Site-Specific Instability in DROSOPHILA MELANOGASTER: The Origin of the Mutation and Cytogenetic Evidence for Site Specificity

During a study of delayed mutations, an unstable X chromosome (Uc) was detected. Spontaneous X-linked recessive lethal mutations were detected in 34 of 993 sperm sampled from 50 males carrying this chromosome. All but three of the 34 lethals originated as clusters in three of the 50 males Cytogenetic and complementation analyses revealed 14 intrachromosomal rearrangements: ten inversions, two reverse repeats, one deficiency and one transposition. Eight of the 14 rearrangements have one break in the 6F1-2 doublet and two rearrangements have a break in 6F1-5 of the X chromosome. The remaining four rearrangements have in addition to the aberrations a lethal point mutation between 6F1 and 6F5. Though each of the lethal lines was established from a single lethal-bearing female, chromosome polymorphism is evident in 17 of the 18 lines having rearrangements, with certain aberrations recurring in several lines. The lethal mutations revert frequently to the nonlethal state, and cytological evidence indicates that more than one mutational event may occur at the unstable locus of the chromosome during one generation. Two lethal lines had more than one type of chromosome rearrangement sharing a common breakpoint. These observations are consistent with the view that the instability of the Uc lines is caused by a transposable element capable of site-specific chromosome breaks and perpetual generation of mutations. The mutagenic and genetic properties of transposable elements can be related to the two-mutation theory of KNUDSON (1971) for cancer initiation.     

Genetic Instability in Drosophila Melanogaster Mediated by Hobo Transposable Elements

Eight independent recessive lethal mutations that occurred on derivatives of an unstable X chromosome (Uc) in Drosophila melanogaster were analyzed by a combination of genetic and molecular techniques. Seven of the mutations were localized to complementation groups in polytene chromosome bands 6E; 7A. In situ hybridization and genomic Southern analysis established that hobo transposable elements were associated with all seven of the mutations. Six mutations involved deletions of DNA, some of which were large enough to be seen cytologically, and in each case, a hobo element was inserted at the junction of the deletion's breakpoints. A seventh mutation was associated with a small inversion between 6F and 7A-B and a hobo element was inserted at one of its breakpoints. One of the mutant chromosomes had an active hobo-mediated instability, manifested by the recurrent production of mutations of the carmine (cm) locus in bands 6E5-6. This instability persisted for many generations in several sublines of an inbred stock. Two levels of instability, high and basal, were distinguished. Sublines with high instability had two hobo elements in the 6E-F region and produced cm mutations by deleting the segment between the two hobos; a single hobo element remained at the junction of the deletion breakpoints. Sublines with low instability had only one hobo element in the 6E-F region, but they also produced deletion mutations of cm. Both types of sublines also acquired hobo-mediated inversions on the X chromosome. Collectively, these results suggest that interactions between hobo elements are responsible for the instability of Uc. It is proposed that interactions between widely separated elements produce gross rearrangements that restructure the chromosome and that interactions between nearby elements cause regional instabilities manifested by the recurrence of specific mutations. These regional instabilities may arise when a copy of hobo transposes a short distance, creating a pair of hobos that can interact to produce small rearrangements.     

Site-specific instability in Drosophila melanogaster: evidence for transposition of destabilizing element

In this study, we show that at least one lethal mutation at the 3F-4A region of the X chromosome can generate an array of chromosome rearrangements, all with one chromosome break in the 3F-4A region. The mutation at 3F-4A (secondary mutation) was detected in an X chromosome carrying a reverse mutation of an unstable lethal mutation, which was mapped in the 6F1-2 doublet (primary mutation). The primary lethal mutation at 6F1-2 had occurred in an unstable chromosome (Uc) described previously (LIM 1979). Prior to reversion, the fF1-2 doublet was normal and stable, as was the 3F-4A region in the X chromosome carrying the primary lethal mutation. The disappearance of the instability having a set of genetic properties at one region (6F1-2) accompanied by its appearance elsewhere in the chromosome (3F-4A) implies that a transposition of the destabilizing element took place. The mutant at 3F-4A and other secondary mutants exhibited all but one (reinversion of an inversion to the normal sequence) of the eight properties of the primary lethal mutations. These observations support the view that a transposable destabilizing element is responsible for the hypermutability observed in the unstable chromosome and its derivatives.     

The Cytogenetic Analysis of a Fractured Gene in Drosophila

The data presented in this study are derived from the analyses of Notch mutants known to be associated with visible cytological deficiencies. One mutant, Df(1)N(62b1), described as a right-side deficiency, bears a deletion that apparently initiates within the Notch locus and extends to the right as far as the locus of dm. Recombination experiments using heterozygotes of Df(1)N(62b1) with a series of intragenic point mutants within the Notch cistron suggest that this deficiency represents a deletion for the right-end portion of the gene. A consideration of the cytology of Df(1)N(62b1) supports the cytogenetic inference that, if a Notch locus-3C7 relationship is valid, the missing portion of the gene as assayed by recombination experiments has an interband position between 3C7 and 8.-The data derived from two left-side deficiencies with a genetic lesion in Notch and a deletion extending to w are somewhat equivocal, but they do support the presumed Notch locus-3C7 band relationship and thereby enhance the likelihood that Df(1)N(62b1) is correctly interpreted.-Cytogenetic information presently available suggests that, although a significant portion of the Notch cistron has a position on the salivary map identified as interband 3C7 to 8, the 3C7 band is part of the total picture of the Notch gene.     

The Role of Dosage of the Region 7d1–7d5-6 of the X Chromosome in the Production of Homeotic Transformations in DROSOPHILA MELANOGASTER

A high frequency of homeotic transformations appears in Df(3)red/+ progeny of Df(1)sn^C128 /+ females. Generally, the metathoracic appendages are partially transformed into mesothoracic ones. Df(1)sn^C128 includes a small region of the X chromosome: 7D1 to 7D5-6. Hypodosage of this region is mainly effective at the level of the maternal genotype, and the effect is probably due to hypodosage of the wild-type allele of the gene fs(1)h. Df(3)red has an effect that is mainly, if not exclusively, zygotic, probably due to hypodosage of the wild-type allele of Rg-bx. The frequencies of transformed flies resulting from the interaction between Df(1)sn^C128 and Df(3)red are not very sensitive to external conditions and genetic background. Studies of the interactions between Df(1)sn^C128 and other mutations or deficiencies of chromosome 3 [Rg-pbx, bx, pbx, Ubx¹, Ubx¹³⁰, Ubx⁸⁰, Df(3)P9] reveal an analogy between the hypodosage effect of region 7D1–7D5-6 and the effects of ether treatment of blastoderm stage eggs. The role of the gene fs(1)h in the process of segment determination is discussed in the light of these results.     

Molecular Characterization of Hobo-Mediated Inversions in Drosophila Melanogaster

The structure of chromosomal inversions mediated by hobo transposable elements in the Uc-1 X chromosome was investigated using cytogenetic and molecular methods. Uc-1 contains a phenotypically silent hobo element inserted in an intron of the Notch locus. Cytological screening identified six independent Notch mutations resulting from chromosomal inversions with one breakpoint at cytological position 3C7, the location of Notch. In situ hybridization to salivary gland polytene chromosomes determined that both ends of each inversion contained hobo and Notch sequences. Southern blot analyses showed that both breakpoints in each inversion had hobo-Notch junction fragments indistinguishable in structure from those present in the Uc-1 X chromosome prior to the rearrangements. Polymerase chain reaction amplification of the 12 hobo-Notch junction fragments in the six inversions, followed by DNA sequence analysis, determined that each was identical to one of the two hobo-Notch junctions present in Uc-1. These results are consistent with a model in which hobo-mediated inversions result from homologous pairing and recombination between a pair of hobo elements in reverse orientation.     

A New Hybrid Rescue Allele in Drosophila melanogaster

Crosses of Drosophila melanogaster females to males of its sibling species Drosophila simulans, Drosophila mauritiana and Drosophila sechellia produce no sons and daughters that are viable only at low temperatures. We describe here a novel rescue allele Df(1)EP307-1-2 isolated on the basis of its suppression of high temperature hybrid female lethality. Df(1)EP307-1-2 also rescues hybrid males to the pharate adult stage, the same stage at which it is lethal to D. melanogaster pure species males. Molecular analysis indicates that Df(1)EP307-1-2 is associated with a deletion of about 61 kb in the 9D region of the X chromosome. The structure of Df(1)EP307-1-2 suggests that it was formed by a process similar to P-element induced male recombination.     

Fine cytogenetical analysis of the band 10A1-2 and the adjoining regions in the Drosophila melanogaster X chromosome

The X chromosome region 9F12-10A7 (7 bands removed by Df(1)v ^l3) was saturated with lethal, semi-lethal, visible and male sterile mutations. A total of 11 complementation groups were found. In the more narrow interval of Df(1)v ^l1 which removes 3 bands (10A1-2, 10A3, 10A4-5) 6 loci were localised. — The band 10A1-2 consists of a sereis of 5 different subunits: (i) silent DNA where no functions were found — at the distal edge of the band; (ii) and (iii) two genes: v and 1(1)BP4; (iv) silent DNA in middle of the band, (v) locus sev on the proximal edge of the band. About 70% of the band's DNA was found to be silent. — Using the set of chromosome rearrangements removing different parts of the band it was shown that these five sequences may function independently from each other.     

Kinase activity and genetic characterization of a growth related antigen of Drosophila

The Drosophila developmental antigen recognized by the monoclonal antibody F7D6 is expressed in dividing embryonic and imaginal cells but is lost from all differentiating tissues except electrogenic cells of the nervous system and spontaneously contracting muscles. The 63 kDa antigen is associated with the inner surface of plasma membranes and is expressed in several classes of tumorous mutants of Drosophila. The monoclonal antibody was used for immunoprecipitating the antigen for biochemical characterization and for screening expression vector cDNA libraries. Here we report that this oncodevelopmental antigen is a phosphoprotein and a serine-threonine specific protein kinase. A 1.6 kb cDNA isolated by immunological screening of an ovarian library hybridized to a single band on polytene chromosomes, localizing the gene to 72F on the left arm of the third chromosome. Immunofluorescence assays of deficiency stocks in the region confirmed the location of the gene and identity of the cDNA clone, and mapped the gene between the left breakpoints of Df(3L) st100.62 and Df(3L) stj7, i.e., between 72F3-7 and 73A1-2. The biochemical and genetic properties indicate that this is a novel growth-related kinase of Drosophila.     

Cytogenetic analysis of variegation suppressors and a dominant temperature-sensitive lethal in region 23–26 of chromosome 2L in Drosophila melanogaster

Three suppressor loci for position-effect variegation, one dominant temperature-sensitive (DTS), three Minute genes, and two recessive visible mutants (ed, tkv) have been cytogenetically localized by using duplications and deficiencies in regions 23-25 of chromosome arm 2L of Drosophila melanogaster. Two of the suppressor loci studied proved to represent haplo-abnormal genes localized in regions 23A6-23F6 and 24E2-25A1, respectively. The third one is a strong triplo-abnormal suppressor mapping in 25F4-26B9 which affects white variegation in wm4h when present in three doses. The l(2)2DTS mutation, which belongs to a group of noncomplementing dominant temperature-sensitive mutations, is localized in the 25A4-B1 region. Furthermore, two Minute genes have been localized in region 24 that are included in Df(2L)M11 and can be separated employing translocation (Y;2)P8 (24E2-4): M(2)LS2 in 24D3-4-24E2-4, and M(2)z in 24E4-5-24F5-7. A third Minute gene (M(2)S1) is localized in 25C3-8-25C9-D1. The usefulness of the isolated chromosomal rearrangements for further genetic studies of region 23-26 is discussed.     

The Cytogenetics of a Recessive Visible Mutant Associated with a Deficiency Adjacent to the Notch Locus in DROSOPHILA MELANOGASTER

The recessive visible faswb allele in Drosophila is an interband deletion between salivary band 3C5, 6 and 7. Heterozygosity for the deletion does not suppress recombination between faswb and mutant sites at Notch adjacent to it.--Df(1)w67k30, deficient for salivary bands 3C2 to 6, is the left of faswb. By crossing over within the homologous bit of interband retained in w67k30 and faswb, the two deficiencies can be linked. Cytologically, 3C7, "fused" to 3C5,6 in faswb, becomes "fused" to 3C1 when the two are coupled. In the double deletion, the recessive visible phenotype of the faswb "allele* is suppressed. Both w67k30 and faswb can be recovered by uncoupling the two deficiencies.--The data suggest that the mutant faswb does not represent a lesion at Notch; the entire gene or locus seems to be present. The interband deletion in faswb has secondarily moved an intact Notch locus to a foreign environment that interferes with its normal function. When faswb is linked to w67k30, the interference is eliminated and normal Notch functions resume.--The position of Notch on the salivary gland chromosome is reviewed in relation to the information obtained in these experiments.     

Interacting Hobo Transposons in an Inbred Strain and Interaction Regulation in Hybrids of Drosophila Melanogaster

A transposable hobo element in the Notch locus of the Uc-1 X chromosome, which does not interfere with the normal expression of the locus, interacts with other hobo elements in the same X chromosome to produce Notch mutations. Almost all of these mutations are associated with deficiencies, inversions or other rearrangements, and hobo elements are present at each of the breakpoints. The Uc-1 X chromosome produces the Notch mutations at a rate of 4-8% in both sexes of flies in a strain that has been inbred for 96 generations. At least two-thirds of the mutations are produced in clusters suggesting that they have originated in mitotic (premeiotic) germ cells of the Uc-1 inbred strain. The interaction of hobo elements in the Uc-1 X chromosome can be repressed by at least two different mechanisms. One found in three inbred strains not related to the Uc-1 strain involves a maternal effect that is not attributable to the actions or products of hobo elements. Repression by this mechanism is manifested by a clear reciprocal cross effect so that the production of Notch mutations is repressed in the daughters of Uc-1 males, but not in the daughters of Uc-1 females. The other mechanism apparently requires genetic factors and/or hobo elements in a particular strain of Oregon-R; complete repression is present in both types of hybrids between Uc-1 and this strain.     

A Cis-Acting Locus That Promotes Crossing over between X Chromosomes in Caenorhabditis Elegans

This study reports the characterization of a cis-acting locus on the Caenorhabditis elegans X chromosome that is crucial for promoting normal levels of crossing over specifically between the X homologs and for ensuring their proper disjunction at meiosis I. The function of this locus is disrupted by the mutation me8, which maps to the extreme left end of the X chromosome within the region previously implicated by studies of X;A translocations and X duplications to contain a meiotic pairing site. Hermaphrodites homozygous for a deletion of the locus (Df/Df) or heterozygous for a deletion and the me8 mutation (me8/Df) exhibit extremely high levels of X chromosome nondisjunction at the reductional division; this is correlated with a sharp decrease in crossing over between the X homologs as evidenced both by reductions in genetic map distances and by the presence of achiasmate chromosomes in cytological preparations of oocyte nuclei. Duplications of the wild-type region that are unlinked to the X chromosome cannot complement the recombination and disjunction defects in trans, indicating that this region must be present in cis to the X chromosome to ensure normal levels of crossing over and proper homolog disjunction. me8 homozygotes exhibit an altered distribution of crossovers along the X chromosome that suggests a defect in processivity along the X chromosome of an event that initiates at the chromosome end. Models are discussed in which the cis-acting locus deleted by the Dfs functions as a meiotic pairing center that recruits trans-acting factors onto the chromosomes to nucleate assembly of a crossover-competent complex between the X homologs. This pairing center might function in the process of homolog recognition, or in the initiation of homologous synapsis.     

Penetrance of craniofacial anomalies in mouse models of Smith-Magenis syndrome is modified by genomic sequence surrounding Rai1: not all null alleles are alike

Craniofacial abnormality is one of the major clinical manifestations of Smith-Magenis syndrome (SMS). Previous analyses in a mixed genetic background of several SMS mouse models--including Df(11)17/+ and Df(11)17-1/+, which have 2-Mb and 590-kb deletions, respectively, and Rai1(-/+)--revealed that the penetrance of the craniofacial phenotype appears to be influenced by deletion size and genetic background. We generated an additional strain with a 1-Mb deletion intermediate in size between the two described above. Remarkably, the penetrance of its craniofacial anomalies in the mixed background was between those of Df(11)17 and Df(11)17-1. We further analyzed the deletion mutations and the Rai1(-/+) allele in a pure C57BL/6 background, to control for nonlinked modifier loci. The penetrance of the craniofacial anomalies was markedly increased for all the strains in comparison with the mixed background. Mice with Df(11)17 and Df(11)17-1 deletions had a similar penetrance, suggesting that penetrance may be less influenced by deletion size, whereas that of Rai1(-/+) mice was significantly lower than that of the deletion strains. We hypothesize that potential trans-regulatory sequence(s) or gene(s) that reside within the 590-kb genomic interval surrounding Rai1 are the major modifying genetic element(s) affecting the craniofacial penetrance. Moreover, we confirmed the influence of genetic background and different deletion sizes on the phenotype. The complicated control of the penetrance for one phenotype in SMS mouse models provides tools to elucidate molecular mechanisms for penetrance and clearly shows that a null allele caused by chromosomal deletion can have different phenotypic consequences than one caused by gene inactivation.     

An "unusual" stress-activated Drosophila virilis 20CD puff: cytologic localization of the thermostable locus and characteristics of the reaction to heat shock

Using a series of chromosome aberrations heat shock (hs) inducible site of 20CD Drosophila virilis hs puff was cytologically localized in distal 20C subdivision: the hs puff was not observed in chromosome 2 carrying Df(2)eb deficiency; the bands in proximal 20D subdivision are unable to provide autonomous activation by hs in aberrant chromosomes having this deficiency or the In(2)eb inversion. Electron microscopical analysis of successive stages of 20CD development showed the hs puff to be initially formed from the right part of 20C 6-7 band. Neighbouring bands of 20C and 20D regions in the most developed puff are only partially decondensed and were detected as discrete compact chromatin clumps.     

Congenital heart defect and mental retardation in a patient with a 13q33.1-34 deletion

13q deletion syndrome is a rare genetic disorder caused by deletions of the long arm of chromosome 13. Patients with 13q deletion display a variety of phenotypic features. We describe a one-year-old female patient with congenital heart defects (CHD), facial anomalies, development and mental retardation. We identified a 12.75Mb deletion in chromosome region 13q33.1-34 with high resolution SNP Array (Human660W-Quad, Illumina, USA). This chromosome region contains about 55 genes, including EFNB2, ERCC5, VGCNL1, F7, and F10. Comparing our findings with previously reported 13q deletion patients with congenital heart defects, we propose that the 13q33.1-34 deletion region might contain key gene(s) associated with cardiac development. Our study also identified a subclinical deficiency of Factors VII and X in our patient with Group 3 of 13q deletion syndrome.     

Analysis of linkage between biochemical and morphological markers of 1R-, 2R-, and 5R-chromosomes in rye with autofertility mutations in main incompatibility loci

In F2 hybrids between self-sterile plants of the Volkhova cultivar and self-fertile lines with established self-fertility mutations (sf-mutations) at the major incompatibility loci S (1R), Z (2R), and T (5R), the effect of sf-mutations on the inheritance of secalin-encoding, isozyme, and morphological markers located on the same chromosomes was investigated. Linkage between loci Prx7 and S and locus Sec3 coding for high-molecular-weight secalins on chromosome 1R was shown for the first time The frequency of recombination between Prx7 and Sec3 and between S and Sec3 was 29.1 +/- 4.8% and 30.9 +/- 7.0%, respectively. Independent inheritance of locus Z and isozyme markers of chromosome 2R, Est3/5 and beta-Glu, from locus Sec2 encoding 75-kDa gamma-secalins was shown; in hybrids, the recombination frequency between Est3/5 and locus Z varied from 19.2 +/- 8.1 to 50%. Independent inheritance of morphological (Ddw and Hs) and isozyme markers (Est4, Est6/9, and Aco2) of chromosome 5R from locus T located on the same chromosome was demonstrated.