Identification of new high affinity targets for Roquin based on structural conservation

Abstract

Post-transcriptional gene regulation controls the amount of protein produced from a specific mRNA by altering both its decay and translation rates. Such regulation is primarily achieved by the interaction of trans-acting factors with cis-regulatory elements in the untranslated regions (UTRs) of mRNAs. These interactions are guided either by sequence- or structure-based recognition. Similar to sequence conservation, the evolutionary conservation of a UTR’s structure thus reflects its functional importance. We used such structural conservation to identify previously unknown cis-regulatory elements. Using the RNA folding program Dynalign, we scanned all UTRs of humans and mice for conserved structures. Characterizing a subset of putative conserved structures revealed a binding site of the RNA-binding protein Roquin. Detailed functional characterization in vivo enabled us to redefine the binding preferences of Roquin and identify new target genes. Many of these new targets are unrelated to the established role of Roquin in inflammation and immune responses and thus highlight additional, unstudied cellular functions of this important repressor. Moreover, the expression of several Roquin targets is highly cell-type-specific. In consequence, these targets are difficult to detect using methods dependent on mRNA abundance, yet easily detectable with our unbiased strategy.

Impact of RNA–Protein Interaction Modes on Translation Control: The Versatile Multidomain Protein Gemin5

“…An example of a protein with various RBDs recognizing stem‐loop structures on the 3′‐UTR of mRNAs is the ring finger and CCCH‐type domains 1, also known as Roquin‐1 (Figure a), a protein mediating T helper‐17 cell differentiation . The observation that Roquin‐1 interacts with RNA structural elements on the 3′‐UTR of proinflammatory cytokines like tumor necrosis factor (TNF) and interleukin‐6 (IL‐6), allowed the identification of novel protein partners by searching RNAs that fold into similar stem‐loops . Subsequently, characterization of RNA reporters carrying these sequences revealed that RNA stability is governed by Roquin‐1.…”

Structural basis for the recognition of transiently structured AU-rich elements by Roquin

“…The target ARE conformers are mutually exclusive and can lead to redundant or competitive regulation of the ARE-containing mRNA through the respective cognate RBP. ( B ) The previously described regulatory ARE region in the UCP3 mRNA 3′-UTR shown as linear sequence and as its structured conformer with the two CDEs bound by Roquin ( 44 ). Below that, the two CDEs are shown as isolated wild type species and as stabilized SLs with the underlying numbering of bases as used in this study.…”

“…The strictness of the CDE sequences in functional targets for Roquin has been a matter of debate for years now, including contradictive findings ( 36 , 38 , 43 , 44 ). Certainly, a major requirement for a functional SL represents its accessibility in vivo , i.e.…”

Roquin1 inhibits the proliferation of breast cancer cells by inducing G1/S cell cycle arrest via selectively destabilizing the mRNAs of cell cycle–promoting genes

“…Roquin1 deficiency induced the death of mouse embryos and severe autoimmune reactions and enteritis [ 13 , 14 ]. Evidence has shown that Roquin1 can induce mRNA decay by binding the stem–loop structure in the 3’UTR of target genes [ 15 – 17 ]. In addition, Roquin1 serves as a regulator of multiple signaling pathways, such as AMP-activated protein kinase (AMPK) [ 18 ], NF-κB [ 19 ], and PI3K-mTOR [ 20 ], to regulate immune responses.…”