(1) Nobumoto Watanabe
Tsukuda Life Science Center
Institute of Physical and Chemical Research (RIKEN)
Sponsor and Host Institution:
Dr. George F. Vande Woude
ABL-Basic Research Program
NCI-Frederick, Cancer Research and Developmental Center
Dates of Visit: June 22 - July 23, 1991
Summary of Activities:
The main objective of this study is collaboration on studies on the function of c-mos protooncogene product with the host scientist. In addition to this main object, I plan to attend the meeting on oncogenes and visit some laboratories to discuss future collaboration.
In the lab of Dr. Vande Woude, mainly, I discussed with the scientists who study on the c-mos oncogene: they kindly showed their experimental procedures during their experiments, and I also showed my procedures for them. We discussed details of the experimental procedures. It was very useful for our collaboration. I also discussed with some scientists in NCI-Frederick and NIH-Bethesda. I have already started the collaboration with one of them.
At the Annual Meeting on Oncogenes, I gave a talk entitled “The Role of CSF (pp39mos) in meiotic release upon fertilization”. I discussed with many scientists who study about c-mos oncogene products. The reports of them were also interesting and it was very useful for my study to attend the meeting.
* Details about this talk are published recently.
Nature 352, 247 (1991)
In Boston, I gave a talk in the Marine Biological Station in Woods Hole and discussed with many good scientists about our future collaboration. The discussion with Dr. Ruderman (Prof. of Harvard Medical School) and her colleague was most useful for me among them, because they had lots of knowledge in the experiments using Xenopus oocytes.
In San Diego, I visited the Salk Institute. I also gave a talk entitled “Mos as CSF” there. The discussion after the seminar was very useful, because there were many scientists who were very familiar about c-mos oncogene (Dr. Hunter, Dr. Verma and the scientists in the lab of Dr. Donoghue (UCSD) etc). The discussion with Dr. Hunter about our future collaboration was also successful.
(2) Takashi Saito
Division of Molecular Genetics Center for Neurobiology and Molecular Immunology, School of Medicine, Chiba University
Sponsor and Host Institution:
Ronald N. Germain
Laboratory of Immunology National Institute of Allergy and Infectious Diseases National Institutes of Health
Dates of Visit: September 25 - October 12. 1991.
Summary of Activities:
The object of this study is to exchange the information and to discuss about the mechanism of signal transduction in T Iymphocytes from the recognition of antigen /MHC by T cell receptor (TCR)/CD3 complex to the expression of T cell function such as the induction of lymphokine gene transcription and secretion throughout the period of this program with researchers in Universities and also in the 21th International Lenkocyte Culture Conference.
(1) Structure and function of TCR/CD3 complex
I discussed with E. Reinherz, J. Ashwell and A. Weissman about the distinct function by different isoforms of TCR complex. There seems to be no more strong evidence to support the original idea that S-7 containing TCR complex is responsible for PI hydrolysis and activation-induced apoptosis. On the other hand, TCR complex containing FC!!
!R appears to have distinct function. Stimulation of Fc!!!R-containing TCR by antigen results in IL2 secretion but not growth inhibition. Fc!!!R can substitute to make functional TCR complex which mediates distinct signal transduction. There are 7-specific exons which are not conserved between species.I discussed with the structure and function of TCR!!
!dimer-expressing thymocytes with collaborators Y. Hashimoto and R. Kubo. Expression of!!!dimer in the absence of other TCR chains seems to occur during certain period of T cell differentiation and not due to particular!!!chain. They may have specific function for allelic exclusion of TCR!!!chain. (2) Signal transduction through TCR complex
On the discussion with L. Samelson and A. Weiss, fyn tyrosine kinase is shown to associate with TCR complex, but is not necessarily associated in all T cells. Trials to demonstrate the activation of fyn by T cell stimulation have been failed. Important contribution of CD45 for T cell activation was confirmed by J. Ashwell. C. June showed thesystem where Ca2+ oscillation in T cells was discussed. Transcriptional regulation of IL2 gene had been discussed with G. Crabtree. NFAT is composed of two components; one is located in cytoplasm and not inducible, the other is inducible and localize in the nuclei. T cell activation induces the cytoplasmic factor to translocate into nuclei. The mechanism of cyclosporin A to block IL2 production seems to inhibit the translocation of the cyioplasmic factor into the nuclei. The mechanisms of suppression by other immunosuppressive drug, FK506 and rapamycin are now under investigation.
Regarding the mechanism of anergy or activation-induced cell death, I discussed with R. Schwartz, M. Lenardo and T. Finkel. T cell activation requires second signal in addition to the signal through TCR/CD3 complex. The molecule responsible for this second signal seems to be CD28. However, there is a receptor with higher affinity than CD28 to the ligand of CD28. Lenardo found that pre-treatment of T cells with IL2 before TCR/CD3 triggering causes cell death. We interpreted this phenomenon as that the induction of apoptosis by TCR stimulation requires the T cells in cell cycle. However, Finkel demonstrated that cross-linking of CD4 with anti-CD4 or HIVgp 120 prior to TCR stimulation induced apoptosis. Generally, cross-linking of CD4 and TCR/CD3 simultaneously activates T cell synergistically. The signal transduction to induce growth inhibition or apoptosis have to be clarified to understand these phenomena.
(3) Jun-ichi Miyazaki
Department of Disease-related Gene Regulation Research (Sandoz), Faculty of Medicine The University of Tokyo
Sponsor and Host Institution:
Dr. Keiko Ozato
Laboratory of Developmental and Molecular Immunology National Institute of Child Health and Human Development National Institutes of Health
Dates of Visit: December 2 - 15, 1991
Summary of Activities:
Our group has been studying the role of major histocompatibility genes in immune regulation, using transgenic technique. We are currently planning to extend the study for clarify the mechanism and the role of transcriptional regulation of MHC class I genes. Dr. Ozato’s group cloned several regulatory genes related to MHC gene regulation. During my stay in her laboratory, I discussed with Dr. Ozato possible collaborative projects, and chose a project concerning a dominant negative nuclear hormone receptor mutant (for H-2RIIBP) as follows:
H-2RIIBP binds to a cis-acting regulatory DNA sequence, designated region II, upstream of MHC class I genes, which I defined when I was in her laboratory five years ago. This gene has become well known in the field. The nuclear hormone receptors heterodimerized with each other and H-2RlIBP is an important player for the dimerization. This heterodimerization requires the C-terminal domain. Her laboratory has shown that H-2RIIBP heterodimerizes with the retinoic acid receptor as well as the thyroid hormone receptor, and activates genes that respond to both retinoic acid and thyroid hormone (Marks et al. unpublished). A mutant H-2RIIBP, which does not have a DNA binding domain but has the dimerization domain, blocks functions of many nuclear hormone receptors in vitro, as expected. With this type of mutants we could learn a lot about overall importance of protein-protein interaction between transcription factors on gene regulation. Therefore, we decided to make transgenic mice harboring this mutant H-2RIIBP cDNA under some tissue specific promoter such as SAP promoter (liver-specific). This approach may be more informative than homologous recombination which I planned initially, since a compensatory effect by other similar genes is not expected to predominate.
According to the above scheme, I have done a part of DNA construction in her laboratory during my stay. Dr. Ozato and I agreed to continue this project as a collaborative work.