The melanoma-associated antigen 1 (MAGEA1) protein stimulates the E3 ubiquitin-ligase activity of TRIM31 within a TRIM31-MAGEA1-NSE4 complex

• Published in: Cell cycle (Georgetown, Tex.) Vol. 14; no. 6; pp. 920 - 930
• Main Authors: Kozakova, Lucie, Vondrova, Lucie, Stejskal, Karel, Charalabous, Panagoula, Kolesar, Peter, Lehmann, Alan R, Uldrijan, Stjepan, Sanderson, Christopher M, Zdrahal, Zbynek, Palecek, Jan J
• Format: Journal Article
• Language: English
• Published: United States Taylor & Francis 15.03.2015
• Subjects: Amino Acid Sequence; Carrier Proteins - metabolism; Chromatography, Liquid; HEK293 Cells; Humans; Immunoprecipitation; Models, Biological; Molecular Sequence Data; Multiprotein Complexes - metabolism; Neoplasm Proteins - chemistry; Neoplasm Proteins - metabolism; Peptide Fragments - chemistry; Peptide Fragments - metabolism; Peptides - chemistry; Peptides - metabolism; Protein Binding; Protein Multimerization; RING Finger Domains; Tandem Mass Spectrometry; Tripartite Motif Proteins; Two-Hybrid System Techniques; Ubiquitin-Protein Ligases - chemistry; Ubiquitin-Protein Ligases - metabolism; melanoma-associated antigen family; NSE1-NSE3-NSE4 complex; PCGF6; TRIM41; protein evolution; NSE, non-SMC element; RING-finger proteins; WH, winged helix; TRIM family; Y2H, yeast 2-hybrid; MAGE, melanoma-associated antigen; TRIM8; ubiquitination; TRAF6; MAGEA1; MHD, MAGE homology domain; TRIM, tripartite motif; E3 ubiquitin ligase; MDM4; NSE4/EID family; SMC, structure maintenance of chromosomes; TRIM31; RNF166
• ISSN: 1538-4101; 1551-4005
• DOI: 10.1080/15384101.2014.1000112

The MAGE (Melanoma-associated antigen) protein family members are structurally related to each other by a MAGE-homology domain comprised of 2 winged helix motifs WH/A and WH/B. This family specifically evolved in placental mammals although single homologs designated NSE3 (non-SMC element) exist in most eukaryotes. NSE3, together with its partner proteins NSE1 and NSE4 form a tight subcomplex of the structural maintenance of chromosomes SMC5-6 complex. Previously, we showed that interactions of the WH/B motif of the MAGE proteins with their NSE4/EID partners are evolutionarily conserved (including the MAGEA1-NSE4 interaction). In contrast, the interaction of the WH/A motif of NSE3 with NSE1 diverged in the MAGE paralogs. We hypothesized that the MAGE paralogs acquired new RING-finger-containing partners through their evolution and form MAGE complexes reminiscent of NSE1-NSE3-NSE4 trimers. In this work, we employed the yeast 2-hybrid system to screen a human RING-finger protein library against several MAGE baits. We identified a number of potential MAGE-RING interactions and confirmed several of them (MDM4, PCGF6, RNF166, TRAF6, TRIM8, TRIM31, TRIM41) in co-immunoprecipitation experiments. Among these MAGE-RING pairs, we chose to examine MAGEA1-TRIM31 in detail and showed that both WH/A and WH/B motifs of MAGEA1 bind to the coiled-coil domain of TRIM31 and that MAGEA1 interaction stimulates TRIM31 ubiquitin-ligase activity. In addition, TRIM31 directly binds to NSE4, suggesting the existence of a TRIM31-MAGEA1-NSE4 complex reminiscent of the NSE1-NSE3-NSE4 trimer. These results suggest that MAGEA1 functions as a co-factor of TRIM31 ubiquitin-ligase and that the TRIM31-MAGEA1-NSE4 complex may have evolved from an ancestral NSE1-NSE3-NSE4 complex.