NATIONAL INSITUTE FOR BASIC BIOLOGY

National Institute for Basic Biology

DIVISION OF CELLULAR COMMUNICATION

(Adjunct)

Professor:
Ritsu Kamiya
Associate Professor:
Katsushi Owaribe
Research Associate:
Takako Minoura
Postdoctral Fellow:
Osamu Kagami 1
Graduate student:
Itsushi Minoura (University of Tokyo)


The research in this laboratory, started in November, 1966, is aimed at an understanding of the molecular mechanisms that regulate the assembly and function of cytoskeletal proteins. Current research effort is centered on the function of actin and flagellar dynein. The organism employed is Chlamydomonas, a biflagellate green alga particularly suited for genetic and molecular biological studies.

A single flagellar axoneme contains at least eleven kinds of dynein heavy chains in inner and outer arms. To understand the specific function of each heavy chain, we have been isolating and characterizing mutants that lack different kinds of axonemal dyneins. Their motility phenotypes have indicated that different dynein species significantly differ in function. For example, the outer arm heavy chains are important for flagellar beating at high frequency, whereas the inner-arm heavy chains are important for producing proper waveforms. Indirect evidence also suggests that the properties of force generation by different heavy chains differ greatly. We are currently carrying out micro-physiological experiments to assess the force production by each species of dynein.

The inner dynein arms are known to contain actin as a subunit. Hence the two independent motility systems of eukaryotes - the actin-based and microtubule-based systems - should somehow cooperate in the inner arm dynein, although the function of actin as a subunit of dynein is totally unknown at present. Recently we found that the mutant ida5, lacking four out of the seven subspecies of inner-arm dyneins, has a mutation in the actin-encoding gene. Intriguingly, Chlamydomonas has only a single gene of conventional actin, and the mutant ida5 was found to express no conventional actin at all. On close inspection, however, the cytoplasm and axoneme of this mutant were found to contain a novel actin-like protein (NAP) which has only a very low (64%) homology to conventional actin. Despite the lack of the conventional actin, ida5 displays normal cell division and grows as rapidly as wild type. However, the mating efficiency of this mutant is much lower than in wild type, because it is deficient in the growth of the fertilization tubule, an F-actin-containing structure that facilitates mating. Thus conventional actin and NAP appear to overlap in some, but not all, cellular functions.

The actin-null mutant ida5 should be of great use in elucidating the functional domains of actin. We have recently succeeded in transforming this mutant with a cloned actin gene, and found that inner arm dynein, as well as the fertilization tubule in gametic cells, became restored in the transformants. We are currently trying to transforming the mutant with artificially modified actin gene, to see if polymerization or interaction with myosin is important for the actin function in the dynein arms. We are also examining expression and localization of actin and NAP in various stages of cell cycle. Studies on those lines will enable us to determine whether actin or NAP is essential for cytokinesis, assembly and function of inner dynein arms, and other fundamental cellular phenomena in Chlamydomonas and other cells.

Fig. 1.
Fertilization tubules in wild-type(A) and ida5 (B) mt+ gametes produced in response to a 1 hour exposure to 10 mM dibutyryl-cAMP and 1 mM IBMX. Bar, 0.3 µm. Wild-type fertilization tubules have been shown to contain F-actin bundles.


Publication List:
Hayashibe, K., Shingyoji, C. and Kamiya, R. (1997). Induction of temporary beating in paralyzed flagella of Chlamydomonas mutants by application of external force. Cell Motil. Cytoskel. 37:232-239.
Kato-Minoura, T., Hirono, M. and Kamiya, R. (1997). Chlamydomonas inner-arm dynein mutant, ida5, has a mutation in an actin-encoding gene. J. Cell Biol. 137: 649-656.
Takada, S. and Kamiya, R. (1997). Beat frequency difference between the two flagella of Chlamydomonas depends on the attachment site of outer dynein arms on the outer-doublet microtubules. Cell Motil. Cytoskel. 36, 68-75.
Wakabayashi, K., Yagi, T. and Kamiya, R. (1997). Ca2+-dependent waveform conversion in the flagellar axoneme of Chlamydomonas mutants lacking the centralpair/radial spoke system. Cell Motil. Cytoskel. 38:22-28.


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Last Modified: 12:00, May 28, 1998