Pipsqueak (Psq) belongs to a family of proteins defined by a phylogenetically old protein-protein interaction motif. Like the GAGA factor and other members of this family, Psq is an important developmental regulator in Drosophila, having pleiotropic functions during oogenesis, embryonic pattern formation, and adult development. The GAGA factor controls the transcriptional activation of homeotic genes and other genes by binding to control elements containing the GAGAG consensus motif. Binding is associated with formation of an open chromatin structure that makes the control regions accessible to transcriptional activators. We show here that Psq contains a novel DNA-binding domain, which binds, like the GAGA factor zinc finger DNA-binding domain, to target sites containing the GAGAG consensus motif. Binding is suppressed, as in the GAGA factor and other proteins of the family, by the associated protein-protein interaction motif. The DNA-binding domain, which we call the Psq domain, is identical with a previously identified region consisting of four tandem repeats of a conserved 50-amino acid sequence, the Psq motif. The Psq domain seems to be structurally related to known DNA-binding domains, both in its repetitive character and in the putative three-␣-helix structure of the Psq motif, but it lacks the conserved sequence signatures of the classical eukaryotic DNA-binding motifs. Psq may thus represent the prototype of a new family of DNA-binding proteins. Members of the BTB1 /POZ protein family play important roles in development and reproduction of Drosophila melanogaster. These proteins contain a protein-protein interaction motif that was first identified in zinc finger proteins encoded by the Drosophila Broad-Complex and tramtrack genes (1, 2), and later also in the bric à brac gene product (3). The domain, which was thereupon designated as BTB (Broad-Complex, Tramtrack, Bric à brac) domain (3), has since been found in proteins of a variety of species, as diverse as slime molds (4) and humans (for a review, see Ref. 5). Many of these proteins are DNA-binding C 2 H 2 zinc finger proteins, but the presence of the domain in a family of pox virus proteins (6) soon indicated that coupling to a DNA-binding domain is not mandatory. The domain is therefore also referred to as the POZ (pox virus, zinc finger) domain (7). In BTB/POZ proteins that contain a zinc finger DNA-binding motif, DNA binding is strongly inhibited by the BTB/POZ domain. This inhibitory effect on DNA binding is also observed in chimeric proteins in which the BTB/POZ domain is associated with a heterologous DNA-binding domain, for instance a POU domain (7). Inhibition of DNA binding appears to be the result of oligomerization through proteinprotein interactions mediated by the BTB/POZ domain.The tendency of BTB/POZ proteins to oligomerize in solution and their localization in distinct nuclear substructures (7-10) suggests that they might act by modifying chromatin structure (5). In fact, such a mode of action is supported by different lines of eviden...

As a member of the trithorax-group, the Trithorax-like (Trl) gene of Drosophila melanogaster contributes to the expression of homeotic genes and many other genes. Trl encodes different isoforms of the GAGA factor which is thought to act as an "antirepressor" of transcription by remodelling chromatin structure and thereby rendering control regions accessible for transcriptional activators. A more global role of the GAGA factor in chromatin structure and function is suggested by various phenotypes of Trl mutations, such as modification of position effect variegation. To better define the molecular basis of these pleiotropic effects, we cloned cDNAs encoding the GAGA isoforms of D. melanogaster and a distantly related species, D. virilis. We also characterized the genomic organization of both the D. melanogaster and D. virilis genes, and analysed the expression patterns of isoform-specific mRNAs. The D. virilis GAGA isoforms show high similarity to their D. melanogaster counterparts, particularly within the BTB/POZ protein-interaction and the zinc finger DNA-binding domains. Interestingly, conservation clearly extends beyond the previously defined limits of these domains. Moreover, the comparison reveals a completely conserved block of amino acid residues located between the BTB/POZ and DNA-binding domains, and a high conservation of the C-terminus specific for one of the GAGA isoforms. Thus, sequences of as yet unknown functions are defined as rewarding targets for further mutational analyses. The high conservation of the GAGA proteins of the two species is in accord with the nearly identical genomic organization and expression patterns of the corresponding genes.