Eukaryotic Initiation Factor 4H Is under Transcriptional Control of p65/NF-κB

Abstract

Protein synthesis is mainly regulated at the initiation step, allowing the fast, reversible and spatial control of gene expression. Initiation of protein synthesis requires at least 13 translation initiation factors to assemble the 80S ribosomal initiation complex. Loss of translation control may result in cell malignant transformation. Here, we asked whether translational initiation factors could be regulated by NF-κB transcription factor, a major regulator of genes involved in cell proliferation, survival, and inflammatory response. We show that the p65 subunit of NF-κB activates the transcription of eIF4H gene, which is the regulatory subunit of eIF4A, the most relevant RNA helicase in translation initiation. The p65-dependent transcriptional activation of eIF4H increased the eIF4H protein content augmenting the rate of global protein synthesis. In this context, our results provide novel insights into protein synthesis regulation in response to NF-κB activation signalling, suggesting a transcription-translation coupled mechanism of control.

CRL3IBTK Regulates the Tumor Suppressor Pdcd4 through Ubiquitylation Coupled to Proteasomal Degradation

“…The shRNA-IBtkα targets the 2077–2098 nucleotides of IBtkα mRNA (NCBI reference sequence: XM_006715453.1). Lentiviral particles were produced in HEK293T cells, as described previously ( 28 , 29 ).…”

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“…HeLa and HEK293T were cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% heat-inactivated foetal calf serum and 2 mM l -glutamine (Lonza Cologne AG, Cologne, Germany); K562 was cultured in RPMI1640 (Lonza Cologne AG, Cologne, Germany) supplemented with 10% heat-inactivated foetal calf serum and 2 mM l -glutamine. Human peripheral blood mononuclear cells (PBMCs) were derived from buffy coats of three healthy donors and isolated by Ficoll Paque gradient (GE Healthcare Europe, Munich, Germany), as previously described [25,26,27]. Briefly, blood samples were diluted 1:1 in PBS and stratified on Ficoll solution with a 3:1 ( v / v ) ratio.…”

“…Protein extracts were obtained as previously described [25,29]. Briefly, HeLa and K562 cells were harvested, washed twice with PBS 1X and lysed in RIPA buffer, containing 1% NP-40, 10 mM Tris-HCl, 150 mM NaCl, and 1 mM EDTA, supplemented with a protease inhibitor cocktail (cOmplete-mini EDTA-free tablets—Roche) on ice for 20 min.…”

Cancer-Associated CD43 Glycoforms as Target of Immunotherapy

“…Peripheral blood mononuclear cells were isolated from healthy donors by Ficoll-Paque gradient centrifugation (GE Healthcare Life Sciences, Piscataway, NJ, USA), as previously described (27-29). Primary cultured NK cell populations were obtained from 10-day co-cultures of PBMCs with irradiated Epstein-Barr virus-positive RPMI 8866 lymphoblastoid cell line, as previously described (30).…”