PCGF2 negatively regulates arsenic trioxide-induced PML-RARA protein degradation via UBE2I inhibition in NB4 cells

• Published in: Biochimica et biophysica acta Vol. 1863; no. 7; pp. 1499 - 1509
• Main Authors: Jo, Sungsin, Lee, Young Lim, Kim, Sojin, Lee, Hongki, Chung, Heekyoung
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.07.2016
• Subjects: Antineoplastic Agents - pharmacology; Arsenic trioxide; Arsenicals - pharmacology; Cell Line, Tumor; Fluorescent Antibody Technique; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; HEK293 Cells; Humans; Immunoprecipitation; Leukemia, Promyelocytic, Acute - drug therapy; Leukemia, Promyelocytic, Acute - enzymology; Leukemia, Promyelocytic, Acute - genetics; Leukemia, Promyelocytic, Acute - pathology; Oncogene Proteins, Fusion - genetics; Oncogene Proteins, Fusion - metabolism; Oxides - pharmacology; PCGF2; PML-RARA; Polycomb Repressive Complex 1 - genetics; Polycomb Repressive Complex 1 - metabolism; Protein Binding; Protein degradation; Proteolysis; RNA Interference; Signal Transduction - drug effects; Sumoylation; Time Factors; Transfection; UBE2I; Ubiquitin-Conjugating Enzymes - genetics; Ubiquitin-Conjugating Enzymes - metabolism; Ubiquitination; AML; PBS; Protein degradation; UBE2I; PCGF2; Arsenic trioxide; PML; BSA; CHX; PML-RARA; NB; PcG; FBS; Sumoylation; qPCR; ATO; ATRA; APL
• DOI: 10.1016/j.bbamcr.2016.03.019
• ISSN: 0167-4889; 0006-3002

Arsenic trioxide (ATO) is a therapeutic agent for acute promyelocytic leukemia (APL) which induces PML-RARA protein degradation via enhanced UBE2I-mediated sumoylation. PCGF2, a Polycomb group protein, has been suggested as an anti-SUMO E3 protein by inhibiting the sumoylation of UBE2I substrates, HSF2 and RANGAP1, via direct interaction. Thus, we hypothesized that PCGF2 might play a role in ATO-induced PML-RARA degradation by interacting with UBE2I. PCGF2 protein was down-regulated upon ATO treatment in human APL cell line, NB4. Knockdown of PCGF2 in NB4 cells, in the absence of ATO treatment, was sufficient to induce sumoylation-, ubiquitylation- and PML nuclear body-mediated degradation of PML-RARA protein. Moreover, overexpression of PCGF2 protected ATO-mediated degradation of ectopic and endogenous PML-RARA in 293T and NB4 cells, respectively. In 293T cells, UBE2I-mediated PML-RARA degradation was reduced upon PCGF2 co-expression. In addition, UBE2I-mediated sumoylation of PML-RARA was reduced upon PCGF2 co-expression and PCGF2-UBE2I interaction was confirmed by co-immunoprecipitation. Likewise, endogenous PCGF2-UBE2I interaction was detected by co-immunoprecipitation and immunofluorescence assays in NB4 cells. Intriguingly, upon ATO-treatment, such interaction was disrupted and UBE2I was co-immunoprecipitated or co-localized with its SUMO substrate, PML-RARA. Taken together, our results suggested a novel role of PCGF2 in ATO-mediated degradation of PML-RARA that PCGF2 might act as a negative regulator of UBE2I via direct interaction. •PCGF2 protein was down-regulated upon ATO treatment•PCGF2 knockdown was sufficient to induce PML-RARA degradation without ATO treatment•PCGF2 overexpression impeded ATO-mediated PML-RARA degradation•PCGF2 inhibited UBE2I activity by direct binding•ATO disrupted PCGF2-UBE2I interaction and enhanced UBE2I recruitment to PML-RARA•PCGF2 negatively regulates ATO-induced PML-RARA degradation via UBE2I inhibition