The large subunit of the human pre-messenger RNA splicing factor U2 small nuclear ribonucleoprotein auxiliary factor (hU2AF65) is required for spliceosome assembly in vitro. A complementary DNA clone encoding the large subunit of Drosophila U2AF (dU2AF50) has been isolated. The dU2AF50 protein is closely related to its mammalian counterpart and contains three carboxyl-terminal ribonucleoprotein consensus sequence RNA binding domains and an amino-terminal arginine- and serine-rich (R/S) domain. Recombinant dU2AF50 protein complements mammalian splicing extracts depleted of U2AF activity. Germline transformation of Drosophila with the dU2AF50 complementary DNA rescues a lethal mutation, establishing that the dU2AF50 gene is essential for viability. R/S domains have been found in numerous metazoan splicing factors, but their function is unknown. The mutation in Drosophila U2AF will allow in vivo analysis of a conserved R/S domain-containing general splicing factor.

P cytotype is a regulatory state, characteristic of Drosophila P-strain females, in which P-element transposition is repressed. P cytotype is established maternally in the germ line but is also dependent on the presence of P elements in the zygote. One aspect of P cytotype involves transcriptional repression of the P-element promoter. Here, we show that transcriptional repression by P cytotype in the female germ line occurs by a general promoter-independent mechanism with heterologous promoters carried in P-element vectors. P-cytotype transcriptional repression results in low levels of pre-mRNA and a reduction in splicing of the P-element third intron (IVS3)-containing mRNA, thus causing an increase in the proportion of 66-kD repressor mRNA. Increased retention of IVS3 in P cytotype would result in an autoregulatory loop of 66-kD repressor production. This combination of germ-line transcriptional repression and splicing control provides a mechanism to maintain repression during the maternal inheritance of P cytotype. These findings suggest that transcriptional repression may play an additional role in the regulation of gene expression, namely allowing alteration of pre-mRNA splicing patterns.[Key Words: Tissue specificity; germ-line transcriptional control; pre-mRNA splicing regulation; germ-lineReceived January 25, 1995; revised version accepted April 13, 1995. P element transposition in Drosophila is controlled in two ways: (1) transposition is restricted to the germ line by alternative RNA splicing (tissue specificity), and (2) transposition only occurs when a P-strain male (carrying P elements} is mated to an M-strain female (lacking P elements) but does not occur in the reciprocal cross. P-strain females are said to possess "P cytotype," the regulatory state by which transposition is repressed. P cytotype is initially inherited maternally and in a manner similar to cytoplasmic inheritance but is ultimately determined zygotically by the presence of chromosomal P elements (for review, see Engels 1983Engels , 1989Rio 1991). At least part of the repressive activity of P cytotype is caused by repressor proteins encoded by the P elements, and this repressive activity can be influenced by genomic position (Robertson and Engels 1989;Misra and Rio 1990;Misra et al. 1993}. Genetic studies of cytotype suggested that repressor synthesis would be autoregulatory in the germ line (Engels 1983; O'Hare and Rubin 1983), but the underlying mechanism has remained elusive.Cytotype repression of P-element mobility has both maternal and zygotic components (Engels 1983(Engels , 1989. Genetic assays for cytotype have allowed the detection of two types of regulatory P elements. The complete 2.9-kb P elements encode the 66-kD repressor protein (Misra and Rio 1990; Gloor et al. 1993}, whereas the smaller internally deleted elements encode truncated repressor proteins, such as the KP protein (Black et al. 1987;Rasmusson et al. 1993). Complete P elements are capable of exhibiting the repressive maternal effect of P cytotype, depending o...

In Drosophila, P-element transposition causes mutagenesis and genome instability during hybrid dysgenesis. The P-element 31-bp terminal inverted repeats (TIRs) contain sequences essential for transposase cleavage and have been implicated in DNA repair via protein-DNA interactions with cellular proteins. The identity and function of these cellular proteins were unknown. Biochemical characterization of proteins that bind the TIRs identified a heterodimeric basic leucine zipper (bZIP) complex between an uncharacterized protein that we termed "Inverted Repeat Binding Protein (IRBP) 18" and its partner Xrp1. The reconstituted IRBP18/Xrp1 heterodimer binds sequence-specifically to its dsDNA-binding site within the P-element TIRs. Genetic analyses implicate both proteins as critical for repair of DNA breaks following transposase cleavage in vivo. These results identify a cellular protein complex that binds an active mobile element and plays a more general role in maintaining genome stability.T ransposable elements contribute significantly to the organization and evolution of all eukaryotic genomes. Recent estimates of transposon content within the Drosophila melanogaster genome are between 5% and 10%, and in humans over half the genome is composed of mobile elements (1, 2). Although many of these elements, including the Drosophila P-element transposon, are still active (3), the cellular mechanisms used to combat the genotoxic effects of DNA double-strand breaks (DSBs) generated by transpositional recombination are not fully understood. The Drosophila P-transposable element provides an excellent model for understanding the ancient mechanisms used by the cell to counteract newly invading parasitic mobile DNA elements (4).The P-element transposon is a mobile DNA element that spread through wild populations of D. melangaster ∼100 y ago after most common laboratory strains were isolated (5, 6). P elements were identified by studying a genetic syndrome called "P-M hybrid dysgenesis." It was observed that males from wild populations (P strains) crossed to females from isolated laboratory stocks (M strains) yielded progeny that had germline mutations, temperature-sensitive sterility, and atypical male recombination (6). Reciprocal crosses yielded phenotypically normal progeny. The P element was shown to be the causative agent of these so-called P-M hybrid dysgenesis phenotypes by molecular analyses showing that P elements were present in variable locations in P strains yet totally absent from most M strains (7,8).The Drosophila P-element transposon encodes a GTP-dependent site-specific DNA transposase/integrase family enzyme (9, 10). At each end of the P-element transposon are perfect 31-bp terminal inverted repeats (TIRs), 11-bp internal inverted repeats that serve as enhancers of transposition, and internal 10-bp transposase binding sites (11-13) (Fig. 1A). The P-element transposase catalyzes DNA cleavage within the 31-bp TIRs to create 17-nt 3′ single-strand extensions at both the donor site and the transposon ends (14, 15)...

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A genome-wide scan in 60 bipolar affective disorder (BPAD) affected sib-pairs (ASPs) identified linkage on chromosome 21 at 21q22 (D21S1446, NPL = 1.42, P = 0.08), a BPAD susceptibility locus supported by multiple studies. Although this linkage only approaches significance, the peak marker is located 12 Kb upstream of S100B, a neurotrophic factor implicated in the pathology of psychiatric disorders, including BPAD and schizophrenia. We hypothesized that the linkage signal at 21q22 may result from pathogenic disease variants within S100B and performed an association analysis of this gene in a collection of 125 BPAD type I trios. S100B single nucleotide polymorphisms (SNPs) rs2839350 (P = 0.022) and rs3788266 (P = 0.031) were significantly associated with BPAD. Since variants within S100B have also been associated with schizophrenia susceptibility, we reanalyzed the data in trios with a history of psychosis, a phenotype in common between the two disorders. SNPs rs2339350 (P = 0.016) and rs3788266 (P = 0.009) were more significantly associated in the psychotic subset. Increased significance was also obtained at the haplotype level. Interestingly, SNP rs3788266 is located within a consensus-binding site for Six-family transcription factors suggesting that this variant may directly affect S100B gene expression. Fine-mapping analyses of 21q22 have previously identified transient receptor potential gene melastatin 2 (TRPM2), which is 2 Mb upstream of S100B, as a possible BPAD susceptibility gene at 21q22. We also performed a family-based association analysis of TRPM2 which did not reveal any evidence for association of this gene with BPAD. Overall, our findings suggest that variants within the S100B gene predispose to a psychotic subtype of BPAD, possibly via alteration of gene expression.