Novel AU-rich proximal UTR sequences (APS) enhance CXCL8 synthesis upon the induction of rpS6 phosphorylation

Abstract

The role of ribosomal protein S6 (rpS6) phosphorylation in mRNA translation remains poorly understood. Here, we reveal a potential role in modulating the translation rate of chemokine (C-X-C motif) ligand 8 ( CXCL8 or Interleukin 8, IL8 ). We observed that more CXCL8 protein was being secreted from less CXCL8 mRNA in primary macrophages and macrophage-like HL-60 cells relative to other cell types. This correlated with an increase in CXCL8 polyribosome association, suggesting an increase in the rate of CXCL8 translation in macrophages. The cell type-specific expression levels were replicated by a CXCL8- UTR-reporter (Nanoluc reporter flanked by the 5’ and 3’ UTR of CXCL8 ). Mutations of the CXCL8 -UTR-reporter revealed that cell type-specific expression required: 1) a 3’ UTR of at least three hundred bases; and 2) an AU base content that exceeds fifty percent in the first hundred bases of the 3’ UTR immediately after the stop codon, which we dub AU-rich proximal UTR sequences (APS). The 5’ UTR of CXCL8 enhanced expression at the protein level and conferred cell type-specific expression when paired with a 3’ UTR. A search for other APS-positive mRNAs uncovered TNF alpha induced protein 6 ( TNFAIP6 ), another mRNA that was translationally upregulated in macrophages. The elevated translation of APS-positive mRNAs in macrophages coincided with elevated rpS6 S235/236 phosphorylation. Both were attenuated by the ERK1/2 signaling inhibitors, U0126 and AZD6244. In A549 cells, rpS6 S235/236 phosphorylation was induced by TAK1, Akt or PKA signaling. This enhanced the translation of the CXCL8 -UTR-reporters. Thus, we propose that the induction of rpS6 S235/236 phosphorylation enhances the translation of mRNAs that contain APS motifs, such as CXCL8 and TNFAIP6 . This may contribute to the role of macrophages as the primary producer of CXCL8, a cytokine that is essential for immune cell recruitment and activation.

Bridge Integrator 2, a New Specific Target for Improving Reproductive Life Span through RPS6 and NNT

“…10, shadowed in the rectangle), suggesting that the conserved activation/inactivation mechanism might rely on these two residues. On the other side, human Bin2 is also (43)(44)(45). In the present study, we showed that Bin2 might be a novel kinase that directly phosphorylates RPS6 (Fig.…”

Inhibiting bridge integrator 2 phosphorylation leads to improved oocyte quality, ovarian health and fertility in aging and after chemotherapy in mice

“…Conceivably, it can regulate various important cellular processes. Actually, RPS6 is known to be regulated by multiple kinases and phosphatases (43)(44)(45). In the present study, we showed that Bin2 might be a novel kinase that directly phosphorylates RPS6 (Fig.…”

Inflammation suppresses DLG2 expression decreasing inflammasome formation

“…Furthermore, a decrease in the phosphorylation of S6, consistent with control of proliferation was detected after DLG2 overexpression. Decreased S6 phosphorylation has been shown to inhibit the synthesis of the chemokine IL-8 (Ang et al 2019 ). Finally, DLG2 loss has been shown to increase cyclin A2 and result in S phase progression and DNA replication (Keane et al 2020 ), a similar function to the integral inflammasome component GSDMD (Wang et al 2018 ).…”