cells undergo sexual differentiation dependent on the gonadal environment. Therefore, spermatogenesis takes place in the presence of X and Y chromosomes, and oogenesis in the presence of two X chromosomes. However, gonadal sex reversal does occur in humans as well as in other mammalian species, and, while the resultant individuals can lead healthy lives, they become infertile or subfertile. Male germ cells carrying an abnormal set of sex chromosomes are efficiently eliminated during spermatogenesis by multilayered surveillance mechanisms in most mammalian species. By contrast, female germ cells carrying an abnormal set of sex chromosomes cause infertility in humans (except for those individuals carrying trisomy-X) but variable fertility states in mice. The sexual dimorphic consequences of genetic defects in the germ cells are not fully understood.

We have previously reported that gonadal primordia of the B6.YTIR mouse develop into ovaries, in which germ cells enter meiosis and go through the meiotic prophase, despite the presence and expression of Sry. The X and Y chromosomes do not pair in most XY oocytes during the first meiotic prophase, unlike in XY spermatocytes. Subsequently, a greater number of XY oocytes are eliminated by the end of meiotic prophase, compared to XX oocytes, which can be attributed to a surveillance mechanism of chromosome asynapsis (described above). Nonetheless, a considerable number of XY oocytes survive to complete the first meiotic prophase and form follicles. When fully-grown oocytes are collected from antral follicles and subjected to meiotic resumption and maturation in vitro, most of them reach the second meiotic metaphase (MII) but very few reach the 2-cell stage after fertilization, thus diminishing their reproductive capacity. We have observed that the MII-spindle is not properly assembled in most oocytes of XY females. Their sister-chromatids separate but often fail to be extruded with the second polar body after oocyte activation or fertilization. Thus, the B6.YTIR mouse model provides an opportunity for studying the mechanism of sister chromatid separation and its coordination with the second meiotic cell division in oocytes.

The developmental incompetence of oocytes from XY females can be attributed to their cytoplasmic defects; when the nuclei of XY oocytes have been transferred into enucleated XX oocytes, the reconstructed oocytes generate healthy offspring after fertilization in vitro and embryo transfer into pseudo-pregnant females. These offspring faithfully inherit the sex chromosomes identified in the MII-oocytes from XY females except for the elimination of YY and YO embryos. It appears that sex chromosome aneuploidy per se does not block oogenesis or embryonic development. We compared gene expression profiles in fully-grown oocytes collected from XX, XO, and XY ovaries. We found that all genes tested show comparable transcript levels between XX and XO oocytes, indicating that mRNA storage levels are regulated despite the difference in X chromosome numbers. By comparison, the transcript levels of many X-linked and autosomal genes are up-regulated or down-regulated in XY oocytes, suggesting that transcriptional regulation processes are altered in XY oocytes. Many of these differentially-expressed genes are included in the gene expression profile of XO oocytes carrying a Y-linked Zfy2 transgene, despite different genetic backgrounds. Differences in mRNA expression profiles may in part differentiate the fertility of XO oocytes and the infertility of XY and XOZfy2 oocytes. We are currently testing the role of several differentially-expressed genes that may be linked to MII-spindle defects and embryonic developmental failure in XY oocytes.


Fig. 2. Cell cycle progression in the oocytes from XX (a-e) and XY (f-k) females after in vitro maturation for 19 h, SrCl2 activation for 2 h, and further culture. Immunolabeling of microtubules (green) and DAPI staining of chromosomes (blue). a. At MII before activation. Meiotic spindle is assembled in a barrel shape on both sides of condensed chromosomes (arrow). b. At AII (2.5 h post-activation). Condensed chromosomes (arrows) are migrated into the two poles of meiotic spindle. c. At TII (4 hpa). A set of condensed chromosomes (arrow) are extruding from the oocyte. d. At PN-stage (4 hpa). One prominent PN (arrow) has been formed. e. At the 2-cell-stage (24 hpa). Two symmetrical blastomeres have been formed after the first cell cleavage. f. At MII (2.5 hpa). Chromosomes are condensed at the second metaphase plate (arrow) encompassed by bulky microspindles. g. At AII (2.5 hpa). Chromosomes are segregating towards the both poles of meitoic spindle but some are bridging between the two groups (arrowhead in inset). h. At TII (2.5 hpa). A group of chromosomes are extruding from the oocyte, but many are bridging between the two groups of chromosomes (in inset). i. At TII- to PN-stage (4 hpa). A pole of meiotic spindle is extruding from the oocyte as if at TII. However, all nuclear materials (arrows) remain within the oocyte. j. At PN-stage (10 hpa). Three PN have been formed (arrows). k. At the 2-cell-stage (24 hpa). Asymmetrical two blastomeres contain multinuclei (arrows). (Villemure et al., 2007).


Selected Publications

  1. Amleh, A., Ledee, N., Saeed, J., Taketo, T. (1996) Competence of XY oocytes from the B6.YDOM sex-reversed female mouse for maturation, fertilization, and embryonic development in vitro. Developmental Biology 178: 263-275.
  2. Amleh, A., Taketo, T. (1998) Live-borns from XX but not XY oocytes in the chimeric ovary composed of B6.YTIR and XX cells. Biology of Reproduction 58: 574-582.
  3. Amleh, A., Smith, L., Chen, H.-Y., Taketo, T. (2000) Both nuclear and cytoplasmic components are defective in oocytes of the B6.YTIR sex-reversed female mouse. Developmental Biology 219: 277-286.
  4. Taketo, T., Lee, C.-H., Zhang, J., Li, Y.-M., Lee, C.-Y.G. and Lau, Y.-F.C. (2005) Expression of SRY proteins in both normal and sex-reversed XY fetal mouse gonads. Developmental Dynamics 233: 612-622.
  5. Villemure, M., Chen, H.-Y., Kurokawa, M., Fissore, R.M., Taketo, T. (2007) The presence of X-and Y-chromosomes in oocytes leads to impairment in the progression of the second meiotic division. Developmental Biology 301: 1-13.
  6. Obata, Y., Villemure, M., Kono, T., Taketo, T. (2008) Transmission of Y-chromosomes from XY female mice was made possible by replacement of cytoplasm during oocyte maturation. Proceedings of the National Academy of Sciences of the United States of America 105: 13918-13923.
  7. Park, S., Zeidan, K.T., Shin, J.S., Taketo, T. (2011) SRY upregulation of SOX9 is inefficient and delayed, allowing ovarian differentiation, in the B6.YTIRgonad. Differentiation 82: 18-27.
  8. Xu, B.-Z., Obata, Y., Cao, F., Taketo T. (2012) The presence of the Y-chromosome, not the absence of the second X-chromosome, alters the mRNAs stored in the fully grown XY mouse oocyte. PLos One 7: e40481.
  9. Xu, B., Noohi, S., Shin, J.S., Tan SL, Taketo T. (2014) Bi-directional communication with cumulus cells involved in the deficiency of XY oocytes in the components essential for proper second meiotic spindle assembly. Developmental Biology 385: 242-252.
  10. Vernet, N., Szot, M., Mahadevaiah, S.K., Ellis, P.J.I., Decarpentrie, F., Ojarikre, O.A., Rattigan, A., Taketo, T., Burgoyne, P.S. (2014) The expression of Y-encoded Zfy2 in XY mouse oocytes triggers subsequent preimplantation embryonic failure and infertility. Development 141: 855-866.


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