DNA polymerases required for repair of UV-induced damage in Saccharomyces cerevisiae

The ability of yeast DNA polymerase mutant strains to carry out repair synthesis after UV irradiation was studied by analysis of postirradiation molecular weight changes in cellular DNA. Neither DNA polymerase alpha, delta, epsilon, nor Rev3 single mutants evidenced a defect in repair. A mutant defective in all four of these DNA polymerases, however, showed accumulation of single-strand breaks, indicating defective repair. Pairwise combination of polymerase mutations revealed a repair defect only in DNA polymerase delta and epsilon double mutants. The extent of repair in the double mutant was no greater than that in the quadruple mutant, suggesting that DNA polymerases alpha and Rev3p play very minor, if any, roles. Taken together, the data suggest that DNA polymerases delta and epsilon are both potentially able to perform repair synthesis and that in the absence of one, the other can efficiently substitute. Thus, two of the DNA polymerases involved in DNA replication are also involved in DNA repair, adding to the accumulating evidence that the two processes are coupled

The Nuclease Activity of the Yeast Dna2 Protein, Which Is Related to the RecB-like Nucleases, Is Essential in Vivo

Saccharomyces cerevisiae Dna2 protein is required for DNA replication and repair and is associated with multiple biochemical activities: DNA-dependent ATPase, DNA helicase, and DNA nuclease. To investigate which of these activities is important for the cellular functions of Dna2, we have identified separation of function mutations that selectively inactivate the helicase or nuclease. We describe the effect of six such mutations on ATPase, helicase, and nuclease after purification of the mutant proteins from yeast or baculovirus-infected insect cells. A mutation in the Walker A box in the C-terminal third of the protein affects helicase and ATPase but not nuclease; a mutation in the N-terminal domain (amino acid 504) affects ATPase, helicase, and nuclease. Two mutations in the N-terminal domain abolish nuclease but do not reduce helicase activity (amino acids 657 and 675) and identify the putative nuclease active site. Two mutations immediately adjacent to the proposed nuclease active site (amino acids 640 and 693) impair nuclease activity in the absence of ATP but completely abolish nuclease activity in the presence of ATP. These results suggest that, although the Dna2 helicase and nuclease activities can be independently affected by some mutations, the two activities appear to interact, and the nuclease activity is regulated in a complex manner by ATP. Physiological analysis shows that both ATPase and nuclease are important for the essential function of DNA2 in DNA replication and for its role in double-strand break repair. Four of the nuclease mutants are not only loss of function mutations but also exhibit a dominant negative phenotype

Some of our advances in breeding fruits and shrubs.

In Bulletin No. 22 we gave in detail our work in crossing the Russian Rosa rugosa with pollen of a number of the best garden roses. The crossing was done in the summer of 1892, and the seed planted the following spring. In the fall of 1893 the plants were potted and wintered in the cellar. The following spring they were planted out in nnrsery rows where they now stand. In the fall of 1894 the tops were cut back to mere stubs which were covered with earth. During the past season (1895,) they have made a rampant growth which has been unfavorable for the blossoming of such young plants. As a rule, the hybrids showing most variation from the Rosa rugosa mother have not bloomed, while those following more nearly the mother in leaf and habit have given more bloom. At this time we will only report two of the wide variations which have blossomed quite freely at this tender age

Second report on the sand cherry as a stock.

In Bulletin No. 22 appeared the first report of an experiment with the Sand Cherry (Prunus pumila) as a stock for the plum and cherry, and it should be read in connection with the following, which gives the result of the second year’s experience. The stocks were grown in 1892 from seed gathered in northwest Nebraska. In the fall of 1892 the largest of the seedlings were taken up for crown-grafting during the winter, leaving the others for budding. The grafts were planted in the spring of 1893, and the budding was done during July of the same year. The trees of suitable size were taken up late in the fall of 1894, the grafts having had two seasons’ growth, and the buds one season’s growth. All the trees had a very strong root-system, consisting mainly of a dense cluster of long cylindrical roots from immediately beneath the crown, no special tap-root being formed. The color was a fine shade of carmine. This red color is a marked characteristic of the sand cherry root

Rose hybrids

About ten years ago the Iowa Agricultural College imported from Russia types of the Rosa rugosa family, varying materially from the varieties introduced from China and Japan. The Russian forms prove hardier in the north, are more graceful in habit, and are finer in bud, flavor and foliage than the Japan varieties. Prof. L. H. Bailey, of the Cornell University Experiment Station, says in American Gardening (June, 1892, p. 342), of these types: “The form of rugosa from Russia, when grown side by side with the ordinary type, is about two weeks later to bloom, and a little darker in color. Where the ordinary rugosa has only two or three buds and flowers in a cluster, this one averages about four or five. The buds show a rich dark red between the narrow sepals, and besides being very long they are very pretty. * * * The blossom from which our engraving is made measures six inches across. * * * The double form of the rose introduced by Professor Budd seems to belong to the rugosa strain, and is known as R. cinnamomea. The blooms are six inches across, quite double, crimson in color, not quite so glowing as the type of rugosa, but more fragrant. The leaves are slightly serrated, bright green and leathery.

The use of the sand cherry for stocks

During the past few years the use of the Sand cherry (Primus pumila), as a stock for the cherry and plum, has been much discussed, but as yet we have had no convincing experience favorable to its use in cherry propagation. But in Utah and in various parts of the prairie States we have plum trees on this stock of several years’ growth, which favor the belief that our cultivated varieties will unite perfectly with its wood, come into bearing earlier, and become dwarfed in size of tree. With a view to more exact conclusions we grew, in 1892, about five thousand stocks from seed gathered in northwest Nebraska. The seed was washed from the pulp, dried for three or four days in the shade, mixed with sand in boxes, and put out for winter freezing. The seedlings made a fine stand and their growth the first season was about equal in height and diameter of stem to our seedlings of native plum. In the fall of 1892 we took up the largest of the seedlings for crown-grafting, leaving the others for budding

Coordination of Nucleases and Helicases during DNA Replication and Double-strand Break Repair

Nucleases and helicases are involved in numerous steps in DNA replication and repair. Nucleases act on intermediates in DNA replication created by DNA polymerases (Chapter 4) and helicases (Chapter 3). They can create substrates for repair as in Okazaki fragment processing (OFP) and homologous recombination. They can also create substrates for activation of a checkpoint response, or participate in downregulation of checkpoints. In the special case of telomere replication, they are also involved in essential processing steps (Chapter 8). Nucleases known to act during DNA replication include Dna2, Rad27, Mre11, Sae2, Exo1, RNaseH, Yen1 andMus81/Mms4. Of these, Dna2, Exo1 and Mre11 are of particular interest because they have been identified as crucial activities that initiate repair of double-strand breaks (DSBs) by homologous recombination and thus form an intrinsic link between DNA replication and repair of DSBs derived from replication fork failure. The action of the nucleases is coordinated with those of a number of helicases and is discussed here in the context of a network of their interactions that combine to maintain genome integrity during DNA replication

Coordination of Nucleases and Helicases during DNA Replication and Double-strand Break Repair

Nucleases and helicases are involved in numerous steps in DNA replication and repair. Nucleases act on intermediates in DNA replication created by DNA polymerases (Chapter 4) and helicases (Chapter 3). They can create substrates for repair as in Okazaki fragment processing (OFP) and homologous recombination. They can also create substrates for activation of a checkpoint response, or participate in downregulation of checkpoints. In the special case of telomere replication, they are also involved in essential processing steps (Chapter 8). Nucleases known to act during DNA replication include Dna2, Rad27, Mre11, Sae2, Exo1, RNaseH, Yen1 andMus81/Mms4. Of these, Dna2, Exo1 and Mre11 are of particular interest because they have been identified as crucial activities that initiate repair of double-strand breaks (DSBs) by homologous recombination and thus form an intrinsic link between DNA replication and repair of DSBs derived from replication fork failure. The action of the nucleases is coordinated with those of a number of helicases and is discussed here in the context of a network of their interactions that combine to maintain genome integrity during DNA replication