Structural and functional implications of the QUA2 domain on RNA recognition by GLD-1.
The STAR family comprises ribonucleic acid (RNA)-binding proteins that play key roles in RNA-regulatory processes. RNA recognition is achieved by a KH domain with an additional α-helix (QUA2) that seems to extend the RNA-binding surface to six nucleotides for SF1 (Homo sapiens) and seven nucleotides for GLD-1 (Caenorhabditis elegans). To understand the structural basis of this probable difference in specificity, we determined the solution structure of GLD-1 KH-QUA2 with the complete consensus sequence identified in the tra-2 gene. Compared to SF1, the GLD-1 KH-QUA2 interface adopts a different conformation resulting indeed in an additional sequence-specific binding pocket for a uracil at the 5'end. The functional relevance of this binding pocket is emphasized by our bioinformatics analysis showing that GLD-1 binding sites with this 5'end uracil are more predictive for the functional response of the messenger RNAs to gld-1 knockout. We further reveal the importance of the KH-QUA2 interface in vitro and that its alteration in vivo affects the level of translational repression dependent on the sequence of the GLD-1 binding motif. In conclusion, we demonstrate that the QUA2 domain distinguishes GLD-1 from other members of the STAR family and contributes more generally to the modulation of RNA-binding affinity and specificity of KH domain containing proteins.
Institute of Molecular Biology and Biophysics, Eidgenössische Technische Hochschule (ETH) Zürich, 8093 Zürich, Switzerland.
Gerrit M
GM
Daubner
Biozentrum, University of Basel, 4056 Basel, Switzerland.
Anneke
A
Brümmer
Friedrich Miescher Institute for Biomedical Research, 4002 Basel, Switzerland.
Cristina
C
Tocchini
Institute of Molecular Biology and Biophysics, Eidgenössische Technische Hochschule (ETH) Zürich, 8093 Zürich, Switzerland.
Stefan
S
Gerhardy
Friedrich Miescher Institute for Biomedical Research, 4002 Basel, Switzerland.
Rafal
R
Ciosk
Biozentrum, University of Basel, 4056 Basel, Switzerland.
Mihaela
M
Zavolan
Institute of Molecular Biology and Biophysics, Eidgenössische Technische Hochschule (ETH) Zürich, 8093 Zürich, Switzerland allain@mol.biol.ethz.ch.
Frédéric H-T
FH
Allain
D020413:3' Untranslated Regions; D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics; D008958:Models, Molecular; D009154:Mutation; D010641:Phenotype; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid; D014498:Uracil / Q000737:chemistry
D020413:3' Untranslated Regions; D029742:Caenorhabditis elegans Proteins; C094448:GLD-1 protein, C elegans; D008565:Membrane Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C076107:tra-2 protein, C elegans; D014498:Uracil