Molecular Characterization of Ribosomal Genes on the Ybb- Chromosome of DROSOPHILA MELANOGASTER

The question of whether the Ybb- chromosome contains ribosomal genes has been examined by using Southern blot analysis and comparing rDNA hybridization patterns for X/X and X/Ybb- DNA. The results demonstrate that the Ybb- chromosome contains sequences that hybridize to an rDNA probe under stringent conditions. Differential hybridization of some of these sequences with DNAs corresponding to different regions of a complete ribosomal gene repeat provides evidence that some of the genes on the Ybb- chromosome are type 2 repeats. Because data obtained by other workers suggest that type 2 repeats are transcribed only to low levels, these repeats may be classed as "nonfunctional". A further finding is that the ribosomal genes on the Ybb- chromosome do not undergo multiple rounds of DNA replication during polytenization of X/Ybb- cells.     

Polytenization of the Ribosomal Genes on the X and Y Chromosomes of DROSOPHILA MELANOGASTER

It has previously been shown (Endow and Glover 1979), that polytenization of the ribosomal genes in D. melanogaster Ore-R X/Y cells and in hybrid X/X cells (Endow 1980) involves replication of genes predominantly from one of the cell's two nucleolus organizers. This analysis takes advantage of strain-specific differences in X and Y chromosome rDNA hybridization patterns detected using the Southern blotting technique. In this report, I extend the previous observations by examining polytene rDNA patterns in wild-type and hybrid X/Y cells. A dominance hierarchy for the X and Y chromosomes from three strains of D. melanogaster is presented and possible mechanisms of replicative dominance are discussed.     

Regulation of Ribosomal RNA Gene Multiplicity in DROSOPHILA MELANOGASTER

The ribosomal RNA (rRNA) genes of Drosophila melanogaster can undergo a disproportionate replication of their number. This occurs when the cluster of rRNA genes (rDNA) of one chromosome is maintained with a homologous chromosome that is completely or partially deficient in its rDNA. Under appropriate genetic conditions, it appears that disproportionate rDNA replication can be generated at the level of both somatic and germ line cells. In the latter case, mutants partially deficient for rDNA can increase their rRNA gene number to the wild type level and transmit this new genotype to successive generations.     

One-Step and Stepwise Magnification of a BOBBED LETHAL Chromosome in DROSOPHILA MELANOGASTER

Bobbed lethal (bbl) chromosomes carry too few ribosomal genes for homozygous flies to be viable. Reversion of bbl chromosomes to bb or nearly bb+ occurs under magnifying conditions at a low frequency in a single generation. These reversions occur too rapidly to be accounted for by single unequal sister chromatid exchanges and seem unlikely to be due to multiple sister strand exchanges within a given cell lineage. Analysis of several one-step revertants indicates that they are X-Y recombinant chromosomes which probably arise from X-Y recombination at bb. The addition of ribosomal genes from the Y chromosome to the bbl chromosome explains the more rapid reversion of the bbl chromosome than is permitted by single events of unequal sister chromatid exchange. Analysis of stepwise bbl magnified chromosomes, which were selected over a period of 4-9 magnifying generations, shows ribosomal gene patterns that are closely similar to each other. Similarity in rDNA pattern among stepwise magnified products of the same parental chromosome is consistent with reversion by a mechanism of unequal sister strand exchange.     

Factors Influencing Disproportionate Replication of the Ribosomal RNA Cistrons in DROSOPHILA MELANOGASTER

This report describes studies of the compensatory response employing D. melanogaster stocks that bear cloned-X chromosomes derived from laboratory populations of strains Oregon R and Canton S. We find that modification of the autosomal background in either the female or the (see PDF) male parent influences the expression of the compensatory response by X chromosomes derived from the Canton S population, whereas Oregon R isolates are unresponsive to these effects. We have also studied compensatory replication in X/O larvae produced from cloned-X derivatives of both Canton S and Oregon R. Canton S larval compensation exceeds that of the adult, whereas in Oregon R the converse is true. We have concluded that both X chromosomal and autosomal factors affect the expression and magnitude of the compensatory response.     

Genes Affecting Productivity in Natural Populations of DROSOPHILA MELANOGASTER

Two hundred second chromosomes were extracted from a Japanese population in October of 1972, and the viabilities and productivities of homozygotes and heterozygotes from them were examined. Viability was measured by the Cy method and productivity by the number of progeny produced per female. The frequency of lethal-carrying chromosomes was 0.315. When the average heterozygote viability was standardized as 1.000, the average homozygote viability was 0.595 including the lethal lines, and 0.866 excluding them. The frequency of recessive sterile chromosomes among 131 non-lethal lines was 0.092 in females and 0.183 in males. There were two instances in which homozygosis for the second chromosome caused sterility in both sexes, which was close to the number expected (2.2) on a random basis of 0.092 x 0.183 x 131. When the average heterozygote productivity of 200 lines was standardized as 1.000, the average homozygote productivity was 0.532 including female steriles, and 0.584 excluding them. The ratio of detrimental load to lethal load was 0.383, while the ratio of partial sterility load to complete sterility load was 5.767. The average viability of lethal heterozygotes was slightly, but not significantly, lower than that of lethal-free heterozygotes, while the average productivity of lethal heterozygotes was significantly lower than that of lethal-free heterozygotes. There was a significant association of sterility in either sex with low viability of homozygotes. However, no statistically significant differences in viability and productivity were detected between sterile heterozygotes and non-sterile heterozygotes. The heterozygous effects of viability and productivity polygenes were examined by regressions of the heterozygotes on the sum of corresponding homozygotes. The regression coefficients were slightly positive for both viability and productivity if lethal and sterile chromosomes were excluded. The correlation between viability and productivity in homozygotes was significantly positive when sterile chromosomes were included, but the significance disappeared when the sterile chromosomes were excluded. In the heterozygotes there were no detectable correlations between them.     

A Meiotic Mutant Affecting Recombination in Female DROSOPHILA MELANOGASTER

mei-S282 is a female meiotic mutant isolated from a natural population of Drosophila melanogaster. It is a recessive mutation located at approximately map position 5 on the third chromosome which has two major effects. It causes a nonuniform decrease in recombination which is most drastic in distal chromosome regions and nondisjunction of all chromosome pairs is elevated at the first meiotic division. Nondisjunctional events are positively correlated; furthermore, nondisjoining chromosomes, themselves nonrecombinant, are preferentially recovered from cells in which nonhomologs are preferentially recovered from cells in which nonhomologs are also non-recombinant.-It is concluded that mei-S282 is a defect which occurs early in meiosis I prior to the time of exchange. In the mutant, the frequency of no-exchange tetrads for each of the major chromosomes is increased-and in cells which contain two or more no-exchange tetrads, an interaction between these chromosomes leads to correlated nondisjunction. mei-S282(+) then, is an exchange precondition necessary for the normal frequency and distribution of exchanges.     

Genetic Analysis of the 5s RNA Genes in DROSOPHILA MELANOGASTER

The 5S RNA genes of Drosophila melanogaster in either an isogenic wild-type or a multiply inverted (SM1) chromosome 2 increase their multiplicity when opposite a deficiency for the 5S gene site. This is analogous to the compensation phenomenon previously described for the 18S and 28S ribosomal RNA genes of the X chromosome nucleolus organizer region. Molecular hybridization of 5S RNA to DNA containing various doses of the 56F1-9 region of chromosome 2 demonstrates that most, if not all, of the 5S genes reside in or near this region. Also, a deficiency missing approximately one-half of the wild-type number of 5S genes was isolated and genetically localized. This mutant has a phenotype like that of bobbed, a mutant known to be partially deficient in 18S and 28S ribosomal RNA genes. Finally, we report the existence of a chromosomal rearrangement which splits the second chromosome into two segments, each containing 5S DNA.     

A Study of Rdna Magnification Phenomenon in a Repair-Recombination Deficient Mutant of DROSOPHILA MELANOGASTER

The rDNA magnification process consists of a rapid and inheritable rDNA increase occurring in bobbed males: in a few generations the bb loci acquire the wild-type rDNA value and reach a bb⁺ phenotype.—We have analyzed the rDNA magnification process in the repair-recombination-deficient mutant mei9^a, both at the phenotypical and rDNA content levels. In mei9^a bb double mutants the recovery of bb⁺ phenotype is strongly disturbed and the rDNA redundancy value fails to reach the wild-type level. The strong effect of this meiotic mutation on rDNA magnification suggests a close relationship between this phenomenon and the repair-recombination processes.     

Mutations in Genes Encoding Essential Mitotic Functions in DROSOPHILA MELANOGASTER

Temperature-sensitive mutations at 15 loci that affect the fidelity of mitotic chromosome behavior have been isolated in Drosophila melanogaster. These mitotic mutants were detected in a collection of 168 EMS-induced X-linked temperature-sensitive (ts) lethal and semilethal mutants. Our screen for mutations with mitotic effects was based upon the reasoning that under semirestrictive conditions such mutations could cause an elevated frequency of mitotic chromosome misbehavior and that such events would be detectable with somatic cell genetic techniques. Males hemizygous for each ts lethal and heterozygous for the recessive autosomal cell marker mwh were reared under semirestrictive conditions, and the wings of those individuals surviving to adulthood were examined for an increased frequency of mwh clones. Those mutations producing elevated levels of chromosome instability during growth of the wing imaginal disc were also examined for their effects on chromosome behavior in the cell lineages producing the abdominal cuticle. Fifteen mutations affect chromosome behavior in both wing and abdominal cells and thus identify loci generally required for the fidelity of mitotic chromosome transmission. Mapping and complementation tests show that these mutations represent 15 loci. One mutant is an allele of a locus (mus-101) previously identified by mutagen-sensitive mutants and a second mutant is an allele of the lethal locus zw 10.--The 15 mutants were also examined cytologically for their effects on chromosomes in larval neuroblasts. Taken together, the results of our cytological and genetical studies show that these mutants identify loci with wild-type functions necessary for either maintenance of chromosome integrity or regular disjunction of chromosomes or chromosome condensation. Thus, these mutations define a broad spectrum of genes required for the normal execution of the mitotic chromosome cycle.     

A Genetic Locus Having TRANS and CONTIGUOUS CIS Functions That Control the Disproportionate Replication of Ribosomal RNA Genes in DROSOPHILA MELANOGASTER

The results of deficiency mapping experiments reveal the presence of a compensatory response (cr+) locus that is located distal to the cluster of ribosomal RNA (rRNA) genes and is responsible for disproportionately replicating these genes when cr+ locus is present in a single dose, as in X/O males or X/sc4-sc8 females. The cr+ locus is novel in that it exhibits both trans and contiguous cis acting properties in somatic cells. It acts in trans to detect the presence of its partner locus in the opposite homolog, and if that partner locus is absent, it acts in cis to drive the disproportionate replication of those rRNA genes (rDNA) that are contiguous with it. The ability of cr+ to function is independent of the number of ribosomal RNA genes present. Furthermore, it can be shown that the cr+ locus is not required for the magnification or reduction of germ line rDNA. Finally, the implication of cr+ for position-effect variegation and the apparent reversion of the abnormal oocyte (abo) phenotype are discussed.     

The Effect of ABO Phenotypic Expression on Ribosomal DNA Instabilities in DROSOPHILA MELANOGASTER

The recessive maternal-effect mutation, abnormal oocyte (abo:2--38), reduces viability in the offspring of homozygous mutant females. Zygotes lacking specific heterochromatic segments of the X or Y chromosomes are most severely affected. We have shown that abo/abo lines can lose the capacity to express the mutant phenotype, and that elevated rDNA redundancies can be observed in such stocks (Krider and Levine 1975). In this study, we describe a microhybridization procedure that facilitates the measurement of rDNA redundancy, using a small number of adult Drosophila. We show that instability of the rDNA content persists in an abo/abo line after loss of the capacity to express the phenotype, and that changes in rDNA amounts occur between successive generations of the stock. Further, we show that the rDNA content of XO progeny from abo/abo females is elevated. The effect is directly correlated with the expression of the abo phenotype, and it is not observed in the XO progeny of abo heterozygous females or abo homozygotes from lines that do not show abo expression.     

Accumulation of Deleterious Genes in a Cage Population of DROSOPHILA MELANOGASTER

Lethal and sterility mutations were accumulated in a cage population which was initiated with lethal- and sterility-free second chromosomes of D. melanogaster. It took about 2,000 days for the frequencies of these genes to reach equilibrium levels, i.e., 18% lethal and 9% male-sterile chromosomes. Two other cage populations which were initiated with random chromosomes sampled from natural populations and kept for more than eleven years in the laboratory showed 19-20% lethal content. The elimination rates of lethals by homozygosis in these populations were smaller than the mutation rate. By using NEI's formulae, the deleterious effect of a lethal gene in heterozygous condition (h) was estimated to be 0.035. The effective population number in the cage populations was estimated to be 1,000-2,900, while the actual population number was 3,500-7,800.     

Centromeric Effect on the Degree of Nonrandom Disjunction in the Female DROSOPHILA MELANOGASTER

From crosses of females possessing a heteromorphic X-chromosome bivalent, FR1/+, the shorter crossover products were recovered on the average more frequently than the longer reciprocals as predicted by Novitski's (1951) hypothesis of nonrandom disjunction (NRD). The present study stemmed from an unexpected result of these crosses. Evidence for a centromeric effect on NRD was obtained, suggested by a negative correlation between the degree of NRD, c, and the distance between the region of exchange and the centromere as inferred from SET's (single exchange tetrads). Studies on sex chromosome systems other than FR1 confirmed these results. An analogous centromeric effect on preferential segregation had been clearly demonstrated in maize (Kikudome 1958, 1959; Rhoades and Dempsey 1966). However, prior to the present investigation, no such effect of the centromere on NRD in Drosophila had been described, although reanalysis of part of the data of Novitski (1951) and Novitski and Sandler (1956) suggests some evidence of a seriation of increasing c values extending from the most distal region of the chromosome toward the centromere. A suggestion that the effect in Drosophila may be related in some way to the time required for chiasma terminalization, i.e., those terminalizing earlier (distally located crossovers) permitting more random disjunction of the chromatids from the asymmetric dyad and those terminalizing later, progressively less random, is considered and rejected since in general the expected pattern of c values for the various double exchange tetrads (DET's) is inconsistent with that prediction and provides evidence suggesting the possibility of reversals, in part, of c values obtained for SET's.     

Mutation Rate and Dominance of Genes Affecting Viability in DROSOPHILA MELANOGASTER

Spontaneous mutations were allowed to accumulate in a second chromosome that was transmitted only through heterozygous males for 40 generations. At 10-generation intervals the chromosomes were assayed for homozygous effects of the accumulated mutants. From the regression of homozygous viability on the number of generations of mutant accumulation and from the increase in genetic variance between replicate chromosomes it is possible to estimate the mutation rate and average effect of the individual mutants. Lethal mutations arose at a rate of 0.0060 per chromosome per generation. The mutants having small effects on viability are estimated to arise with a frequency at least 10 times as high as lethals, more likely 20 times as high, and possibly many more times as high if there is a large class of very nearly neutral mutations.-The dominance of such mutants was measured for chromosomes extracted from a natural population. This was determined from the regression of heterozygous viability on that of the sum of the two constituent homozygotes. The average dominance for minor viability genes in an equilibrium population was estimated to be 0.21. This is lower than the value for new mutants, as expected since those with the greatest heterozygous effect are most quickly eliminated from the population. That these mutants have a disproportionately large heterozygous effect on total fitness (as well as on the viability component thereof) is shown by the low ratio of the genetic load in equilibrium homozygotes to that of new mutant homozygotes.