No abstract

We have isolated 37 radiation-sensitive mutants of the basidiomycete Coprinus cinereus. Each mutation is recessive, and the collection defines at least ten complementation groups for survival of gamma irradiation. Four complementation groups define the genes rad3, rad9, rad11 and rad12, which are required both for survival of gamma irradiation and for meiosis. Mutants in each of these four groups fail to complete meiosis and produce mushrooms with greatly reduced numbers of viable spores. Propidium iodide staining of meiotic nuclei showed a characteristic terminal appearance for each mutant: few cells of any of the meiotic mutants progress beyond prophase I, and both condensation and fragmentation or dispersal of meiotic chromatin are frequently observed. Scanning electron micrographs showed that the meiotic mutants make varying numbers (0-6) of basidiospore initials and that few of these initials develop into mature spores. When initials are present they are always symmetrically arrayed on the basidium, regardless of initial number. In quantitative measurements of gamma ray sensitivity, double mutants of every tested combination of rad3, rad9, rad11 and rad12 consistently showed the same gamma ray sensitivity as the more sensitive single mutant parent of the cross. Therefore, these four genes are in the same pathway for the repair of gamma radiation damage, and this pathway also represents one or more functions essential for meiosis.

Treatment of Nitella antheridia with colchicine results in various sperm abnormalities, depending upon duration of exposure and subsequent recovery . Early effects of treatment include disappearance of spindle fibers and a cessation of ordered cell wall formation in dividing cells . Sperm released from antheridia treated for 24 hr and allowed to recover for 4-5 days possess branched flagella . After a recovery period of 6-10 days the sperm appear normal ; however, following longer recovery periods, the sperm exhibit variations in size and number of flagella . Branched flagella contain a variety of microtubule patterns ranging from branches containing a single microtubule to flagella with an excess of microtubules . Spermatids which differentiate in the presence of colchicine lack flagella and a microtubular sheath . Nuclear contents undergo condensation stages ; however, the nucleus as a whole does not undergo the orderly elongation and coiling characteristic of untreated Nitella spermatids . Long-term colchicine treatment followed by a recovery period produces atypical microtubules and microtubular aggregations in the spermatid . The results indicate that colchicine affects not only polymerization of microtubule subunits but also factors responsible for their ordered spatial relationships in the cell . The presence of microtubules is a prerequisite for normal morphological changes during spermiogenesis .

No abstract

We have been able to purify, in bulk, the cytoplasmic microtubule proteins of eggs and embryos of Drosophila melanogaster by means of in vitro self-assembly of microtubules from subunits present in a high-speed supernatant fraction of eggs or embryos. This provides the first successful application of this method to purification of microtubule protein from a source other than vertebrate brain, and the first purification of insect microtubule proteins. Our electron micrographs show that the in vitro assembled microtubules are morphologically typical and apparently are comprised of the expected 13 protofilaments. The protein we obtain from such preparations binds [3H]colchicine and has a sedimentation value of 6.4 S-6.9 S which is close to the predicted value for microtubule protein dimer. Both alpha- and beta-microtubule proteins are evident in sodium dodecyl sulfate polyacrylamide electropherograms of the isolated proteins. The apparent molecular weights of these species on dodecyl sulfate polyacrylamide gels are 54,000 and 52,000, respectively. These values as well as the amino acid composition and N-terminal methionine of the Drosophilia proteins are very closely comparable to microtubule proteins from other, unrelated organisms.