Delta-lactoferrin induces cell death via the mitochondrial death signaling pathway by upregulating bax expression

Abstract

Delta-lactoferrin (∆Lf) is a transcription factor belonging to the lactoferrin family, the expression of which inhibits cell proliferation and leads to Skp1 and DcpS gene transactivation. In this study, we showed that ∆Lf expression also induces cell death via apoptosis in HEK 293 and MCF7 cells using a cell viability assay and DNA fragmentation. Western blot analyses showed that apoptosis was caspase-9, 7 and 8 dependent. Proteolytic cleavage of the endonuclease PARP was significantly increased. The levels of expression of Bcl family members were detected by immunochemistry and showed that the Bcl-xl/Bax and Bcl-2/Bax protein ratios were decreased. We determined that the pro-apoptotic effects of ∆Lf are mainly mediated by the activation of the mitochondria-dependent death-signaling pathway. Apoptosis induction by ∆Lf is concomitant with increased cellular levels of Bax protein. Analysis of the Bax promoter region detected a ∆Lf response element located at -155 bp from the transcription start site. Both luciferase reporter gene and chromatin immunoprecipitation assays confirmed that ∆Lf interacts in vitro and in vivo specifically with this sequence. Its deletion, realized using directed mutagenesis, totally abolished ∆Lf transcriptional activity, identifying it as a ∆Lf-responsive element. These results indicate that the Bax gene is a novel ∆Lf target. Moreover we also showed that the O-GlcNAc/P interplay, which controls ∆Lf transcriptional activity, modulates Bax transactivation.

Detection and identification of O‐GlcNAcylated proteins by proteomic approaches

“…These interplays offer theoretically infinite combinations of PTMs on protein isoforms in the human proteome. The diversity of O ‐GlcNAcylated proteins, the dynamism of O ‐GlcNAcylation and its interaction with other PTMs make this glycosylation a regulator of most of the basic and fundamental cellular processes such as cell signaling , cell cycle , and apoptosis . Deregulation of O ‐GlcNAc cycling has been linked with the etiology of diabetes , cancers , cardiovascular diseases , and neuronal disorders .…”

Evaluation of the Chemical Reporter Analog PNP‐6AzGlcNAc as an O‐GlcNAcase Substrate

“…1 The O-GlcNAc modification is nutrient sensitive, 2 and part of a signaling network that plays key roles in many fundamental biological processes, including signal transduction 3-5 epigenetic regulation, 6,7 stress response, 8,9 and protein degradation, 10,11 and apoptosis. 12,13 Only two human enzymes, O-GlcNAc transferase (OGT) 14,15 and OGA 16 are responsible for the addition and removal of O-GlcNAc, respectively. Aberrant O-GlcNAc profiles have been suggested to play a mechanistic role in the etiology and pathological progression of several diseases, including diabetes, [17][18][19] cancers, [20][21][22] and neurodegenerative disorders.…”

Modification by SUMOylation Controls Both the Transcriptional Activity and the Stability of Delta-Lactoferrin

“…Two regions of Lf in which a strong concentration of positive charges were found could be good candidates [ 10 ]. ΔLf exhibits antitumoral activities and we previously showed that overexpression of ΔLf leads to cell cycle arrest at the G1/S transition and apoptosis [ 11 , 12 ]. ΔLf mainly exerts its anti-proliferative and pro-apoptotic activities via its role as a transcription factor.…”

“…ΔLf mainly exerts its anti-proliferative and pro-apoptotic activities via its role as a transcription factor. Indeed, ΔLf transactivates different target genes such as Skp1 , DcpS , Bax , SelH , GTF2F2 and UBE2E1 [ 9 , 12 – 14 ]. A genome-wide pathway analysis and our quantitative proteomic analysis showed that the re-introduction of Lf isoforms in cancerous cells modified essential genes and/or signaling networks responsible mainly for cell survival, apoptosis and RNA processing [ 14 , 15 ].…”