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The recently discovered MCPIP1 (monocyte chemoattractant protein-induced protein 1), a multidomain protein encoded by the MCPIP1 (ZC3H12A) gene, has been described as a new differentiation factor, a ribonuclease, and a deubiquitination-supporting factor. However, its role in cancer is poorly recognized. Our recent analysis of microarrays data showed a lack of expression of the MCPIP1 transcript in primary neuroblastoma, the most common extracranial solid tumor in children. Additionally, enforced expression of the MCPIP1 gene in BE(2)-C cells caused a significant decrease in neuroblastoma proliferation and viability. Aim of the present study was to further investigate the role of MCPIP1 in neuroblastoma, using expression DNA microarrays and microRNA microarrays. Transient transfections of BE(2)-C cells were used for overexpression of either wild type of MCPIP1 (MCPIP1-wt) or its RN-ase defective mutant (MCPIP1-ΔPIN). We have analyzed changes of transcriptome and next, we have used qRT-PCR to verify mRNA levels of selected genes responding to MCPIP1 overexpression. Additionally, protein levels were determined for some of the selected genes. The choline transporter, CTL1, encoded by the SLC44A1 gene, was significantly repressed at the specific mRNA and protein levels and most importantly this translated into a decreased choline transport in MCPIP1-overexpressing cells. Then, we have found microRNA-3613-3p as the mostly altered in the pools of cells overexpressing the wild type MCPIP1. Next, we analyzed the predicted targets of the miR-3613-3p and validated them using qRT-PCR and western blot. These results indicate that the expression of miR-3613-3p might be regulated by MCPIP1 by cleavage of its precursor form.

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The GD2 ganglioside expressed on neuroectodermally derived tumors, including neuroblastoma and melanoma, is weakly immunogenic in tumor-bearing patients and induces predominantly immunoglobulin (Ig)-M antibody responses in the immunized host. Here, we investigated whether interconversion of GD2 into a peptide mimetic form would induce GD2 cross-reactive IgG antibody responses in mice. Screening of the X 15 phage display peptide library with the anti-GD2

BackgroundMCPIP is a novel CCCH zinc finger protein described as an RNase engaged in the regulation of immune responses. The regulation of expression of the gene coding for MCPIP - ZC3H12A is poorly explored.ResultsHere we report that the proinflammatory cytokine IL-1β rapidly induces the synthesis of MCPIP in primary monocyte-derived macrophages and HepG2 cells. This up-regulation takes place through the MAP kinase pathway and following activation of the transcription factor Elk-1. Using a ZC3H12A reporter construct we have shown that a ZC3H12A promoter region, stretching from -76 to +60, mediates activation by IL-1β. This region contains binding sites for Elk-1 and its partner SRF. Chromatin immunoprecipitation analysis confirms in vivo binding of both transcription factors to this region of the ZC3H12A promoter.ConclusionsWe conclude that the transcription factor Elk-1 plays an important role in the activation of ZC3H12A expression in response to IL-1β stimulation.

The main physiological function of MCPIP1 (regnase-1) is negative regulation of inflammation. Moreover, roles of regnase-1 in apoptosis and differentiation have also been described, but its involvement in cancer is yet to be fully recognized. Earlier, we showed a lack of expression of MCPIP1 in both primary tumors and several neuroblastoma cell lines. Additionally, we reported that levels of MCPIP1 and the key neuroblastoma oncoprotein-MYCN were inversely correlated in BE(2)-C clones overexpressing the MCPIP1 gene. Here, we show that exogenous expression of the MCPIP1 protein decreases MYCN mRNA and protein levels without changing the MYCN mRNA half-life. Furthermore, it was shown that MCPIP1-wt exogenous expression affects levels and phosphorylation of MYCN partners such as Aurora A (Thr288), CDC2 (Tyr15 and Thr161), GSK3β (Ser9), and key cellular components of Akt/mTOR signaling, which regulate MYCN stability and activation. In accordance with the obtained results, we found increased phosphorylation of MYCN protein at Thr58 that causes destabilization of the oncoprotein. Moreover, it is shown that exogenous expression of MCPIP1 does not cause apoptosis. Our data extend knowledge on roles of MCPIP1 in our model and link the protein to regulation of expression and stability of MYCN through decrease of signaling via Akt/mTOR pathway. J. Cell. Biochem. 118: 1741-1755, 2017. © 2016 Wiley Periodicals, Inc.

The importance of monocyte chemotactic protein-1-induced protein 1 (MCPIP1) in the negative regulation of inflammatory reactions has already been extensively studied. However, its role in cancer is not yet established. We studied MCPIP1 gene expression in primary human neuroblastomas and several neuroblastoma cell lines. Our results showed a lack of MCPIP1 expression in primary neuroblastoma tumors. Moreover, it was found that the low expression of the protein measured in human neuroblastoma cell lines might be important for neuroblastoma survival, since enforced MCPIP1 gene expression in human neuroblastoma BE(2)-C cells caused a significant decrease in neuroblastoma cell viability and proliferation.

Monoclonal antibodies targeting GD2 ganglioside (GD2)have recently been approved for the treatment of high risk neuroblastoma and are extensively evaluated in clinics in other indications. This study illustrates how a therapeutic antibody distinguishes between different types of gangliosides present on normal and cancer cells and informs how synthetic peptides can imitate ganglioside in its binding to the antibody. Using high resolution crystal structures we demonstrate that the ganglioside recognition by a model antibody (14G2a) is based primarily on an extended network of direct and water molecule mediated hydrogen bonds. Comparison of the GD2-Fab structure with that of a ligand free antibody reveals an induced fit mechanism of ligand binding. These conclusions are validated by directed mutagenesis and allowed structure guided generation of antibody variant with improved affinity toward GD2. Contrary to the carbohydrate, both evaluated mimetic peptides utilize a "key and lock" interaction mechanism complementing the surface of the antibody binding groove exactly as found in the empty structure. The interaction of both peptides with the Fab relies considerably on hydrophobic contacts however, the detailed connections differ significantly between the peptides. As such, the evaluated peptide carbohydrate mimicry is defined primarily in a functional and not in structural manner. Molecular & Cellular