Margaret McCarron

Organization of the Rosy locus in Drosophila melanogaster.

A. Chovnick, W. Gelbart* and M. McCarron Genetics and Cell Biology Section Biological Sciences Group The University of Connecticut Storrs. Connecticut 06268 One of the critical issues in eucaryotic molecular genetics is the nature of control elements and their relationship to the structural elements that they regulate. A particularly favorable system for investi- gating such questions is provided by the rosy locus of Drosophila melanogaster. Originally recovered and defined as recessive brownish eye color mutants, rosy mutants were shown subsequently to be deficient in red (drosop- terin) pigment and to exhibit no detectable xan- thine dehydrogenase (XDH) activity (Glassman and Mitchell, 1959). Figure 1 summarizes reactions used in this laboratory to assay Drosophila XDH (Forrest, Glassman and Mitchell, 1956; Glassman and Mitchell, 1959). Of particular interest is the fact that zygotes possessing little or no XDH activity are unable to complete development and die before eclosion on standard Drosophila culture medium supplemented with purine (Glassman, 1965). Two observations serve to place the coding information for XDH in or near rosy: first, variation in dosage of ry+ alleles, from O-3 doses, appears to be the limit- ing factor in determining level of XDH activity/fly in otherwise wild-type flies (Grell, 1962; Glassman, Karam and Keller, 1962); second, the genetic basis for variation in electrophoretic mobility of XDH seen in wild-type strains maps to the immediate vicinity of the rosy locus (Yen and Glassman, 1965). All these observations have been confirmed and extended in this laboratory (Chovnick, 1966; Fin- nerty, Baillie and Chovnick, 1970; McCarron, Gel- bart and Chovnick, 1974). Electrophoretic mobility variants of XDH which map to the rosy locus are readily isolated from laboratory stocks and natural populations of Dro- sophila melanogaster. From these sources, we have established a number of wild-type isoalleles of the rosy locus. These are maintained as stable lines which possess XDH molecules with distinctive electrophoretic mobilities and thermal properties. Moreover, the XDH enzyme activity level associated with each of these wild-type alleles is also a distinc- tive, stable phenotypic character. Table 1 summa- rizes our present array of ry+ isoalleles. The XDH produced by ry+O serves as a mobility standard and is designated XDH1.Oo. Under standard conditions of

Gene expression in Drosophila: post-translational modification of aldehyde oxidase and xanthine dehydrogenase.

SummaryMaroon-like homozygotes are completely deficient for xanthine dehydrogenase (XDH) and aldehyde oxidase (AO), however ma-l is not a structural locus for either enzyme. Quantitative immunoelectrophoresis of ma-l and wild type extracts suggests that the ma-l function must be post-translational. To determine whether the ma-l function involves some direct physical changes in XDH and/or AO the enyzmes were characterized with respect to temperature sensitivity and behavior in gel sieving electrophoresis. Since the XDH and AO from complementary ma-l heterozygotes is more thermolabile and different in shape from wild type XDH and AO, we conclude that ma-l is involved in a post-translational modification of these enzymes.

Structural and Functional Organization of a Gene in Drosophila Melanogaster

In recent years, considerable interest and research effort has been directed towards an understanding of the mechanisms underlying the control of gene expression in higher organisms. Several experimental approaches have been employed to this end, and the results of such studies have been the subject of recent review (Axel et al., 1979). Major emphasis in this laboratory has been directed towards the development of an experimental system in Drosophila melanogaster involving the genetic dissection of a specific genetic unit. The end-product of this effort will be a gene whose DNA is marked and mapped to permit eventual identification of the various sequences that control its expression.

P-element transposase induces male recombination in Drosophila melanogaster.

Male recombination in P-M dysgenic crosses has been viewed as a reflection of P-element transposase interacting with P elements. However, recent studies suggest that the transposase may catalyse double-stranded breaks in chromosomal DNA. We have, therefore, introduced P(delta 2-3 ry+) (99B), a single non-mobile P-element transposase source, into the long-standing laboratory true M strains of a flanking lethal crossover selective system, thus facilitating the examination of rare male recombination events as an assay for transposase activity. We find that the rate of male recombination in the presence of this non-mobile P element is greater than twenty times the background rate of male recombination in the control examined prior to introduction of the transposase source.

Studies on Recombination in Higher Organisms

Random-strand and half-tetrad recombination studies of rosy and maroon-like mutants have permitted us to investigate linked exchange in higher organisms. These studies involve the systematic recovery and analysis of the products of exchange events restricted to exceedingly short genetic intervals. Our observations lead us to conclude that all recombination involves a nonreciprocal transfer of information in the immediate region of the exchange event. These studies have been reported in great detail, and they have been the subject of recent review (Chovnick et al., 1971; Finnerty, 1974).

ELECTROPHORETIC VARIANTS AS A TOOL IN THE ANALYSIS OF GENE ORGANIZATION IN HIGHER ORGANISMS

ABSTRACT . The rosy locus of chromosome 3 in Drosophila melanogaster controls the biosynthesis of the enzyme xanthine dehydrogenase. This report describes our progress in an investigation of the structural and functional organization of this locus as a model system for the study of higher organism gene organization. A number of wild-type isoalleles of the rosy locus have been isolated which are associated with the production of electrophoretically distinguishable XDH molecules. Large scale recombination experiments were carried out involving null enzyme mutants induced on electrophoretically distinct wild-type isoalleles. The genetic sites responsible for the mobility differences are followed as unselected markers in these crosses. Since electrophoretic variants represent alterations in the portion of the locus that encodes structural information, the resultant map of electrophoretic sites defines the minimal limits of that portion of the rosy locus. The present report additionally presents evidence that the structural element of the rosy locus is a single, uninterrupted sequence coding for a polypeptide molecule that is present in XDH as a dimer.