Structural basis for the assembly of the Sxl–Unr translation regulatory complex

Abstract

Genetic equality between males and females is established by chromosome-wide dosage-compensation mechanisms. In the fruitfly Drosophila melanogaster, the dosage-compensation complex promotes twofold hypertranscription of the single male X-chromosome and is silenced in females by inhibition of the translation of msl2, which codes for the limiting component of the dosage-compensation complex. The female-specific protein Sex-lethal (Sxl) recruits Upstream-of-N-ras (Unr) to the 3' untranslated region of msl2 messenger RNA, preventing the engagement of the small ribosomal subunit. Here we report the 2.8 Å crystal structure, NMR and small-angle X-ray and neutron scattering data of the ternary Sxl-Unr-msl2 ribonucleoprotein complex featuring unprecedented intertwined interactions of two Sxl RNA recognition motifs, a Unr cold-shock domain and RNA. Cooperative complex formation is associated with a 1,000-fold increase of RNA binding affinity for the Unr cold-shock domain and involves novel ternary interactions, as well as non-canonical RNA contacts by the α1 helix of Sxl RNA recognition motif 1. Our results suggest that repression of dosage compensation, necessary for female viability, is triggered by specific, cooperative molecular interactions that lock a ribonucleoprotein switch to regulate translation. The structure serves as a paradigm for how a combination of general and widespread RNA binding domains expands the code for specific single-stranded RNA recognition in the regulation of gene expression.

One, Two, Three, Four! How Multiple RRMs Read the Genome Sequence

“…It also reveals intermolecular cooperativity between the Sxl RRMs and the UNR CSD1 in RNA binding (Hennig et al, 2014). In this ternary complex, the Sxl RRMs bind to the 5 0 -half of the 18-mer RNA, while CSD1 interacts with the 3 0 -end of the RNA (Fig.…”

“…Recognition of neighboring regulatory sites by trans-acting factors in a coordinated fashion, nicely illustrated by the recent ternary complex structures of Sxl RRM-UNR CSD1-RNA (Hennig et al, 2014) and ASD-1-SUP-12-RNA (Kuwasako et al, 2014), is in fact a recurrent phenomenon in eukaryotic posttranscriptional control. It is therefore clear that the interpretation of genomic data requires a certain level of precaution and it may fail to identify the exact binding position of the precipitated protein.…”

“…(E) Structure of ternary complex of ASD-1 and SUP-12 RRMs in complex with RNA (Kuwasako et al, 2014). (F) Crystal structure of ternary complex of Sxl tandem RRMs (in gray), UNR CSD1 (in cyan) and RNA (shown as yellow sticks) (Hennig et al, 2014).…”

“…6F). The RNA is sandwiched between the Sxl tandem RRMs and UNR CSD by a set of unique, intertwined protein-RNA-protein interactions resembling a "triple zipper," which mediates cooperative RNA recognition by Sxl RRM1 and UNR CSD1 (Hennig et al, 2014).…”

Specific RNP capture with antisense LNA/DNA mixmers

“…The interaction of Sxl with poly(U) motifs has been well-characterized biochemically and structurally (Inoue et al 1990;Valcárcel et al 1993;Handa et al 1999;Hennig et al 2014). In addition to its interaction with msl-2 mRNA (Bashaw and Baker 1995;Kelley et al 1995Kelley et al , 1997Bashaw and Baker 1997), Sxl also binds to poly(U) motifs present in its own mRNA (Cline 1984;Bell et al 1991) and in the transformer mRNA (tra) (Boggs et al, 1987).…”