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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.

© 2020 by the authors., Prostate cancer is the most frequent malignancy in European men and the second worldwide. One of the major oncogenic events in this disease includes amplification of the transcription factor cMYC. Amplification of this oncogene in chromosome 8q24 occurs concomitantly with the copy number increase in a subset of neighboring genes and regulatory elements, but their contribution to disease pathogenesis is poorly understood. Here we show that TRIB1 is among the most robustly upregulated coding genes within the 8q24 amplicon in prostate cancer. Moreover, we demonstrate that TRIB1 amplification and overexpression are frequent in this tumor type. Importantly, we find that, parallel to its amplification, TRIB1 transcription is controlled by cMYC. Mouse modeling and functional analysis revealed that aberrant TRIB1 expression is causal to prostate cancer pathogenesis. In sum, we provide unprecedented evidence for the regulation and function of TRIB1 in prostate cancer., We are grateful to the Carracedo lab for valuable input, to Ana M. Aransay and F. Elortza for technical advice. P.S. was funded by the European Training Networks Project (H2020-MSCA-ITN-308 2016 721532). V.T. is funded by Fundación Vasca de Innovación e Investigación Sanitarias, BIOEF (BIO15/CA/052), the AECC J.P. Bizkaia, the Basque Department of Health (2016111109) and the MICINN RTI2018-097267-B-I00. A. Macchia was funded by a FPI predoctoral fellowship from MICINN (PRE2018-083607). EB is funded by the MICINN (BFU2016-76872-R (FEDER/EU), PID2019-108112RB-I00, Severo Ochoa Excellence Accreditation SEV-2016-0644 and Excellence Networks SAF2017-90794-REDT). The work of A. Carracedo is supported by the Basque Department of Industry, Tourism and Trade (Elkartek), the department of education (IKERTALDE IT1106-16) and health (RIS3), the BBVA foundation and the MICINN (SAF2016-79381-R (FEDER/EU; PID2019-108787RB-I00) Severo Ochoa Excellence Accreditation SEV-2016-0644 and Excellence Networks SAF2016-81975-REDT and RED2018-102769-T), European Training Networks Project (H2020-MSCA-ITN-308 2016 721532), the AECC (IDEAS175CARR, GCTRA18006CARR), La Caixa Foundation (ID 100010434), under the agreement LCF/PR/HR17/ and the European Research Council (Starting Grant 336343, Consolidator Grant 819242). CIBERONC was co-funded with FEDER funds and funded by ISCIII.

Neutrophils are implicated in the pathogenesis of atherosclerosis but are seldom detected in atherosclerotic plaques. We investigated whether neutrophil-derived microvesicles may influence arterial pathophysiology. Here we report that levels of circulating neutrophil microvesicles are enhanced by exposure to a high fat diet, a known risk factor for atherosclerosis. Neutrophil microvesicles accumulate at disease-prone regions of arteries exposed to disturbed flow patterns, and promote vascular inflammation and atherosclerosis in a murine model. Using cultured endothelial cells exposed to disturbed flow, we demonstrate that neutrophil microvesicles promote inflammatory gene expression by delivering miR-155, enhancing NF-κB activation. Similarly, neutrophil microvesicles increase miR-155 and enhance NF-κB at disease-prone sites of disturbed flow in vivo. Enhancement of atherosclerotic plaque formation and increase in macrophage content by neutrophil microvesicles is dependent on miR-155. We conclude that neutrophils contribute to vascular inflammation and atherogenesis through delivery of microvesicles carrying miR-155 to disease-prone regions., The role of neutrophils in the development of atherosclerosis has long been an enigma, with few neutrophils detected within the plaque. Here, the authors show that microvesicles released from neutrophils increase vascular inflammation and enhance atherosclerotic plaque formation through delivery of miR-155.

Introduction MiR-101-3p has been implicated in the regulation of macrophage cholesterol efflux and pro-inflammatory genes by directly targeting ABCA1 and MKP-1, which are both downregulated in atherosclerosis. By using bioinformatic analysis we identified TRIB1 as a direct target of miR-101. The TRIB1 gene plays major roles in myeloid cells, regulating inflammation and lipid metabolism. Its deficiency is associated with a severe reduction of anti-inflammatory tissue macrophages (M2-like cells) and an increase in plasma triglycerides. Here we aimed to uncover the role of miR-101-3p/TRIB1 interaction and its consequences on human macrophage immunometabolism. Methods MiRanda target prediction algorithm was used to identify macrophage-specific miRNAs targeting TRIB1, along with additional web-based tools. MiR-101-3p was selected based on its conservation among species as well as its high predictive score and free energy. A luciferase reporter assay was used to validate the miR-101-3p/TRIB1 interaction, employing a miR-101 mimic and inhibitor. Gene expression (RT-qPCR) and protein analysis (Western Blot) was carried out in human monocyte-derived macrophages (hMDMs) following transient manipulation of TRIB1 and miR-101 levels. Results miR-101-3p has a functional binding site for the 3’UTR of TRIB1 and when overexpressed in hMDMs is able to reduce TRIB1 expression at both mRNA and protein levels. Over-expression of TRIB1 in hMDMs alters the levels of pro-inflammatory genes and scavenger receptors, inducing an M2-like phenotype. Conversely, overexpression of miR-101 is associated with a pro-inflammatory phenotype (M1-like). Conclusion miR-101-3p offers a potential target to simultaneously enhance the expression of ABCA1, MKP-1 and TRIB1, thus improving macrophage lipid metabolism and opposing inflammation, thereby attenuating atherosclerosis development. Conflict of interest No