A developmental and molecular analysis of Cdc2 mutations in Drosophila melanogaster

Genome ◽  
1993 ◽  
Vol 36 (4) ◽  
pp. 676-685 ◽  
Author(s):  
N. J. Clegg ◽  
I. P. Whitehead ◽  
J. A. Williams ◽  
G. B. Spiegelman ◽  
T. A. Grigliatti
Keyword(s):  
Drosophila Melanogaster ◽  
Null Allele ◽  
Donor Site ◽  
Pupal Stage ◽  
P Element ◽  
Splice Donor Site ◽  
Coding Region ◽  
Cdc2 Gene ◽  
Exon 1 ◽  
Single Base Pair

In fission yeast, the product of the cdc2 gene is required both for entry into S phase and mitosis. Homologs of cdc2 have been isolated from a number of metazoans, but in general they have not been amenable to genetic analysis. Here we describe P element transposon tagging of Cdc2 in Drosophila melanogaster and the analysis of 10 Cdc2 mutants. The recessive lethality of Cdc2P is associated with a P element located in the 5′ untranslated region of the gene. Seven other alleles have unique single base pair substitutions in the coding region of Cdc2. One allele, Cdc2B47, is mutated in the splice donor site of exon 1. Most mutations in Cdc2, including the presumptive null allele Cdc2B47, die at the pupal stage, suggesting that the maternally supplied Cdc2 gene product drives earlier cell divisions. The phenotypes of our mutants are consistent with a role for Cdc2 in cell proliferation; however, we did not observe any perturbation of the endoreduplication cycle associated with the acquisition of polyteny.Key words: Cdc2, Drosophila, mutations, sequence.

scholarly journals Structure and transcription of the Drosophila melanogaster vermilion gene and several mutant alleles

1990 ◽  
Vol 10 (4) ◽  
pp. 1423-1431
Author(s):  
L L Searles ◽  
R S Ruth ◽  
A M Pret ◽  
R A Fridell ◽  
A J Ali
Keyword(s):  
Drosophila Melanogaster ◽  
Transcription Initiation ◽  
Donor Site ◽  
Initiation Site ◽  
Splice Donor Site ◽  
Wild Type ◽  
Upstream Site ◽  
Reading Frame ◽  
Exon 1 ◽  
A Minor

The nucleotide sequence and intron-exon structure of the Drosophila melanogaster vermilion (v) gene have been determined. In addition, the sites of several mutations and the effects of these mutations on transcription have been examined. The major v mRNA is generated upon splicing six exons of lengths (5' to 3') 83, 161, 134, 607, 94, and 227 nucleotides (nt). A minor species of v mRNA is initiated at an upstream site and has a 5' exon of at least 152 nt which overlaps the region included in the 83-nt exon of the major v RNA. The three v mutations, v1, v2, and vk, which can be suppressed by mutations at suppressor of sable, su(s), are insertions of transposon 412 at the same position in exon 1, 36 nt downstream of the major transcription initiation site. Despite the 7.5-kilobase insertion in these v alleles, a reduced level of wild-type-sized mRNA accumulates in suppressed mutant strains. The structure and transcription of several unsuppressible v alleles have also been examined. The v36f mutation is a B104/roo insertion in intron 4 near the splice donor site. A mutant carrying this alteration accumulates a very low level of mRNA that is apparently polyadenylated at a site within the B104/roo transposon. The v48a mutation, which deletes approximately 200 nt of DNA, fuses portions of exons 3 and 4 without disruption of the translational reading frame. A smaller transcript accumulates at a wild-type level, and thus an altered, nonfunctional polypeptide is likely to be synthesized in strains carrying this mutation.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Structure and transcription of the Drosophila melanogaster vermilion gene and several mutant alleles.

1990 ◽  
Vol 10 (4) ◽  
pp. 1423-1431 ◽  
Author(s):  
L L Searles ◽  
R S Ruth ◽  
A M Pret ◽  
R A Fridell ◽  
A J Ali
Keyword(s):  
Drosophila Melanogaster ◽  
Transcription Initiation ◽  
Donor Site ◽  
Initiation Site ◽  
Splice Donor Site ◽  
Wild Type ◽  
Upstream Site ◽  
Reading Frame ◽  
Exon 1 ◽  
A Minor

The nucleotide sequence and intron-exon structure of the Drosophila melanogaster vermilion (v) gene have been determined. In addition, the sites of several mutations and the effects of these mutations on transcription have been examined. The major v mRNA is generated upon splicing six exons of lengths (5' to 3') 83, 161, 134, 607, 94, and 227 nucleotides (nt). A minor species of v mRNA is initiated at an upstream site and has a 5' exon of at least 152 nt which overlaps the region included in the 83-nt exon of the major v RNA. The three v mutations, v1, v2, and vk, which can be suppressed by mutations at suppressor of sable, su(s), are insertions of transposon 412 at the same position in exon 1, 36 nt downstream of the major transcription initiation site. Despite the 7.5-kilobase insertion in these v alleles, a reduced level of wild-type-sized mRNA accumulates in suppressed mutant strains. The structure and transcription of several unsuppressible v alleles have also been examined. The v36f mutation is a B104/roo insertion in intron 4 near the splice donor site. A mutant carrying this alteration accumulates a very low level of mRNA that is apparently polyadenylated at a site within the B104/roo transposon. The v48a mutation, which deletes approximately 200 nt of DNA, fuses portions of exons 3 and 4 without disruption of the translational reading frame. A smaller transcript accumulates at a wild-type level, and thus an altered, nonfunctional polypeptide is likely to be synthesized in strains carrying this mutation.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Genetic and Molecular Characterization of sting, a Gene Involved in Crystal Formation and Meiotic Drive in the Male Germ Line of Drosophila melanogaster

Genetics ◽  
1999 ◽  
Vol 151 (2) ◽  
pp. 749-760 ◽  
Author(s):  
Armin Schmidt ◽  
Gioacchino Palumbo ◽  
Maria P Bozzetti ◽  
Patrizia Tritto ◽  
Sergio Pimpinelli ◽  
...  
Keyword(s):  
Amino Acids ◽  
Drosophila Melanogaster ◽  
Null Allele ◽  
Germ Line ◽  
Meiotic Drive ◽  
P Element ◽  
Crystal Formation ◽  
Male Sterile ◽  
Male Sterile Mutation

Abstract The sting mutation, caused by a P element inserted into polytene region 32D, was isolated by a screen for male sterile insertions in Drosophila melanogaster. This sterility is correlated with the presence of crystals in spermatocytes and spermatids that are structurally indistinguishable from those produced in males carrying a deficiency of the Y-linked crystal (cry) locus. In addition, their morphology is needle-like in Ste+ flies and star-shaped in Ste flies, once again as observed in cry– males. The sti mutation leads to meiotic drive of the sex chromosomes, and the strength of the phenomenon is correlated with the copy number of the repetitive Ste locus. The same correlation is also true for the penetrance of the male sterile mutation. A presumptive sti null allele results in male sterility and lethal maternal effect. The gene was cloned and shown to code for a putative protein that is 866 amino acids long. A C-terminal domain of 82 amino acids is identified that is well conserved in proteins from different organisms. The gene is expressed only in the germline of both sexes. The interaction of sting with the Ste locus can also be demonstrated at the molecular level. While an unprocessed 8-kb Ste primary transcript is expressed in wild-type males, in X/Y homozygous sti males, as in X/Y cry– males, a 0.7-kb mRNA is produced.

scholarly journals The Gene Search System: A Method for Efficient Detection and Rapid Molecular Identification of Genes in Drosophila melanogaster

Genetics ◽  
1999 ◽  
Vol 151 (2) ◽  
pp. 725-737 ◽  
Author(s):  
Gakuta Toba ◽  
Takashi Ohsako ◽  
Naomasa Miyata ◽  
Tsuyoshi Ohtsuka ◽  
Ki-Hyeon Seong ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Core Promoter ◽  
Sequence Similarity ◽  
Direct Sequencing ◽  
P Element ◽  
Drosophila Genome ◽  
Coding Region ◽  
New Genes ◽  
Gene Search ◽  
Efficient Detection

Abstract We have constructed a P-element-based gene search vector for efficient detection of genes in Drosophila melanogaster. The vector contains two copies of the upstream activating sequence (UAS) enhancer adjacent to a core promoter, one copy near the terminal inverted repeats at each end of the vector, and oriented to direct transcription outward. Genes were detected on the basis of phenotypic changes caused by GAL4-dependent forced expression of vector-flanking DNA, and the transcripts were identified with reverse transcriptase PCR (RT-PCR) using the vector-specific primer and followed by direct sequencing. The system had a greater sensitivity than those already in use for gain-of-function screening: 64% of the vector insertion lines (394/613) showed phenotypes with forced expression of vector-flanking DNA, such as lethality or defects in adult structure. Molecular analysis of 170 randomly selected insertions with forced expression phenotypes revealed that 21% matched the sequences of cloned genes, and 18% matched reported expressed sequence tags (ESTs). Of the insertions in cloned genes, 83% were upstream of the protein-coding region. We discovered two new genes that showed sequence similarity to human genes, Ras-related protein 2 and microsomal glutathione S-transferase. The system can be useful as a tool for the functional mapping of the Drosophila genome.

Tau gene mutations and neurodegeneration

2001 ◽  
Vol 67 ◽  
pp. 59-71 ◽  
Author(s):  
Michel Goedert ◽  
Maria Grazia Spillantini
Keyword(s):  
Tau Protein ◽  
Donor Site ◽  
Gene Mutations ◽  
Corticobasal Degeneration ◽  
Chromosome 17 ◽  
Splice Donor Site ◽  
Coding Region ◽  
Tau Pathology ◽  
Tau Isoforms ◽  
Exon 10

Abundant neurofibrillary lesions made of the microtubule-associated protein tau constitute a defining neuropathological characteristic of Alzheimer's disease. Filamentous tau protein deposits are also the defining neuropathological characteristic of other neurodegenerative diseases, many of which are frontotemporal dementias or movement disorders, such as Pick's disease, progressive supranuclear palsy and corticobasal degeneration. It is well established that the distribution of tau pathology correlates with the presence of symptoms of disease. However, until recently, there was no genetic evidence linking dysfunction of tau protein to neurodegeneration and dementia. This has now changed with the discovery of close to 20 mutations in the tau gene in frontotemporal dementia with Parkinsonism linked to chromosome 17. All cases with tau mutations examined to date have shown an abundant filamentous tau pathology in brain cells. Pathological heterogeneity is determined to a large extent by the location of mutations in tau. Known mutations are either coding region or intronic mutations located close to the splice-donor site of the intron downstream of exon 10. Most coding region mutations produce a reduced ability of tau to interact with microtubules. Several of these mutations also promote sulphated glycosaminoglycan-induced assembly of tau into filaments. Intronic mutations and some coding region mutations produce increased splicing in of exon 10, resulting in an overexpression of four-repeat tau isoforms. Thus a normal ratio of three-repeat to four-repeat tau isoforms is essential for preventing the development of tau pathology. The new work has shown that dysfunction of tau protein can cause neurodegeneration and dementia.

scholarly journals A novel pathway for alternative splicing: identification of an RNA intermediate that generates an alternative 5' splice donor site not present in the primary transcript of AMPD1.

1990 ◽  
Vol 10 (10) ◽  
pp. 5271-5278 ◽  
Author(s):  
I Mineo ◽  
P R Clarke ◽  
R L Sabina ◽  
E W Holmes
Keyword(s):  
Alternative Splicing ◽  
Donor Site ◽  
Specific Pattern ◽  
Splice Donor Site ◽  
Splice Donor ◽  
Primary Transcript ◽  
Exon 2 ◽  
Tissue Specific ◽  
Exon 1 ◽  
Alternatively Spliced

AMP deaminase (AMPD) is a central enzyme in eucaryotic energy metabolism, and tissue-specific as well as stage-specific isoforms are found in many vertebrates. This study demonstrates the AMPD1 gene product in rat is alternatively spliced. The second exon, a 12-base miniexon, was found to be excluded or included in a tissue-specific and stage-specific pattern. This example of cassette splicing utilizes a unique pathway through an RNA intermediate that generates an alternative 5' splice donor site at the point where exon 2 is ligated to exon 1. In the analogous intermediate of human AMPD1, the potential 5' splice donor site created at the boundary of exon 1 and exon 2 was a poor substrate for splicing because of differences in exon 2 sequences, and human AMPD1 was not alternatively spliced. These results demonstrate that in some cases alternative splicing may proceed through an RNA intermediate that generates an alternative splice donor site not present in the primary transcript. Discrimination between alternative 5' splice donor sites in the RNA intermediate of AMPD1 is apparently controlled by tissue-specific and stage-specific signals.

scholarly journals The congested-like tracheae Gene of Drosophila melanogaster Encodes a Member of the Mitochondrial Carrier Family Required for Gas-Filling of the Tracheal System and Expansion of the Wings After Eclosion

Genetics ◽  
1997 ◽  
Vol 147 (4) ◽  
pp. 1755-1768 ◽  
Author(s):  
Kirsten Hartenstein ◽  
Pradip Sinha ◽  
Arati Mishra ◽  
Heide Schenkel ◽  
Istvan Török ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Tandem Repeats ◽  
Polytene Chromosomes ◽  
P Element ◽  
Coding Region ◽  
Tracheal System ◽  
Mitochondrial Carrier ◽  
Transmembrane Segments ◽  
Hatching Time ◽  
Mitochondrial Carrier Family

Abstract A recessive semi-lethal mutation resulting from the insertion of a P-lacW transposon at the cytological position 23A on the polytene chromosomes of Drosophila melanogaster was found to affect the unfolding and expansion of the wings resulting in a loss of venation and a marked decrease in their size. Lethality was polyphasic with numerous animals dying during early larval development and displaying apparently collapsed tracheal trees. The gene was therefore designated as congested-like tracheae, or colt. The colt mutation resulted from the insertion of a P-lacW transposon within the coding region of a 1.4kb transcript. Wild-type function was restored by inducing a precise excision of the P-lacWtransposon, while a deletion of the colt locus, produced by imprecise excision of the P element, showed a phenotype similar to that of the original P insert. The colt gene consists of a single exon and encodes a protein of 306 amino acids made of three tandem repeats, each characterized by two predicted transmembrane segments and a loop domain. The COLT protein shares extensive homology with proteins in the mitochondrial carrier family and particularly with the DIF-1 protein of Caenwhabditis ekgans, which has been shown to be maternally required for embryonic tissue differentiation. Our analysis revealed that zygotic colt function is dispensable for normal embryonic morphogenesis but is required for gas-filling of the tracheal system at hatching time of the embryo and for normal epithelial morphogenesis of the wings.

Sites of P element insertion and structures of P element deletions in the 5' region of Drosophila melanogaster RpII215

1986 ◽  
Vol 6 (10) ◽  
pp. 3312-3319
Author(s):  
L L Searles ◽  
A L Greenleaf ◽  
W E Kemp ◽  
R A Voelker
Keyword(s):  
Drosophila Melanogaster ◽  
P Element ◽  
Nucleotide Sequencing ◽  
Insertion Mutation ◽  
Activity Levels ◽  
Coding Region ◽  
Element Insertion ◽  
Insertion And Deletion ◽  
Transcribed Sequences ◽  
Specific Sequences

Several P element insertion and deletion mutations near the 5' end of Drosophila melanogaster RpII215 have been examined by nucleotide sequencing. Two different sites of P element insertion, approximately 90 nucleotides apart, have been detected in this region of the gene. Therefore, including an additional site of P element insertion within the coding region, there are at least three distinct sites of P element insertion at RpII215. Both 5' sites are within a noncoding portion of transcribed sequences. The sequences of four revertants of one P element insertion mutation (D50) indicate that the P element is either precisely deleted or internally deleted to restore RpII215 activity. Partial internal deletions of the P element result in different RpII215 activity levels, which appear to depend on the specific sequences that remain after excision.

scholarly journals Sites of P element insertion and structures of P element deletions in the 5' region of Drosophila melanogaster RpII215.

1986 ◽  
Vol 6 (10) ◽  
pp. 3312-3319 ◽  
Author(s):  
L L Searles ◽  
A L Greenleaf ◽  
W E Kemp ◽  
R A Voelker
Keyword(s):  
Drosophila Melanogaster ◽  
P Element ◽  
Nucleotide Sequencing ◽  
Insertion Mutation ◽  
Activity Levels ◽  
Coding Region ◽  
Element Insertion ◽  
Insertion And Deletion ◽  
Transcribed Sequences ◽  
Specific Sequences

Several P element insertion and deletion mutations near the 5' end of Drosophila melanogaster RpII215 have been examined by nucleotide sequencing. Two different sites of P element insertion, approximately 90 nucleotides apart, have been detected in this region of the gene. Therefore, including an additional site of P element insertion within the coding region, there are at least three distinct sites of P element insertion at RpII215. Both 5' sites are within a noncoding portion of transcribed sequences. The sequences of four revertants of one P element insertion mutation (D50) indicate that the P element is either precisely deleted or internally deleted to restore RpII215 activity. Partial internal deletions of the P element result in different RpII215 activity levels, which appear to depend on the specific sequences that remain after excision.

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