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Chlamydomonas inner-arm dynein mutant, ida5, has a mutation in an actin-encoding gene.

Kato-Minoura T, Hirono M, Kamiya R - J. Cell Biol. (1997)

Bottom Line: These mutants were found to have mutations in the conventional actin gene, such that its product is totally lost; ida5 has a single-base deletion that results in a stop codon at a position about two-thirds from the 5' end of the coding region, and ida5-t lacks a large portion of the entire actin gene.The net growth rate of ida5 and ida5-t cells did not differ from that of wild type, but the mating efficiency was greatly reduced.These results suggest that NAP can carry out some, but not all, functions performed by conventional actin in the cytoplasm and raise the possibility that Chlamydomonas can live without ordinary actin.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, School of Science, Nagoya University, Japan.

ABSTRACT
Chlamydomonas flagellar inner-arm dynein consists of seven subspecies (a-g), of which all but f contain actin as subunits. The mutant ida5 and a new strain, ida5-t, lack four subspecies (a, c, d, and e). These mutants were found to have mutations in the conventional actin gene, such that its product is totally lost; ida5 has a single-base deletion that results in a stop codon at a position about two-thirds from the 5' end of the coding region, and ida5-t lacks a large portion of the entire actin gene. Two-dimensional gel electrophoresis patterns of the axonemes and inner-arm subspecies b and g of ida5 lacked the spot of actin (isoelectric point [pI] = approximately 5.3) but had two novel spots with pIs of approximately 5.6 and approximately 5.7 instead. Western blot with different kinds of anti-actin antibodies suggested that the proteins responsible for the two novel spots and conventional actin are different but share some antigenicity. Since Chlamydomonas has been shown to have only a single copy of the conventional actin gene, it is likely that the novel spots in ida5 and ida5-t originated from another gene(s) that codes for a novel actin-like protein(s) (NAP), which has hitherto been undetected in wild-type cells. These mutants retain the two inner-arm subspecies b and g, in addition to f, possibly because NAP can functionally substitute for the actin in these subspecies while they cannot in other subspecies. The net growth rate of ida5 and ida5-t cells did not differ from that of wild type, but the mating efficiency was greatly reduced. This defect was apparently caused by deficient growth of the fertilization tubule. These results suggest that NAP can carry out some, but not all, functions performed by conventional actin in the cytoplasm and raise the possibility that Chlamydomonas can live without ordinary actin.

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Fertilization tubules in wild-type (A) and ida5 (B) mt+  gametes produced in response to a 1-h exposure to 10 mM dibutyryl-cAMP and 1 mM IBMX. Bar, 0.3 μm.
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Figure 7: Fertilization tubules in wild-type (A) and ida5 (B) mt+ gametes produced in response to a 1-h exposure to 10 mM dibutyryl-cAMP and 1 mM IBMX. Bar, 0.3 μm.

Mentions: To confirm the above prediction, we induced fertilization tubule growth by treating gametes with dibutyrylcAMP and IBMX (Pasquale and Goodenough, 1987) and observed them with a differential-interference microscope. After about an hour of treatment, fertilization tubules were clearly seen in >70% of the mt+ wild-type gametes, whereas no fertilization tubules were found in mt+ ida5 gametes. EM indicated that a small fraction of ida5 gametes in fact produced fertilization tubules of abnormal shapes (Fig. 7). In contrast with the slender fertilization tubules of wild type, the ida5 fertilization tubules were round and stubby. Appendices of round appearance were also observed in mt− ida5 gametes treated with dibutyryl-cAMP and IBMX, as well as in the mt− wild-type gametes thus treated (Pasquale and Goodenough, 1987).


Chlamydomonas inner-arm dynein mutant, ida5, has a mutation in an actin-encoding gene.

Kato-Minoura T, Hirono M, Kamiya R - J. Cell Biol. (1997)

Fertilization tubules in wild-type (A) and ida5 (B) mt+  gametes produced in response to a 1-h exposure to 10 mM dibutyryl-cAMP and 1 mM IBMX. Bar, 0.3 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139884&req=5

Figure 7: Fertilization tubules in wild-type (A) and ida5 (B) mt+ gametes produced in response to a 1-h exposure to 10 mM dibutyryl-cAMP and 1 mM IBMX. Bar, 0.3 μm.
Mentions: To confirm the above prediction, we induced fertilization tubule growth by treating gametes with dibutyrylcAMP and IBMX (Pasquale and Goodenough, 1987) and observed them with a differential-interference microscope. After about an hour of treatment, fertilization tubules were clearly seen in >70% of the mt+ wild-type gametes, whereas no fertilization tubules were found in mt+ ida5 gametes. EM indicated that a small fraction of ida5 gametes in fact produced fertilization tubules of abnormal shapes (Fig. 7). In contrast with the slender fertilization tubules of wild type, the ida5 fertilization tubules were round and stubby. Appendices of round appearance were also observed in mt− ida5 gametes treated with dibutyryl-cAMP and IBMX, as well as in the mt− wild-type gametes thus treated (Pasquale and Goodenough, 1987).

Bottom Line: These mutants were found to have mutations in the conventional actin gene, such that its product is totally lost; ida5 has a single-base deletion that results in a stop codon at a position about two-thirds from the 5' end of the coding region, and ida5-t lacks a large portion of the entire actin gene.The net growth rate of ida5 and ida5-t cells did not differ from that of wild type, but the mating efficiency was greatly reduced.These results suggest that NAP can carry out some, but not all, functions performed by conventional actin in the cytoplasm and raise the possibility that Chlamydomonas can live without ordinary actin.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, School of Science, Nagoya University, Japan.

ABSTRACT
Chlamydomonas flagellar inner-arm dynein consists of seven subspecies (a-g), of which all but f contain actin as subunits. The mutant ida5 and a new strain, ida5-t, lack four subspecies (a, c, d, and e). These mutants were found to have mutations in the conventional actin gene, such that its product is totally lost; ida5 has a single-base deletion that results in a stop codon at a position about two-thirds from the 5' end of the coding region, and ida5-t lacks a large portion of the entire actin gene. Two-dimensional gel electrophoresis patterns of the axonemes and inner-arm subspecies b and g of ida5 lacked the spot of actin (isoelectric point [pI] = approximately 5.3) but had two novel spots with pIs of approximately 5.6 and approximately 5.7 instead. Western blot with different kinds of anti-actin antibodies suggested that the proteins responsible for the two novel spots and conventional actin are different but share some antigenicity. Since Chlamydomonas has been shown to have only a single copy of the conventional actin gene, it is likely that the novel spots in ida5 and ida5-t originated from another gene(s) that codes for a novel actin-like protein(s) (NAP), which has hitherto been undetected in wild-type cells. These mutants retain the two inner-arm subspecies b and g, in addition to f, possibly because NAP can functionally substitute for the actin in these subspecies while they cannot in other subspecies. The net growth rate of ida5 and ida5-t cells did not differ from that of wild type, but the mating efficiency was greatly reduced. This defect was apparently caused by deficient growth of the fertilization tubule. These results suggest that NAP can carry out some, but not all, functions performed by conventional actin in the cytoplasm and raise the possibility that Chlamydomonas can live without ordinary actin.

Show MeSH
Related in: MedlinePlus