Abstract
We previously reported that IL-6R, STAT3 and miR-34a form a positive feedback-loop, which promotes epithelial to mesenchymal transition (EMT), invasion, and metastasis of colorectal cancer (CRC) [1]. In that study only the membrane-bound form of the IL-6R was shown to be repressed by miR-34a. Here, we show that also the mRNA encoding the soluble IL6R (s-IL-6R) is directly targeted and repressed by miR-34a. Accordingly, the concentration of s-IL6R protein was decreased in conditioned media of CRC cell lines ectopically expressing miR-34a. The s-IL-6R mediates IL-6 trans-signaling, which also affects cells that do not express the IL-6R. Since IL-6 trans-signaling is involved in numerous inflammatory disease states these findings may be relevant for future therapeutic approaches.
Highlights
Signaling mediated via the interleukin-6 (IL-6)/ interleukin-6 receptor (IL-6R) plays a pivotal role during immune responses and in cancer [2,3,4]
To investigate whether the endogenous transcript isoform that encodes s-IL-6R is regulated by miR-34a, we designed qPCR primers that exclusively recognize the membrane-bound IL-6R (m-IL-6R) or the s-IL6R mRNA isoform (Figure 3A)
We determined the effect of ectopic miR-34a expression on s-IL-6R protein expression by transfecting SW480 and SW620 cells with pre-miR-34a oligonucleotides. s-IL6R specific ELISA analyses showed that in both cell lines ectopic expression of miR-34a significantly decreased the concentration of s-IL-6R in conditioned media (Figure 3C, 3D)
Summary
Signaling mediated via the interleukin-6 (IL-6)/ interleukin-6 receptor (IL-6R) plays a pivotal role during immune responses and in cancer [2,3,4]. Besides classical IL-6 signaling, which involves the membrane-bound IL-6R (m-IL-6R), IL-6 trans-signaling has been described [6]. The later involves the s-IL-6R, which is shedded/ released by cells and in complex with IL-6 binds to and activates gp130. Thereby, IL-6 trans-signaling can affect cells that do not express the IL-6R [7]. The human s-IL6R protein is either generated by skipping exon 9 via alternative splicing and subsequent earlier termination of translation caused by a stop codon in the alternative reading frame in exon 10 (Figure 1A) or by cleavage of the m-IL-6R protein by the α-secretase ADAM17 upstream of the transmembrane (TM) domain (Figure 1B) [8]. The N-terminal IL-6R fragment is shedded, whereas the C-terminal fragment is cleaved by γ-secretase in the transmembrane (TM) domain and removed by lysosomal degradation
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