Welcome to Komiya's lab.
Our Current Research Projects
1. Germ cell formation and differentiation in Xenopus laevis.
Germ cells are set aside from somatic cells in early embryogenesis. There are largely two modes in which germ cells are specified. One is “inducing” or “epigenesis” mode, in which germ cells are specified by the influence of factors secreted from surrounding cells. Many species such as mouse or human adopt this, and it is considered to be an ancient mode. The other is “determinant” or “preformation” mode, in which germ cells are specified by the inheritance of germ cell determinant that has been asymmetrically localized in the oocytes. Many model animals such as C. elegans, Drosophila, zebrafish and Xenopus adopt this mode. Although the two modes are clearly different in appearance, conserved mechanisms underlying germ cell specification among all animal species has been revealed. One of the conserved characteristic features of germ cells is the transcriptional and translational repression of the genes that are primarily required for somatic cell specification or differentiation. In Drosophila and nematode, pgc and PIE-1, respectively, negatively regulate RNA polymerase II by preventing phosphorylation of the polymerase either directly or in directly by modulating their regulatory proteins, leading to the downregulation of general transcription . And, in mouse, Blimp1, a transcriptional repressor, has been proved to be involved in the repression of Hox genes. Moreover, in addition to Blimp1, nanos, which has been considered to be a translational repressor, interacts with the genes for chromosome remodeling, thus consequently, more higher ordered transcriptional repression is implicated. In fact, in Drosophila, germ cells lacking nanos have shown to trans-differentiate into somatic cells if they are prevented to undergo apoptosis, indicating that proper gene repression is required not only for specification but also for maintaining the state of germ cells. In the animals adopting determinant mode, such important gene products mostly as mRNAs are maternally provided and localized asymmetrically during the oogenesis to form the germ cell determinant, germ plasm. In Xenopus, germ plasm is rich in mitochondria, endoplasmic reticulum and proteins such as xpat . It also contains about 20 species of mRNAs such as xcat2, xpat, DEADSouth, Dead end and XGRIP2, which are indispensable for the specification or differentiation of germ cells. As the Xenopus germ plasm is localized just under the vegetal cortex of the fertilized eggs, the germ plasm-inherited cells, primordial germ cells (PGCs), arise from vegetal blastomeres, and later, actively migrate to the dorsal sides, where future gonads are developed. Most of the germ plasm-localizing mRNAs are transcribed at stage I-II in the oogenesis and transported to the vegetal cortex by way of mitochondrial cloud, the pathway of which was defined as early pathway or METRO with some exceptions for the transporting pathway. The mechanism for this mRNA transportation system to the germ plasm has been investigated from the viewpoint of cis-elements of the mRNAs as well as trans-factors that bind to the cis-elements and transport them. Generally, 3-untranslated region (UTR) of the mRNAs has been known to be responsible for the cis-elements for the transportation and a large number of examinations were performed about the molecular mechanism for germ plasm localization signal and binding factors.
Our findings in this field are as follows:
2. Testis development in mice and Xenopus.
Adult germ cells are specially differentiated cells by which genomic information is passed from generation to generation. Above all, male gametes, sperms, have characteristic morphology and function, which contribute to successfully carry out the fertilization, and are stepwise formed in the seminiferous tubules of the adult testis from spermatogonia, germ-line stem cells, through spermatocytes, spermatids and finally to spermatozoa. During the spermatogenesis steps, many genes are transcribed and translated at proper time and space. During the spermiogenesis, the final stage of the spermatogenesis, spermatids that complete the meiotic division differentiate into spermatozoa with several typical reactions. Firstly, the condensation of chromatin is organized by changing the chromatin binding protein from histons to transition proteins and finally to protamin. Secondly, drastic morphological changes occur by activating proteolytic pathways to induce autophagy, and by recruiting cytoskeleton system to cast useless structures as residual bodies. Thirdly, energy-producing mitochondria are densely packed to the midpiece and a flagellum is constructed to the tail. Fourth, functions required for fertilization; i.e., acrosome for passing zona pellucida, proteins for cell-membrane fusion and protein kinases for activating eggs, are equipped. Obtaining the knowledge of the function of the genes required for the each step is essential to understand the molecular mechanism of spermiogenesis, as well as to find fertility treatment in medical fields.
Previously we reported the development of two high throughput-screening methods, the large-scale immunohistological screening and the in situ hybridization system. Both methods utilize 96-well plates in which tissue sections are held. When performing the in situ hybridization method, all procedures, including template preparation, DIG-RNA-probe synthesis, hybridization and detection were carried out by 96-well format. Using an equalized cDNA library as the source for probes, we could screen various tissue samples fast and efficiently, and subsequently we could clone the cDNAs whose mRNAs were cell-type or region specific.
3. Intestinal development in mice.
#Intesitinal paneth cells: The Paneth cells are considered to be specially differentiated cells against microorganisms in a small intestine. Because it secrets a number of anti-microbe molecules, including alpha-defensins, also known as cryptdins, lysozym M, phospholipase A2 and angiogenin, thereby contributing to maintenance of the gastrointestinal barrier. Thereby, identifying molecules expressed in Paneth cells is a challenging attempt to as the first step for discovering unknown anti-microbe molecules. And further to examine the biological function of the molecules may reveal the underlying mechanism of unknown host-microbe interactions.
4. Systematic identification of the genes expressed in the tissues such as intestine, testis, brain, etc. performed by the large-scale in situ hybridization system that I have developed.
Monitoring the gene expression is crucial to understand the function of the genes that are involved in the normal and abnormal biological phenomena such as morphogenesis, maintenance of tissues, and/or development of malignant tissues. Recently, the techniques such as differential hybridization method using the DNA microarray or qualitative PCR analysis enable us to measure the quantity of the expressed transcript of catalogued cDNAs in the various tissues or in the embryos of certain developmental stages. The information derived from the experiments has been accumulating as the gene expression databases. Also recently more than 100,000 of the full-length cDNAs with functional annotation was open to the pubic. As a matter of course, such resources are now made use of to a wide variety of biological fields. Moreover, now, accurate genetic network in the biological processes is undertaken by the computer simulation utilizing such resources.
However, as biological processes depend on cell-to-cell interactions such as induction, cell-to-cell contact, lateral inhibition, etc., the information of overall gene expression of cellular magnification must be required. From the stand point of view, several databases of lower animals, for example, those of the Caenorhabditis elegans, the Drosophila melanogaster, and of the Xenopus laevis provide us the comprehensive cellular gene expression information. In those, expression pattern was obtained from the whole mount in situ hybridization techniques, immunohistochemical method using antibodes, and/or enhancer trapping of LacZ staining. Above all, in situ hybridization method is available directly from the cDNA that has been cloned. However in adapting these techniques in higher animals such as mouse or human, some difficulties must be overcome to carry out the systematic and large-scale in situ hybridization. First, gathering a lot of embryos is laborious. Secondly, before the implantation stage, embryos are too small to manipulate during the whole mount in situ hybridization processes. Lastly, after late stage embryos or in adult, it is very difficult to subject the tissues to the whole mount in situ hybridization. Instead, hybridization to suitable tissue sections may be practicable at least in the later stage embryos or in adult tissues.
Stuff
Associate Professor:
Tohru Komiya Ph.D
Career
1. Basical medical doctor course in Tokyo University
Research Scientist:
Yoko Tanigawa:
Master course 2nd grade :
Shin Fujita
Ayumi Takeuchi
Master course 1st grade :
Tomoaki Hiramatsu
Bachelor:
Maya Ishikawa
Moeka Fujisawa