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R S Hawley

Publications and source records attributed to R S Hawley.

62 records · Page 4Linked to original sources
PubMed1986

DNA ligase from Drosophila melanogaster embryos. Purification and physical characterization.

A DNA ligase has been purified approximately 2,100-fold, to near-homogeneity, from Drosophila melanogaster 6-12-h embryos and was shown to catalyze the formation of 3',5'-phosphodiester bonds. Polypeptides with molecular weights 83,000, 75,000, and 64,000 were observed when the purified enzyme was electrophoresed under denaturing conditions. These polypeptides were shown by partial proteolysis studies and two-dimensional gel analysis to be structurally related. The two smaller polypeptides were presumably derived from the largest, 83,000 molecular weight protein, by proteolysis during purification or in vivo. All three polypeptides formed enzyme-adenylylate complexes in the absence of DNA. Drosophila DNA ligase had a Stokes radius of 45 A, a sedimentation coefficient of 4.3 S, and a frictional ratio of 1.6, yielding a calculated molecular weight of 79,800. These studies indicate that DNA ligase from Drosophila embryos is a monomer. The purified ligase was free of detectable ATPase, nuclease, topoisomerase, and DNA polymerase activities. The enzyme exhibited an absolute requirement for ATP in the joining reaction. A divalent metal was required and N-ethylmaleimide inhibited the reaction. Formation of phosphodiester bonds by Drosophila ligase required the presence of 5'-phosphoryl and 3'-hydroxyl termini. The purified enzyme restored biological activity to endonucleolytically cleaved pBR322 DNA. The specific activity of Drosophila DNA ligase was highest in unfertilized eggs. Developing embryos had 5-10-fold more ligase activity than at any later time in development.

Adenosine Triphosphate↗
PubMed1986

Autosomal modifiers of the bobbed phenotype are a major component of the rDNA magnification paradox in Drosophila melanogaster.

rDNA magnification in Drosophila melanogaster is defined experimentally as the ability of bb/Ybb- males to produce exceptional progeny that are wild type with respect to rDNA associated phenotypes. Here, we show that some of these bobbed-plus progeny result not from genetic reversion at the bb locus but rather from variants at two or more autosomal loci that ameliorate the bobbed phenotype of rDNA deficient males in Drosophila. In doing so we resolve several aspects of a long-standing paradox concerning the phenomenon of rDNA magnification. This problem arose from the use of two genetic assays, which were presumed to be identical, but paradoxically, produced conflicting data on both the kinetics of reversion and the stability of magnified bb+ chromosomes. We resolve this problem by demonstrating that in one assay bobbed-plus progeny arise primarily by genetic reversion at the bobbed locus, whereas in the other assay bobbed-plus progeny arise both by reversion and by an epistatic effect of autosomal modifiers on the bobbed phenotype. We further show that such modifiers can facilitate the appearance of phenotypically bobbed-plus progeny even under conditions where genetic reversion is blocked by magnification defective mutants. Finally, we present a speculative model relating the action of these modifiers to the large increases in rDNA content observed in males undergoing magnification.

Animals↗
PubMed1985

Repair-defect mutations inhibit rDNA magnification in Drosophila and discriminate between meiotic and premeiotic magnification.

We have examined rDNA magnification in Drosophila melanogaster males carrying one of 11 recombination- or repair-defective mutations representing seven loci. We show that mutations defined by a defect in postreplication repair (mus-101, mei-41, and mus-108) are also defective in rDNA magnification, whereas mutations that do not affect postreplication repair have little or no effect on magnification. mei-41 inhibits only premeiotic magnification events, while mus-108 blocks both premeiotic and meiotic events. This suggests that meiotic and premeiotic events share some but not all functions. A molecular analysis of rDNA magnification reveals that in mus-108 males, changes in the rDNA restriction pattern can occur within one or a few generations under magnifying conditions. We interpret these data in terms of the role of DNA repair systems in rDNA magnification and in terms of stable maintenance of tandemly repeated genes.

Animals↗
PubMed1985

A two-stage model for the control of rDNA magnification.

Males of the genotype bb/Ybb- have been shown to produce both magnified (bbm+) and, less frequently, reduced (bbrl) X chromosomes. An analysis of the progeny of single magnifying bb/Ybb- males reveals that bbm+ revertants may be recovered either as rare single events or, more frequently, in large clusters. To analyze the role of the bb phenotype in the induction of rDNA magnification we have constructed a series of bb and bb+ derivatives of Ybb-. Males carrying an X chromosomal bb allele and one of these derivatives (bb/bbYbb- or bb/bb+Ybb-) produce small numbers (one to two) of bbm+ progeny at a frequency similar to that observed for bb/Ybb- males but do not produce large clusters of bbm+ revertants. In addition, bb/bb+Ybb- males produce essentially equal numbers of magnified (bbm+) and reduced (bbrl) X chromosomes. These data, together with a consideration of the growth properties of the male germline in Drosophila, suggest that magnification/reduction may occur at two different times during development. Those events that give rise to large clusters, and, thus, necessarily arise early in germ cell development, appear to be dependent on the bb phenotype. However, those events that give rise to single bbm+ chromosomes arise late in spermatogenesis, probably at meiosis, and are independent of the bb phenotype.

Animals↗
PubMed1983

The effect of mei-41 on rDNA redundancy in Drosophila melanogaster.

The recombination and repair defective mutant, mei-41, exhibits three rather striking effects on the genetic properties and chromosomal stability of rDNA in Drosophila. First, mei-41 inhibits rDNA magnification. However, mei-9, another recombination and repair defective mutation has no similar effect. This indicates that magnification requires some, but not all, of the gene products necessary for meiotic exchange. Second, under magnifying conditions, mei-41 induces interchanges between the X rDNA and either arm of the Ybb- chromosome. These interchanges occur at high frequency and are independent of rDNA orientation. Third, in mei-41 bb+/Ybb+ males, bobbed mutants in the X, but not the Y, also arise at high frequency. Evidence suggests that these events involve the rDNA type I insertion. The recombination and repair defective properties of mei-41 together with our results regarding its unusual and specific effects involving rDNA are explained in a simple model that has general implications for chromosome structure.

Animals↗
PubMed1980

Chromosomal sites necessary for normal levels of meiotic recombination in Drosophila melanogaster. I. Evidence for and mapping of the sites.

Meiotic exchange was measured in females heterozygous for a normal sequence X chromosome and for each of eleven T(1;4)s and each of sixteen T(1;Y)s. The results indicate that the X chromosome can be divided into five intervals, such that heterozygosity for a breakpoint in one interval strongly suppresses exchange within that interval, but has little or no effect on exchange in other intervals. The boundaries between these intervals are identified and mapped to regions 3C4-6/7, 7A-7E, 11A and proximal to 18C on the standard salivary map; each boundary is located at (or within a small region containing) a major constriction (i.e., a block of intercalary heterochromatin).--Exchange was examined in females heterozygous for translocations broken within the constriction at 11A. The results imply that a boundary occupies only a subregion of the entire constriction and is subdivisible by translocation breakpoints. Several other properties of boundaries have been elucidated. Finally, the relationship of these data to a simple model of meiotic pairing proposed by I. Sandler (1956) and to the role of intercalary heterochromatin in the meiotic process is discussed.

Animals↗