Abstract
In the absence of the scanning ribosomes that unwind mRNA coding sequences and 5′UTRs, mRNAs are likely to form secondary structures and intermolecular bridges. Intermolecular base pairing of non polysomal mRNAs is involved in stress granule (SG) assembly when the pool of mRNAs freed from ribosomes increases during cellular stress. Here, we unravel the structural mechanisms by which a major partner of dormant mRNAs, YB-1 (YBX1), unwinds mRNA secondary structures without ATP consumption by using its conserved cold-shock domain to destabilize RNA stem/loops and its unstructured C-terminal domain to secure RNA unwinding. At endogenous levels, YB-1 facilitates SG disassembly during arsenite stress recovery. In addition, overexpression of wild-type YB-1 and to a lesser extent unwinding-defective mutants inhibit SG assembly in HeLa cells. Through its mRNA-unwinding activity, YB-1 may thus inhibit SG assembly in cancer cells and package dormant mRNA in an unfolded state, thus preparing mRNAs for translation initiation.
Highlights
In proliferating cells such as cancer cells, mRNAs are mostly associated with scanning ribosomes to synthesize proteins, and only a small fraction of mRNAs remain dormant [1]
We considered the putative role of YB-1, an abundant mRNA-binding protein, in stress granule (SG) assembly
YB-1 possesses three distinct domains [10,14]: (i) an unstructured alanine/proline-rich domain which may interact with protein partners, (ii) a highly conserved -barrel structure, i.e., the cold-shock domain (CSD) that binds to RNA and DNA and (iii) a long unstructured C-terminal domain (CTD) that harbors well separated clusters of negatively and positively charged residues that may be involved in selfadhesive interactions [49]
Summary
In proliferating cells such as cancer cells, mRNAs are mostly associated with scanning ribosomes to synthesize proteins, and only a small fraction of mRNAs remain dormant [1]. As most mRNAs rely on cap-dependent translation in mammalian cells, polysome dissociation after stress induction leads to an increase in non polysomal mRNA in the cytoplasm (stalled translation initiation complexes, mRNPs without ribosomes). The association of non polysomal mRNA with self-adhesive RNA-binding proteins such as G3BP-1 subsequently promotes the assembly of liquid-like mRNArich condensates in the cytoplasm called stress granules (SGs) [3,4,5,6]. In addition to the important contribution of self-adhesive RNA-binding proteins (RBPs), intermolecular RNA–RNA interactions contribute to SG assembly [7]. The confinement of mRNAs in SGs increases the occurrence of intermolecular base pairing between mRNAs. intramolecular base-pairing may direct self-adhesive RBPs in SGs owing to their high affinity for mRNA secondary structures [8]. In agreement with the critical role of RNA:RNA interactions in SG assembly, the overexpression of the RNA helicases, eIF4A and DDX19A, was shown to negatively regulate SG assembly [9]
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