Article

The Ubiquitin Conjugating Enzyme, UbcM2, Engages in Novel Interactions with Components of Cullin-3 Based E3 Ligases †

Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
Biochemistry (Impact Factor: 3.02). 04/2009; 48(15):3527-37. DOI: 10.1021/bi801971m
Source: PubMed

ABSTRACT

The class III ubiquitin conjugating enzymes (E2s) are distinguished from other E2s by the presence of unique N-terminal domains, and the utilization of importin-11 for transport into the nucleus in an activation dependent fashion. To begin determining the physiological roles of these enzymes, we carried out a yeast two-hybrid screen with the class III E2, UbcM2. This screen retrieved RCBTB1, a putative substrate adaptor for a cullin3 (CUL3) E3 ligase. We initially established through biochemical studies that RCBTB1 has the properties of a CUL3 substrate adaptor. Further analysis of the UbcM2-RCBTB1 complex led to the discovery and characterization of the following novel interactions: (i) UbcM2 binds an N-terminal domain of CUL3 requiring the first 57 amino acids, the same domain that binds to RCBTB1 and other substrate adaptors; (ii) UbcM2 does not bind mutants of CUL3 that are deficient in substrate adaptor recruitment; (iii) UbcM2 interacts with CUL3 independent of a bridging RING-finger protein; and (iv) can engage the neddylated (i.e., activated) form of CUL3. We also present evidence that UbcM2 can bind to the N-terminal halves of multiple cullins, implying that this E2 is a general cofactor for this class of ligases. Together, these studies represent the first evidence that UbcM2, in concert with substrate adaptors, engages activated CUL3 ligases, thus suggesting that class III E2s are novel regulators of cullin ligases.

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    • "This effect of Imp-11 depletion on nuclear KEAP1 accumulation is likely due to disrupted import of UBE2E3 (Plafker et al., 2004). Increased cytoplasmic UBE2E3 binds to and stabilizes CUL3 KEAP1 in the cytoplasm (Figure 7B; Plafker et al., 2009), thereby reducing KEAP1 dissociation from CUL3, a necessary step for bulk KEAP1 import (Sun et al., 2007). These studies additionally identify a new function for an import receptor. "

    Full-text · Dataset · Feb 2015
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    • "This effect of Imp-11 depletion on nuclear KEAP1 accumulation is likely due to disrupted import of UBE2E3 (Plafker et al., 2004). Increased cytoplasmic UBE2E3 binds to and stabilizes CUL3 KEAP1 in the cytoplasm (Figure 7B; Plafker et al., 2009), thereby reducing KEAP1 dissociation from CUL3, a necessary step for bulk KEAP1 import (Sun et al., 2007). These studies additionally identify a new function for an import receptor. "
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    • "The mouse versions are referred to as UbcM3, UBE2E2, and UbcM2, respectively, and each is identical to its human counterpart [17] (see Table 1). These enzymes share a host of properties, including: 1) a steady-state nuclear distribution [18,19], 2) entering the nucleus via the importin-11 transport receptor [18,19], 3) binding common E3 ligase partners (e.g., [20,21]), and 4) interacting with the N-terminal domain of various cullin proteins [21]. An additional distinguishing feature among these enzymes is that each has a unique N-terminal domain of 40–60 residues [17]. "
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