Tryptophan-mediated interactions between tristetraprolin and the CNOT9 subunit are required for CCR4-NOT deadenylase complex recruitment

Journal article

The zinc-finger protein tristetraprolin (TTP) binds to AU-rich elements present in the 3' untranslated regions of transcripts that mainly encode proteins of the inflammatory response. TTP-bound mRNAs are targeted for destruction via recruitment of the eight-subunit deadenylase complex 'carbon catabolite repressor protein 4 (CCR4) -negative on TATA-less (NOT)' which catalyzes the removal of mRNA poly-(A) tails, the first obligatory step in mRNA decay. Here we show that a novel interaction between TTP and the CCR4-NOT subunit, CNOT9, is required for recruitment of the deadenylase complex. In addition to CNOT1, CNOT9 is now included in the identified CCR4-NOT subunits shown to interact with TTP. We find that both the N- and C-terminal domains of TTP are involved in an interaction with CNOT9. Through a combination of SPOT peptide array, site-directed mutagenesis and bio-layer interferometry, we identified several conserved tryptophan (Trp) residues in TTP that serve as major sites of interaction with two Trp-binding pockets of CNOT9, previously found to interact with another modulator GW182. We further demonstrate that these interactions are also required for recruitment of the CCR4-NOT complex and TTP-directed decay of an mRNA containing an AU-rich element in its 3'-untranslated region. Together the results reveal new molecular details for the TTP-CNOT interaction that shape an emerging mechanism whereby TTP targets inflammatory mRNAs for deadenylation and decay.

Authors: Bulbrook, D; Brazier, H; Mahajan, P; Kliszczak, M; Fedorov, O

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Elsevier
• Journal: Journal of Molecular Biology
• Volume: 430
• Issue: 5
• Pages: 722-736
• Publication date: 2017-12-29
• Acceptance date: 2017-12-20
• DOI: 10.1016/j.jmb.2017.12.018
• ISSN: 0022-2836
• Pmid: 29291391
• Language: English
• Keywords: AU-rich elements; mRNA; post-translational control; deadenylation; inflammatory