[A Study of the Fertility of a Drosophila melanogaster MS Strain with Impaired Transposition Control of the gypsy Mobile Element].

The flamenco locus is one of the main components of the piRNA pathway of regulation of mobile genetic elements (MGEs) in Drosophila melanogaster. Mutations at this locus lead to an increase in the transposition activity of MGEs and, as a result, to genetic instability. In this paper, the fertility of a genetically unstable MS strain obtained more than 25 years ago and characterized by a mutation in the flamenco locus and the presence of a functionally active copy of gypsy retrotransposon was investigated. Complex violations of the ovarian morphology were revealed in the MS strain in females: defects in the follicular layer and ring channels, as well as degradation of trophocytes, which in turn led to a decrease in reproductive abilities. Analysis of the MS strain transcriptome showed a decrease in the expression level of 40 genes encoding chorionic proteins and expression specificity at different stages of follicle development. In the F1 and F2 hybrid females from the crosses of MS females with wild type males, restoration of reproductive abilities was observed, despite the fact that half of the F2 females had the flamenco genotype and genetic instability caused by transposition of gypsy (according to the ovo^(D) test). Moreover, the frequency of gypsy transposition in the hybrid F2 females with the flamenco genotype doubled in comparison with the MS strain females. Thus, the MS strain had acquired partial suppression of the flamenco phenotype and accumulated several recessive mutations in the genes that control oogenesis after cultivation for over 25 years.

Expression of hp1 family genes and their plausible role in formation of flamenco phenotype in D. melanogaster.

Results of expression analysis of transcription of the flamenco locus that controls transposition of the mobile genetic element gypsy, RNA interference system genes ago3, zuc, aub, and HP1 heterochromatin protein family genes hp1a, hp1b, hp1c, hp1d (rhino), and hp1e in D. melanogaster SS strain mutant on the flamenco gene are presented. We show that the number of transcripts in the SS strain that are formed in the flamenco locus is unchanged in some freely chosen points, and this is different from the wild-type strain where a decreased number of transcripts is observed, which clearly is a result of processing of the flamenco locus primary transcript, a predecessor of piRNA. At the same time, expression of genes of the RNA interference system is not affected, but there is a reduced level of hp1d gene expression in ovary tissue. We suggest that the hp1d gene product is directly or indirectly involved in the flamenco locus primary transcript processing.

[Study of the transcriptional and transpositional activities of the Tirant retrotransposon in Drosophila melanogaster strains mutant for the flamenco locus].

Transpositions of the gypsy retrotransposon in the Drosophila melanogaster genome are controlled by the flamenco locus, which is represented as an accumulation of defective copies of transposable elements. In the present work, genetic control by the flamenco locus of the transcriptional and transpositional activities of the Tirant retrotransposon from the gypsy group was studied. Tissue-specific expression of Tirant was detected in the tissues of ovaries in a strain mutant for the flamenco locus. Tirant was found to be transpositionally active in isogenic D. melanogaster strains mutant for the flamenco locus. The sites of two new insertions have been localized by the method of subtractive hybridization. It has been concluded from the results obtained that the flamenco locus is involved in the genetic control of Tirant transpositions.

[Analysis of the structure and expression of the DIP1 gene in Drosophila melanogaster strains mutant for the flamenco gene].

DIP1 gene transcription was analyzed with the use of RT-PCR in three Drosophila melanogaster strains with the flamenco- phenotype (flam(SS), flam(MS), and flam(Ore)) and in one flamenco+ strain at the stages of embryos (0-24 h), third-instar larvae, and adult flies. The mutant strains flam(SS) and flam(Ore) lack an active copy of transposon gypsy. Theflam(MS) strain was obtained by introducing an active copy of gypsy in flies of theflam(SS) strain and is characterized by a high rate of gypsy transpositions. The experiments showed that at least five forms of DIP1 gene transcripts are produced. The form of cDNA corresponding to CDS DIP1-d was discovered only in embryos. It was found that DIP1 gene transcription depends on the age of flies: at the larval stage the level of transcription is significantly reduced. However, no reduction of gene transcription is observed in theflam(Ore) strain. These results suggest that the flamenco- phenotype may be associated with an alteration of DIP1 gene transcription, as in differentflamenco- strains the DIP1 gene expression is changed differently.

[Characteristics of the structural organization of the DIP1 gene in Drosophila melanogaster strains mutant for the flamenco gene].

Molecular cloning of the DIP1 gene located in the 20A4-5 region has been performed from the following strains with the flamenco phenotype: flamSS (SS) and flamMS (MS) characterized by a high transposition rate of retrotransposon gypsy (mdg4), flampy + (P) carrying the insertion of a construction based on the P element into the region of the flamenco gene, and flamenco+. The results of restriction analysis and sequencing cloned DNA fragments has shown that strains flamSS, flamMS, flampy +(P), and flamenco+ considerably differ from one another in the structure of DIP1. Strains flamss and flamMS have no Dral restriction site at position 1765 in the coding region of the gene, specifically, in the domain determining the signal of the nuclear localization of the DIP1 protein. This mutation has been found to consist in a nucleotide substitution in the recognition site of DraI restriction endonuclease, which is transformed from TTTAAA into TTTAAG and, hence, is not recognized by the enzyme. This substitution changes codon AAA into AAG and is translationally insignificant, because both triplets encode the same amino acid, lysine. The Dral gene of strainsflamSS andflamMS has been found to contain a 182-bp insertion denoted IdSS (insertion in DIP1 strain SS); it is located in the second intron of the gene. The IdSS sequence is part of the open reading frame encoding the putative transposase of the mobile genetic element HB1 belonging to the Tcl/mariner family. This insertion is presumed to disturb the conformations of DNA and the chromosome, in particular, by forming loops, which alters the expression of DIPI and, probably, neighboring genes. In strains flamenco+ and flampy + (P), the IdSS insertion within the HB1 sequence is deleted. The deletion encompasses five C-terminal amino acid residues of the conserved domain and the entire C-terminal region of the putative HB1 transposase. The obtained data suggest that DIP1 is involved in the control of gypsy transpositions either directly or through interaction with other elements of the genome.

[Study of male mating behavior in some Drosophila melanogaster strains in experiments with fertilized females]. / Izuchenie eituala ukhazhivaniia samtsov nekotorykh linii Drosophila melanogaster v éksperimentakh s oplodotvorennymi samkami.

Male courtship ritual is among the main behavioral characteristics of Drosophila. This is a complex, genetically determined process consisting of four general stages: orientation, vibration, licking, and attempts at copulation (or successful copulation). Several genes are known that control some stages of this behavior. Most of them have pleiotropic effects and are involved in other biological processes. Earlier, we have shown that a mutation in locus flamenco (20A1-3), which controls transposition and infectivity of retrotransposon gypsy (MDG4), is involved in the genetic control of behavior. In strains mutant for this locus, the male mating activity is decreased and the structure of courtship ritual is changed. To understand the mechanisms of these changes, it is important to study all behavioral stages in genetically identical strains. For this purpose, the normal allele of gene flamenco from the X chromosome of the wild-type strain (stock) Canton S was introduced into strain SS carrying flamMS. This offers new opportunities in studying the role of gene flamenco in the control of mating behavior in Drosophila.

[Male reproductive behavior in Drosophila melanogaster strains with different alleles of the flamenco gene]. / Reproduktivnoe povedenie samtsov v liniiakh Drosophila melanogaster, otlichaiushchikhsia po alleliam gena flamenco.

The allelic state of gene flamenco has been determined in a number of Drosophila melanogaster strains using the ovoD test. The presence of an active copy of gypsy in these strains was detected by restriction analysis. Then male reproduction behavior was studied in the strains carrying a mutation in gene flamenco. In these experiments mating success has been experimentally estimated in groups of flies. It has been demonstrated that the presence of mutant allele flamMS decreases male mating activity irrespective of the presence or absence of mutation white. The active copy of gypsy does not affect mating activity in the absence of the mutation in gene flamenco. Individual analysis has demonstrated that that mutation flamMS results in characteristic changes in courtship: flamMS males exhibit a delay in the transition from the orientation stage to the vibration stage (the so-called vibration delay). The role of locus flamenco in the formation of male mating behavior in Drosophila is discussed.

[Some behavioral features in Drosophila melanogaster lines carrying a flamenco gene mutation]. / Nekotorye povedencheskie osobennosti v liniiakh Drosophila melanogaster mutantnykh po genu flamenco.

Olfactory sensitivity and locomotor activity was assayed in Drosophila melanogaster strains carrying a mutation of the flamenco gene, which controls transposition of the mobile genetic element 4 (MGE4) retrotransposon the gypsy mobile element. A change in olfactory sensitivity was detected. The reaction to the odor of acetic acid was inverted in flies of the mutator strain (MS), which carried the flam mutation and active MGE4 copies and were characterized by genetic instability. Flies of the genetically unstable strains displayed a lower locomotor activity. The behavioral changes in MS flies can be explained by the pleiotropic effect of the flam mutation or by insertion mutations which arise in behavior genes as a result of genome destabilization by MGE4.

[Mating success and courtship ritual in strains of Drosophila melanogaster carrying mutation flamenco]. / Uspeshnost' sparivaniia i osobennosti rituala ukhazhivaniia v liniiakh Drosophila melanogaster, mutantnykh po genu flamenco.

Mating success was examined in groups of Drosophila melanogaster carrying mutation flamMS (SS, MSn1-2, and MSn1-3) and in wild-type flies. The proportion of normally copulating males was significantly lower in the mutant strains. The reduction in mating efficiency was caused by changes in male behavior rather than in female attractiveness. Individual analyses showed that male mating behavior in strains carrying flamMS was qualitatively and quantitatively different from that in the wild-type strain Canton S. The proportion of males that performed consecutive courtship stages was significantly lower in the mutant strains. The sequence and duration of some courtship stages (in particular, orientation and wing vibration) in mutant flies was shown to be altered. The significance of the flamenco locus in regulation of processes occurring at the organismal level are discussed.

[Analysis of spontaneous and ethylmethanesulfonate-induced visible mutations in the genetically unstable MS, w strain of Drosophila melanogaster]. / Analiz spontannykh i indutsirovannykh étilmetansul'fonatom vidimykh mutatsii v geneticheski nestabil'noi linii MS, w Drosophila melanogaster.

We analyzed genetic effects of ethylmethanesulfonate (EMS) on mature sperm of males of the Drosophila melanogaster MS, w strain. The frequency of induced mutations was estimated in the X chromosome, in five loci of chromosome 2 (b, j, pr, cn, and vg), and in eight loci of chromosome 3 (ru, h, th, st, cu, sr, es, and ca). Mutability was significantly increased in the genetically unstable MS, w strain in comparison with the reference laboratory ss, w strain. In MS, w, genetic instability was characteristic of more than 50% EMS-induced mutations. Analysis of the mutation spectrum revealed a specific distribution along the chromosomes. In MS, w, exhibition of spontaneous and induced instability was shown to be similar. This could imply a common mechanism of control of mutability involving transpositions of mobile genetic elements.

[Ethylmethane sulfonate- and X-ray-induced mutability of the loci of chromosome III in normal strains of Drosophila melanogaster and strains defective in excision repair]. / Indutsirovannaia étilmetansul'fonatom i rentgenovskimi luchami mutabel'nost' lokusov III khromosomy v normal'noi i defektnoi po ékstsizionnoi reparatsii liniiakh Drosophila melanogaster.

The frequencies of spontaneous and ethylmethanesulfonate and X-ray-induced mutations in III chromosome of strains D-32 (wild type) and mei-9LI deficient in excision repair have been studied. Mutations have been induced in mature spermatozoa and analysed using multiply marked strain rucuca. It has been shown that the spectra of mutability and frequencies of mutations don't differ in both strains. It indicates the absence of specificity of mutagens studied.

[Analysis of the visible mutations induced by x-rays and ethyl methanesulfonate in the mature spermatozoa of Drosophila melanogaster]. / Analiz vidimykh mutatsii, indutsirovannykh rentgenovskimi luchami i étilmetansul'fonatom v zrelykh spermiiakh Drosophila melanogaster.

The recessive visible mutations spectrum of chromosome II induced by X-rays and ethylmethanesulfonata (EMS) in mature Drosophila melanogaster spermatozoa has been studied. Treatment of both mutagens resulted in mutations in all 5 genes in stock mei-9LI and only 4--in D-32. The comparison of mutation frequencies of the same genes in two stocks under EMS-treatment demonstrated the statistical difference of mutation frequencies j, pr, cn of two stocks, genes b and vg did not differ. Under the influence of X-rays the differencies have been observed only for gene b. In stock D-32 the mutation frequency differes from the control for b and vg (EMS treatment) and j, pr, vg (under the action of X-rays), in mei-9LI--the mutation frequency of all 5 genes (under the X-rays) and 4 of 5 genes (EMS treatment).