Genomic structure and characterization of the Drosophila S3 ribosomal/DNA repair gene and mutant alleles

The Drosophila S3 protein is known to be associated with ribosomes, where it is thought to play a role in the initiation of protein translation. The S3 protein also contains a DNA repair activity, efficiently processing 8-oxoguanine residues in DNA via an N-glycosylase/apurinic-apyrimidinic (AP) lyase activity. The gene that encodes S3 has previously been localized to one of the Minute loci on chromosome 3 in Drosophila. This study focused on the genomic organization of S3 at M(3)95A, initial promoter characterization, and analysis of three mutant alleles at this locus. The S3 gene was found to be a single-copy gene 2 to 3 kb in length and containing a single intron. The upstream 1.6-kb region was analyzed for promoter activity, identifying a presumptive regulatory domain containing potential enhancer and suppressor elements. This finding is of interest, as the S3 gene is constitutively expressed throughout development and mRNA is most likely maternally inherited. Lastly, three Minute alleles from the same locus were sequenced and two alleles found to contain a 22-bp deletion in exon 2, resulting in a truncated S3 protein, although wildtype levels of S3 mRNA and protein were detected in the viable heterozygous Minute alleles, possibly reflecting dosage compensation.     

A Drosophila model for xeroderma pigmentosum and Cockayne's syndrome: haywire encodes the fly homolog of ERCC3, a human excision repair gene.

The haywire gene of Drosophila encodes a protein with 66% identity to the product of the human ERCC3 gene, associated with xeroderma pigmentosum B (XP-B) and Cockayne's syndrome (CS). XP is a human autosomal recessive disease characterized by extreme sensitivity to ultraviolet irradiation and marked susceptibility to skin cancer. In addition, XP and CS patients often exhibit a variety of defects, ranging from central nervous system disorders to hypogonadism. Phenotypes of haywire mutants mimic some of the effects of XP. Many haywire alleles are recessive lethal, viable alleles cause ultraviolet sensitivity, and files expressing marginal levels of haywire display motor defects and reduced life span. Progeny of females carrying a maternal effect allele show central nervous system defects.     

Defects in the DNA repair and transcription gene ERCC2(XPD) in trichothiodystrophy.

Trichothiodystrophy (TTD) is a rare autosomal recessive disorder characterized by brittle hair with reduced sulfur content, ichthyosis, peculiar face, and mental and growth retardation. Clinical photosensitivity is present in approximately 50% of TTD patients but is not associated with an elevated frequency of cancers. Previous complementation studies show that the photosensitivity in nearly all of the studied patients is due to a defect in the same genetic locus that underlies the cancer-prone genetic disorder xeroderma pigmentosum group D (XP-D). Nucleotide-sequence analysis of the ERCC2 cDNA from three TTD cell strains (TTD1V1, TTD3VI, and TTD1RO) revealed mutations within the region from amino acid 713-730 and within previously identified helicase functional domains. The various clinical presentations and DNA repair characteristics of the cell strains can be correlated with the particular mutations found in the ERCC2 locus. Mutations of Arg658 to either His or Cys correlate with TTD cell strains with intermediate UV-sensitivity, mutation of Arg722 to Trp correlates with highly UV-sensitive TTD cell strains, and mutation of Arg683 to Trp correlates with XP-D. Alleles with mutation of Arg616 to Pro or with the combined mutation of Leu461 to Val and deletion of 716-730 are found in both XP-D and TTD cell strains.     

Analysis of Drosophila proboscipedia mutant alleles.

Proboscipedia (PB) is a HOX protein required for adult maxillary palp and proboscis formation. To identify domains of PB important for function, 21 pb point mutant alleles were sequenced. Twelve pb alleles had DNA sequence changes that encode an altered PB protein product. The DNA sequence changes of these 12 alleles fell into 2 categories: missense alleles that effect the PB homeodomain (HD), and nonsense or frameshift alleles that result in C-terminal truncations of the PB protein. The phenotypic analysis of the pb homeobox missense alleles suggests that the PB HD is required for maxillary palp and proboscis development and pb - Sex combs reduced (Scr) genetic interaction. The phenotypic analysis of the pb nonsense or frameshift alleles suggests that the C-terminus is an important region required for maxillary palp and proboscis development and pb-Scr genetic interaction. PB and SCR do not interact directly with one another in a co-immunoprecipitation assay and in a yeast two-hybrid analysis, which suggests the pb-Scr genetic interaction is not mediated by a direct interaction between PB and SCR.     

Molecular genetics of Drosophila alpha-actinin: mutant alleles disrupt Z disc integrity and muscle insertions

We have isolated a Drosophila melanogaster alpha-actinin gene and partially characterized several mutant alleles. The Drosophila protein sequence is very similar (68% identity) to those of chicken alpha-actinin isoforms, but less closely related (30% identity) to Dictyostelium alpha-actinin. The gene is within subdivision 2C of the X chromosome, coincident with 15 lethal (1)2Cb mutations. At least four alleles, l(1)2Cb1, l(1)2Cb2, l(1)2Cb4, and l(1)2Cb5 are interrupted by rearrangement breakpoints and must be null. In all four cases, hemizygous mutants complete embryogenesis and do not die until the second day of larval growth, signifying that either the role of alpha-actinin in nonmuscle cells is redundant or that a distinct and only distantly related gene encodes the non-muscle isoform. Allelic but less severely affected fliA mutants are apparently due to point mutations, and develop into adults having thoracic muscle abnormalities. EM of mutant muscles reveals that Z discs and myofibrillar attachments are disrupted, whereas epithelial "tendon" cells are less affected. We discuss these phenotypes in the light of presumed in vivo alpha-actinin functions.