Multi-subunit and single-chain E3 ligases that regulate T cell function. (a) Multi-subunit E3 ligases (Skp1-Cullin-F-box (SCF), Cullin-Elongin BC-SOCS/VHL (CBC), and U-box) are anchored by a Cullin scaffold protein and recruit an E2 ubiquitin-conjugating enzyme via a Roc1 or Rbx RING protein (as shown in blue). Substrate specificity is determined by the binding of the target protein (either with or without the carboxyl terminus of Hsc70-interacting protein (CHIP) and Hsc70 containing pre-ubiquitin complex) to a particular F-box (e.g., Skp2), suppressor of cytokine signaling (SOCS), or von Hippel-Lindau (VHL) protein (red), and is mediated by a Skp1 or Elongin BC adapter protein [6,15,29]. (b) Single-chain E3 ligases contain RING or homologous to E6-associated protein carboxyl terminus (HECT) E2 recruitment (blue) and substrate binding (red) domains within one polypeptide [31,50,93]. The question mark on the putative GRAIL target protein indicates that no substrate has yet been identified. C2, Ca2+ binding; PA, protease-associated; Pro, proline rich; TKB, tyrosine kinase binding; TM, transmembrane; UBA, ubiquitin-associated; WW, two tryptophan repeat.