Biological role of miR-204 and miR-211 in melanoma
1 Oncogenomics Unit, CRL-ISPRO, 56124 Pisa, Italy
2 Institute of Clinical Physiology, CNR, 56124 Pisa, Italy
3 Institute of Informatics and Telematics, CNR, 56124 Pisa, Italy
Keywords: melanoma, small RNA-seq, BRAF inhibitors, miR-204, miR-211
Received: March 28, 2018
Accepted: May 18, 2018
Published: August 22, 2018
Copyright: Vitiello et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License 3.0 (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
In this short report, we pinpoint some technical and conceptual flaws that we found in the article entitled “miR-204-5p and miR-211-5p contribute to BRAF inhibitor resistance in melanoma” (Díaz-Martínez et al., Cancer Research 2018). We also discuss how, in our opinion, these flaws led Díaz-Martínez and colleagues to incorrect conclusions about the biological role that miR-204 and miR-211 play in melanoma and about the terms of their involvement in the phenomenon of resistance to BRAF inhibitors.
With the aim to identify the microRNAs involved in resistance to vemurafenib, in the research article entitled “miR-204-5p and miR-211-5p contribute to BRAF inhibitor resistance in melanoma” Díaz-Martínez and colleagues performed small RNA sequencing on A375 parental cells and the resistant A375-VR population, looking for differentially expressed microRNAs .
miR-204 was chosen because its levels are ~2-fold higher in A375-VR vs A375, as detected by small RNA-seq and confirmed by qRT-PCR. Consistently with our previously published data (Vitiello et al., Context-dependent miR-204 and miR-211 affect the biological properties of amelanotic and melanotic melanoma cells, Oncotarget ), Díaz-Martínez and colleagues show that in A375 cells (but not in A375-VR cells) the ERK pathway negatively regulates miR-204. They also claim that miR-204 is positively involved in resistance to vemurafenib. However, this claim is formally supported only by the mild decrease in proliferation that A375-VR show when transfected with a miR-204 inhibitor and exposed to vemurafenib (Figure 5E in reference 1) [3, 4]. Conversely, we and others have extensively demonstrated both in vitro and using patient data that miR-204 rather exerts its activity in sensitive cells, where its induction upon vemurafenib treatment is very robust, it targets AP1S2 (a validated pro-motility target not considered by Díaz-Martínez and colleagues) and it potentiates the anti-motility effects of the drug, in turn behaving as an oncosuppressor [2, 5].
miR-211, the other member of the same microRNA family, was also prioritized in light of its higher expression level in A375-VR vs A375 (~80-fold according to small RNA-seq, ~2-fold according to qRT-PCR) . Since in A375 cells the basal levels of miR-211 are substantially lower than those of miR-204 (please refer to Figure 1 and its caption for details about the analysis of microRNA expression levels), these data raise multiple concerns.
First, it is unclear why the authors discarded miR-504 due to its low expression levels and yet they went after miR-211 that is expressed even less (Figure 1).
Second, the accuracy of mature miR-211 detection by qRT-PCR is questionable. No evidence is provided about the specificity of the Taqman probes used, in spite of the fact that miR-211 is very similar in sequence to miR-204. In addition, the location of the primers used for the qRT-PCR detection of TRPM1 host gene is suboptimal (Figure 2). miR-204 is likely detected instead of or together with miR-211 and this is why they both show a ~2-fold increase in expression according to qRT-PCR.
Finally, we question the biological relevance of a microRNA that is still expressed at very low levels even when upregulated. Since they belong to the same family, it is not surprising that miR-211 behaves like miR-204, if exogenously overexpressed [1, 2]. However, we and others have shown that the appropriate biological context to study endogenous miR-211 are not amelanotic cells, like A375 cells, but melanotic ones: only there miR-211 shows high basal levels (actually higher than those of miR-204) and is able to limit the efficacy of vemurafenib, by exerting its MITF-dependent pro-pigmentation activity [2, 6, 7].
CONFLICTS OF INTEREST
The authors declare no potential conflicts of interest.
Last Modified: 2018-08-24 01:32:37 EDT