Annual Report 2002
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CENTER FOR RADIOISOTOPE FACILITIES (CRF)

Head:YAMAMORI, Tetsuo (Professor, concurrent post)
Associate Professor:OGAWA, Kazuo (Radiation Protection Supervisor)
Technical Staffs:MATSUDA, Yoshimi (Radiation Protection Supervisor)
KATO, Yosuke (Radiation Protection Supervisor)
MOROOKA, Naoki (Radiation Protection Supervisor)
Supporting Staff:ITO, Takayo
IIDA, Yumi
KATAGIRI, Izumi

I. Research supporting activity

Technical and supporting staffs of the CRF are serving the purchase of radioisotopes from JRA (Japan Radioisotope Association) and the transfer of radioisotope wastes to JRA. The physical maintenance of the controlled areas where radioisotopes are used is also one of our business.

In this summer (July 1, 2002), a new radioisotope facility opened at the area E about 1km apart from the area A where four controlled areas have been built: Center, NIBB-sub, LGER (Laboratory of Gene Expression and Regulation)-sub, and NIPS (National Institute for Physiological Science)-sub. Each facility is maintained under the two radiation protection supervisors. Matsuda and Kato are named to the supervisors in the area A, and Ogawa and Morooka in the area E. Ito and Iida support them at the area A, and Katagiri at the area E.

Users going in and out the controlled areas counted by the monitoring system are 6,700 persons in 2002. The items in each controlled area is presented in Figure 1.

Figure 1. Percentage of users going in and out the controlled areas in 2002.

II. Academic activity

Academic activity by teaching staff is focused on the analysis of the structure and function of a dynein motor protein. Dyneins are a group of microtubule-activated ATPases that serve to convert chemical energy into mechanical energy and divided into axonemal and cytoplasmic dyneins. Figure 2 shows the localization of two isoforms of dynein in the outer arms of sperm axonemes (Ogawa et al., 1977) and the mitotic apparatus of cleaving egg (Mohri et al., 1976) visualized by anti-axonemal dynein (Fragment A) antibodies.

The native dyneins are very large and range in molecular mass up to 1 to 2 mega Da. They are complex proteins containing heavy, intermediate, and light chains defined by the molecular mass. Our present project is the molecular cloning of polypeptides contained in outer arm dynein of sea urchin sperm flagella to understand the mechanism how dynein interacts with microtubules, resulting in producing the force.

Outer arm dynein consists of two heavy chains with ATPase activity. The motor activity is closely related to this polypeptide. The first successful molecular cloning of this huge polypeptide (520 kDa) was performed in our laboratory in 1991. Since then cDNA clones for axonemal and cytoplasmic dyneins have been isolated in a variety of organisms. The sequences of heavy chains, without exception, contain four P-loop motives referred to as ATP-binding sites in the midregion of the molecules. Figure 3A and B draw the structure of heavy chain deduced from the amino acid sequence (Ogawa, 1992). Outer arm dynein contains three intermediate chains (IC1, IC2, and IC3) that range in molecular mass from 70 to 120 kDa. IC2 and IC3 were cloned by Ogawa et al. (1995) and contain the WD repeats in the carboxy-terminal halves of the molecules. By contrast, IC1 is not a member of the WD family. IC1 has a unique sequence such that the N-terminal part is homologous to the sequence of thioredoxin, the middle part consists of three repetitive sequences homologous to the sequence of NDP kinase, and the C-terminal part contains a high proportion of negatively charged glutamic acid residues (Ogawa et al., 1996). Thus, IC1 is a novel dynein intermediate chain distinct from IC2 and IC3 and may be a multifunctional protein.

Six light chains with molecular masses of 23.2, 20.8, 12.3, 11.5, 10.4, and 9.3 kDa are associating with outer arm dynein. We have already isolated cDNA clone of five LCs. LC1 (23.2 kDa) and LC3 (12.3 kDa) are highly homologous to mouse Tctex2 and Tctex1, respectively, which are encoded by the t complex region that is involved in transmission ratio distortion (TRD), male sterility and the development of germ cells. Our finding raises the possibility that axonemal dynein proteins are involved in this phenomenon. TRD may be caused by the dysfunction of multiple axonemal dynein proteins.

Figure 2. Localization of two dynein isotypes on outer arm of sperm axonemes and mitotic apparatus of cleaving egg.
Figure 3. Structure of dynein heavy chain. A; Analysis of amino acid sequence of heavy chain reveals that it consists of three major domains referred to as N, M, and C from the N-terminus. B and C; M and C domains make larger domain (motor domain) by intramolecular association.

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