Die Rolle der roX RNA in Struktur und Funktion des Dosis Kompensationskomplexes.
Biochemie
Zusammenfassung der Projektergebnisse
Complex eukaryotic genomes encode abundant long, non-coding (lnc) RNAs. Our knowledge of the molecular mechanisms underlying chromatin-based lncRNA function is scarce. We will study a paradigmatic, multi-enzyme chromatin regulator, which absolutely requires lncRNA for assembly and function. The dosage compensation complex (DCC) of Drosophila melanogaster consists of five male-specific-lethal (MSL) proteins that assemble with non-coding roX RNA to form a ribonucleoprotein regulatory complex with special properties: the DCC doubles the transcription of most genes on the single X chromosome in male cells to match the combined output of the two Xs in females. Although the MSL protein alone can associate with each other and are active enzymes in vitro, the association of roX is a vital requirement for DCC function in male flies. RoX function, however, is mysterious. Secondary structure remodeling of roX by the DExH helicase MLE initiates association of MSL proteins towards assembly of a functional DCC. We hypothesize that the MSL proteins acquire their dedicated functionality within the DCC by association with roX. We aim to elucidate the molecular principles of lncRNA roX integration into the DCC to understand the roX dependency of DCC assembly and activity. To this end, we will study the mechanisms of roX incorporation in detail, optimise the assembly of MSL proteins with specific RNA towards a functional DCC and characterize the roX-MSL interactions through specific protein-RNA interaction assays (involving crosslinking, sequencing and mass spectrometry). Our final goal is to compare MSL assemblies in the presence and absence of specific RNA and to correlate structural changes with refined MSL functions. Our study will shed light on a fascinating, highly evolved chromatin regulator. More importantly, we strive to uncover regulatory principles of lncRNA function of general relevance and broad applicability for the emerging field of lncRNA biology.
Projektbezogene Publikationen (Auswahl)
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Structure, dynamics and roX2-lncRNA binding of tandem double-stranded RNA binding domains dsRBD1,2 of Drosophila helicase Maleless. Nucleic Acids Research, 47(8), 4319-4333.
Ankush, Jagtap Pravin Kumar; Müller, Marisa; Masiewicz, Pawel; von Bülow, Sören; Hollmann, Nele Merret; Chen, Po-Chia; Simon, Bernd; Thomae, Andreas W.; Becker, Peter B. & Hennig, Janosch
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Two-step mechanism for selective incorporation of lncRNA into a chromatin modifier. Nucleic Acids Research.
Müller, Marisa; Schauer, Tamas; Krause, Silke; Villa, Raffaella; Thomae, Andreas W. & Becker, Peter B.
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Identification of Intrinsic RNA Binding Specificity of Purified Proteins by in vitro RNA Immunoprecipitation (vitRIP). BIO-PROTOCOL, 11(5).
Müller, Marisa; Schauer, Tamas & Becker, Peter
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Structural basis of RNA-induced autoregulation of the DExH-type RNA helicase maleless. Molecular Cell, 83(23), 4318-4333.e10.
Jagtap, Pravin Kumar Ankush; Müller, Marisa; Kiss, Anna E.; Thomae, Andreas W.; Lapouge, Karine; Beck, Martin; Becker, Peter B. & Hennig, Janosch
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Processivity and specificity of histone acetylation by the male-specific lethal complex. Nucleic Acids Research, 52(9), 4889-4905.
Kiss, Anna E.; Venkatasubramani, Anuroop V.; Pathirana, Dilan; Krause, Silke; Sparr, Aline Campos; Hasenauer, Jan; Imhof, Axel; Müller, Marisa & Becker, Peter B.
