Structure and dynamics of the quaternary hunchback mRNA translation repression complex

Author:

Macošek Jakub1ORCID,Simon Bernd1,Linse Johanna-Barbara2,Jagtap Pravin Kumar Ankush1ORCID,Winter Sophie L1ORCID,Foot Jaelle1,Lapouge Karine3,Perez Kathryn3,Rettel Mandy4,Ivanović Miloš T2,Masiewicz Pawel1,Murciano Brice1,Savitski Mikhail M4,Loedige Inga1,Hub Jochen S2,Gabel Frank5,Hennig Janosch16ORCID

Affiliation:

1. Structural and Computational Biology Unit, European Molecular Biology Laboratory Heidelberg, Heidelberg 69117, Germany

2. Theoretical Physics and Center for Biophysics, Saarland University, Saarbrücken 66123, Germany

3. Protein Expression and Purification Core Facility, European Molecular Biology Laboratory Heidelberg, Heidelberg 69117, Germany

4. Proteomics Core Facility, European Molecular Biology Laboratory Heidelberg, Heidelberg 69117, Germany

5. Institut Biologie Structurale, University Grenoble Alpes, CEA, CNRS, Grenoble 38044, France

6. Chair of Biochemistry IV, Biophysical Chemistry, University of Bayreuth, Universitätsstrasse 30, 95447 Bayreuth, Germany

Abstract

Abstract A key regulatory process during Drosophila development is the localized suppression of the hunchback mRNA translation at the posterior, which gives rise to a hunchback gradient governing the formation of the anterior-posterior body axis. This suppression is achieved by a concerted action of Brain Tumour (Brat), Pumilio (Pum) and Nanos. Each protein is necessary for proper Drosophila development. The RNA contacts have been elucidated for the proteins individually in several atomic-resolution structures. However, the interplay of all three proteins during RNA suppression remains a long-standing open question. Here, we characterize the quaternary complex of the RNA-binding domains of Brat, Pum and Nanos with hunchback mRNA by combining NMR spectroscopy, SANS/SAXS, XL/MS with MD simulations and ITC assays. The quaternary hunchback mRNA suppression complex comprising the RNA binding domains is flexible with unoccupied nucleotides functioning as a flexible linker between the Brat and Pum-Nanos moieties of the complex. Moreover, the presence of the Pum-HD/Nanos-ZnF complex has no effect on the equilibrium RNA binding affinity of the Brat RNA binding domain. This is in accordance with previous studies, which showed that Brat can suppress mRNA independently and is distributed uniformly throughout the embryo.

Funder

Deutsche Forschungsgemeinschaft

Publisher

Oxford University Press (OUP)

Subject

Genetics

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