BTBD1 and BTBD2 colocalize to cytoplasmic bodies with the RBCC/tripartite motif protein, TRIM5δ

• Published in: Experimental cell research 2003-08, Vol.288 (1), p.84-93
• Main Authors: Xu, Lixin, Yang, Lihong, Moitra, Prasun K, Hashimoto, Keiko, Rallabhandi, Prasad, Kaul, Sunil, Meroni, Germana, Jensen, Jane P, Weissman, Allan M, D’Arpa, Peter
• Format: Article
• Language: English
• Subjects: BTB/POZ; BTBD1; BTBD2; Cytoplasmic bodies; RBCC domain; TRIM5; Tripartite domain
• ISSN: 0014-4827; 1090-2422
• DOI: 10.1016/S0014-4827(03)00187-3

We previously identified BTBD1 and BTBD2 as novel topoisomerase I-interacting proteins that share 80% amino acid identity. Here we report the characterization of their subcellular localization. In a number of mouse and human cells, BTBD1 and BTBD2 (BTBD1/2) colocalized to punctate or elongated cytoplasmic bodies (< 5 μm long and several per cell) that were larger and more elongated in cancer cell lines than in fibroblasts and myoblasts. A search for potential colocalizing proteins identified TRIM family members that localize to morphologically similar cytoplasmic bodies, which were then tested for colocalization with BTBD1/2. TRIM5δ, expressed as a GFP fusion, colocalized with BTBD1/2 immunostaining and appeared to serve as a scaffold for the assembly of endogenous BTBD1/2 proteins. TRIM family members contain a RING domain, B-box(es), and coiled-coil regions, which have a characteristic order and spacing (RBCC domain). RING-dependent ubiquitin ligase activity and multimerization via the coiled-coil region may be defining properties of the RBCC/TRIM protein family. We found that TRIM5δ with a deleted coiled-coil region or a mutated RING domain failed to colocalize with BTBD1/2. Additionally, TRIM5δ ubiquitylated itself in a RING finger- and UbcH5B-dependent manner. BTBD1/2 each contain a PHR-similarity region, repeated twice on the putative ubiquitin ligases PAM, highwire and RPM-1, which also contain a RING and B-box. Thus, four protein modules found on each of these putative ubiquitin ligases, a RING, a B-box and two PHR repeats, are present on BTBD1/2 and TRIM5δ that are colocalized to cytoplasmic bodies.