Abstract
The pseudokinase TRIB1 controls cell function in a range of contexts, by regulating MAP kinase activation and mediating protein degradation via the COP1 ubiquitin ligase. TRIB1 regulates polarization of macrophages and dysregulated Trib1 expression in murine models has been shown to alter atherosclerosis burden and adipose homeostasis. Recently, TRIB1 has also been implicated in the pathogenesis of prostate cancer, where it is often overexpressed, even in the absence of genetic amplification. Well described TRIB1 effectors include MAP kinases and C/EBP transcription factors, both in immune cells and in carcinogenesis. However, the mechanisms that regulate TRIB1 itself remain elusive. Here, we show that the long and conserved 3’untranslated region (3’UTR) of TRIB1 is targeted by miRNAs in macrophage and prostate cancer models. By using a systematic in silico analysis, we identified multiple “high confidence” miRNAs potentially binding to the 3’UTR of TRIB1 and report that miR-101-3p and miR-132-3p are direct regulators of TRIB1 expression and function. Binding of miR-101-3p and miR-132-3p to the 3’UTR of TRIB1 mRNA leads to an increased transcription and secretion of interleukin-8. Our data demonstrate that modulation of TRIB1 by miRNAs alters the inflammatory profile of both human macrophages and prostate cancer cells.
Highlights
Tribbles-1 (TRIB1) is an evolutionary conserved pseudokinase that in mammals controls a wide range of interacting signaling pathways, such as mitogen activated protein kinases and the PI3kinase pathway
Dual immunofluorescence staining of Kupffer cells (Figures 1A, B) and adipose-tissue macrophages (ATMs) (Supplementary Figure 2), revealed that while altered Trib1 expression has no effect on the number of F4/80 positive macrophages in these tissues (Figure 1C and Supplementary Figure 2B), it significantly altered their phenotype (Figures 1D, E and Supplementary Figure 2C)
As the human TRIB1 mRNA is characterized by a 2 kb long and conserved 3’untranslated region (3’UTR), we proposed that regulatory sequences within this may contribute to the control of TRIB1 mRNA stability, including a role for miRNAs
Summary
Tribbles-1 (TRIB1) is an evolutionary conserved pseudokinase that in mammals controls a wide range of interacting signaling pathways, such as mitogen activated protein kinases and the PI3kinase pathway. The TRIB1 protein contains an N-terminal PEST region, a pseudokinase domain and a C-terminal COP1-binding region. TRIB1 has been reported to be overexpressed in prostate cancer, where it controls the expression of endoplasmic reticulum chaperone proteins and induces M2-macrophage differentiation [10, 11]. TRIB1 is highly unstable, reported to have an mRNA half-life shorter than 1 hour [12]. TRIB1 expression is highly variable among different cell types and tissues, suggesting it might be subject to cell type-dependent posttranscriptional regulation [13, 14]. The 2kb long, 3’untraslated region (3’UTR) of TRIB1 mRNA represents more than 50% of the entire sequence and is highly conserved among different animal species. A major gene regulatory mechanism described in eukaryotes is mediated by microRNAs (miRNAs) by the process of RNA interference (RNAi). miRNAs are small non-coding RNAs that commonly bind to the 3’UTR of the target gene, and, via association with effector proteins, form the RNA-induced silencing complex (RISC) that leads to RNA degradation and inhibition of protein translation [16]
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