Physical and Functional Interaction of the HECT Ubiquitin-protein Ligases E6AP and HERC2

Kühnle, Simone; Kogel, Ulrike; Glockzin, Sandra; Marquardt, Andreas; Ciechanover, Aaron; Matentzoglu, Konstantin; Scheffner, Martin

doi:10.1074/jbc.m110.205211

Cited by 67 publications

(82 citation statements)

References 37 publications

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“…Thus, we adopted this strategy to study the interaction of E6AP with UbcH7 and UbcH5b (Fig. S1), which both support E6AP-mediated ubiquitination in vitro (19,35). As shown by isothermal titration calorimetry and size exclusion chromatography, the ubiquitin loading status of UbcH7 does not have a major influence on the ability of UbcH7 to interact with the isolated HECT domain of E6AP (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Whereas some mechanisms controlling transcription of the UBE3A gene have been identified (e.g., the paternal allele is silenced by a UBE3A antisense transcript) (14,18), only little is known about how the E3 activity of E6AP is regulated. We recently reported that E6AP binds to HERC2, a member of the HECT E3 family, and that HERC2 acts as an allosteric activator of E6AP (19). The physiological relevance of this interaction is indicated by the finding that a point mutation in the HERC2 gene, resulting in a mutant HERC2 protein with increased turnover rate and hence decreased protein levels, underlies the development of a neurodevelopmental disorder with AS-like features (20).…”

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confidence: 99%

“…When analyzing the E6AP-HERC2 interaction, we observed that a ubiquitin variant, in which the so-called canonical hydrophobic patch of ubiquitin is mutated (Ub_hpI), is only poorly used by E6AP for ubiquitination and that HERC2 can partially rescue this disability of E6AP (19). This observation prompted us to take a closer look at the role of ubiquitin in E6AP-mediated ubiquitination.…”

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confidence: 99%

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Role of ubiquitin and the HPV E6 oncoprotein in E6AP-mediated ubiquitination

Mortensen

Schneider

Barbic

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Deregulation of the ubiquitin ligase E6 associated protein (E6AP) encoded by the UBE3A gene has been associated with three different clinical pictures. Hijacking of E6AP by the E6 oncoprotein of distinct human papillomaviruses (HPV) contributes to the development of cervical cancer, whereas loss of E6AP expression or function is the cause of Angelman syndrome, a neurodevelopmental disorder, and increased expression of E6AP has been involved in autism spectrum disorders. Although these observations indicate that the activity of E6AP has to be tightly controlled, only little is known about how E6AP is regulated at the posttranslational level. Here, we provide evidence that the hydrophobic patch of ubiquitin comprising Leu-8 and Ile-44 is important for E6AP-mediated ubiquitination, whereas it does not affect the catalytic properties of the isolated catalytic HECT domain of E6AP. Furthermore, we show that the HPV E6 oncoprotein rescues the disability of full-length E6AP to use a respective hydrophobic patch mutant of ubiquitin for ubiquitination and that it stimulates E6AP-mediated ubiquitination of Ring1B, a known substrate of E6AP, in vitro and in cells. Based on these data, we propose that E6AP exists in at least two different states, an active and a less active or latent one, and that the activity of E6AP is controlled by noncovalent interactions with ubiquitin and allosteric activators such as the HPV E6 oncoprotein.I n eukaryotes, posttranslational modification of proteins by ubiquitin plays a pivotal role in the regulation of many cellular processes, including cell cycle, DNA metabolism (e.g., DNA repair, transcription), and various signal transduction pathways (1-4). The specificity of the ubiquitin-conjugation system is mainly ensured by E3 ubiquitin ligases, which mediate the recognition of target proteins. Based on the presence of distinct domains and their mode of action, E3 proteins can be grouped into three families, RING/RING-like E3s, RING-in-between-RING (RBR) E3s, and HECT E3s (5-7). All E3s have interaction sites for both substrate proteins and E2 ubiquitinconjugating enzymes. However, whereas in the case of RBR E3s and HECT E3s, ubiquitin is transferred from the E3 to substrates, RING/RING-like E3s function as adaptors between substrates and E2s (i.e., ubiquitin is transferred from the E2 to the substrate).E6AP, the founding member of the HECT E3 family, was originally identified as an interacting protein of the E6 oncoprotein of cancer-associated human papillomaviruses (HPVs) (8, 9). The E6-E6AP complex targets the tumor suppressor p53 and other proteins-which in the absence of E6 are not targeted by E6AP-for ubiquitination and degradation thereby contributing to HPV-induced cervical carcinogenesis (10, 11). In 1997, it was recognized that alterations in the UBE3A gene, which encodes E6AP, resulting in loss of E6AP expression or in the expression of E6AP variants with compromised E3 activity, are the cause of the Angelman syndrome (AS), a neurodevelopmental disorder (12-14). Recently, it was rep...

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Role of ubiquitin and the HPV E6 oncoprotein in E6AP-mediated ubiquitination

Mortensen

Schneider

Barbic

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…HERC2 stimulates the ubiquitin-ligase activity of UBE3A/E6AP through its RLD2 domain. 4 An impairment of UBE3A/E6AP E3 ligase activity due to a defect of HERC2 is expected to result in a decreased ubiquitination and degradation of UBE3A/E6AP substrates. The impact of HERC2 on UBE3A/E6AP and other specific substrates may contribute to the severity of the disease observed in the patient described here.…”

Section: Accumulation Of Herc2 Substrates Involved In Dna Repairmentioning

confidence: 99%

“…2 HERC2 encodes a 528 kDa E3 ubiquitin ligase, that interacts with E6AP/UBE3A, another ubiquitin ligase, and targets BRCA1 and XPA for degradation. [3][4][5] Until recently, HERC2 mutations were not associated with a human disorder. Lately, a single homozygous mutation of HERC2 was found to segregate in Old Order Amish families with a neurodevelopmental disorder that has some phenotypic similarities to Angelman Syndrome.…”

Section: Introductionmentioning

confidence: 99%

Complete loss of function of the ubiquitin ligase HERC2 causes a severe neurodevelopmental phenotype

et al. 2016

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The ubiquitin-proteasome pathway is involved in the pathogenesis of several neurogenetic diseases. We describe a Mauritanian patient harboring a homozygous deletion restricted to two contiguous genes HERC2 and OCA2 and presenting with severe developmental abnormalities. The deletion causes the complete loss of HERC2 protein function, an E3-ubiquitin ligase. HERC2 is known to target XPA and BRCA1 for degradation and a mechanism whereby it is involved in DNA repair and cell cycle regulation. We showed that loss of HERC2 function leads to the accumulation of XPA and BRCA1 in the patient's fibroblasts and generates decreased sensitivity to apoptosis and increased level of DNA repair. Our data describe for the first time the phenotypic consequences, both at the clinical and cellular levels, of a complete loss of HERC2 function in a patient. They strongly suggest that profound ubiquitin ligase -associated dysfunction is responsible for the severe phenotype in this patient, and that dysfunction of this pathway may be involved in other patients with similar neurodevelopmental diseases.

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Targeting HECT‐type E3 ligases – insights from catalysis, regulation and inhibitors

2017

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Edited by Wilhelm JustUbiquitination plays a pivotal role in most cellular processes and is critical for protein degradation and signalling. E3 ligases are the matchmakers in the ubiquitination cascade, responsible for substrate recognition and modification with specific polyubiquitin chains. Until recently, it was not clear how the catalytic activity of E3s is modulated, but major recent studies on HECT E3 ligases is filling this void. These enzymes appear to be held in a closed, inactive conformation, which is relieved by biochemical manoeuvres unique to each member, thus ensuring exquisite regulation and specificity of the enzymes. The new advances and their significance to the function of HECT E3s are described here, with a particular focus on the Nedd4 family members.

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Physical and Functional Interaction of the HECT Ubiquitin-protein Ligases E6AP and HERC2

Cited by 67 publications

References 37 publications

Role of ubiquitin and the HPV E6 oncoprotein in E6AP-mediated ubiquitination

Role of ubiquitin and the HPV E6 oncoprotein in E6AP-mediated ubiquitination

Complete loss of function of the ubiquitin ligase HERC2 causes a severe neurodevelopmental phenotype

Targeting HECT‐type E3 ligases – insights from catalysis, regulation and inhibitors

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