The N-Terminal Part of Drosophila CP190 Is a Platform for Interaction with Multiple Architectural Proteins.

CP190 is a co-factor in many Drosophila architectural proteins, being involved in the formation of active promoters and insulators. CP190 contains the N-terminal BTB/POZ (Broad-Complex, Tramtrack and Bric a brac/POxvirus and Zinc finger) domain and adjacent conserved regions involved in protein interactions. Here, we examined the functional roles of these domains of CP190 in vivo. The best-characterized architectural proteins with insulator functions, Pita, Su(Hw), and dCTCF, interacted predominantly with the BTB domain of CP190. Due to the difficulty of mutating the BTB domain, we obtained a transgenic line expressing a chimeric CP190 with the BTB domain of the human protein Kaiso. Another group of architectural proteins, M1BP, Opbp, and ZIPIC, interacted with one or both of the highly conserved regions in the N-terminal part of CP190. Transgenic lines of D. melanogaster expressing CP190 mutants with a deletion of each of these domains were obtained. The results showed that these mutant proteins only partially compensated for the functions of CP190, weakly binding to selective chromatin sites. Further analysis confirmed the essential role of these domains in recruitment to regulatory regions associated with architectural proteins. We also found that the N-terminal of CP190 was sufficient for recruiting Z4 and Chromator proteins and successfully achieving chromatin opening. Taken together, our results and the results of previous studies showed that the N-terminal region of CP190 is a platform for simultaneous interaction with various DNA-binding architectural proteins and transcription complexes.

The Hox proteins Ubx and AbdA collaborate with the transcription pausing factor M1BP to regulate gene transcription.

In metazoans, the pausing of RNA polymerase II at the promoter (paused Pol II) has emerged as a widespread and conserved mechanism in the regulation of gene transcription. While critical in recruiting Pol II to the promoter, the role transcription factors play in transitioning paused Pol II into productive Pol II is, however, little known. By studying how Drosophila Hox transcription factors control transcription, we uncovered a molecular mechanism that increases productive transcription. We found that the Hox proteins AbdA and Ubx target gene promoters previously bound by the transcription pausing factor M1BP, containing paused Pol II and enriched with promoter-proximal Polycomb Group (PcG) proteins, yet lacking the classical H3K27me3 PcG signature. We found that AbdA binding to M1BP-regulated genes results in reduction in PcG binding, the release of paused Pol II, increases in promoter H3K4me3 histone marks and increased gene transcription. Linking transcription factors, PcG proteins and paused Pol II states, these data identify a two-step mechanism of Hox-driven transcription, with M1BP binding leading to Pol II recruitment followed by AbdA targeting, which results in a change in the chromatin landscape and enhanced transcription.

Structural insights into highly similar spatial organization of zinc-finger associated domains with a very low sequence similarity.

ZAD is a C4 zinc-coordinating domain often found at the N-terminus mostly of arthropodan transcription factors with multiple C2H2 zinc-finger domains involved in the regulation of chromosome architecture and promotor activity. ZADs predominantly form homodimers and have low primary sequence similarity. We obtained three crystal structures of the most phylogenetically distant Drosophila ZADs and structure of the only known ZAD-like domain from a mammalian protein (ZNF276). All ZAD structures demonstrate unity of the spatial fold as well as some unique structural features. The specific homodimerization of ZAD is primarily determined by the position and size of secondary structural elements and is further strengthened by a number of unique interactions between subunits. Structural comparison allowed for unraveling key sequence features underlying the similarity of the spatial fold. These features result in a broad variety of ZADs in Arthropod C2H2 proteins, allowing for the emergence of a wide range of highly specific homodimers.