Non-canonical substrate recognition by the human WDR26-CTLH E3 ligase regulates prodrug metabolism

Karthik V. Gottemukkala; Jakub Chrustowicz; Dawafuti Sherpa; Sara Sepic; Duc Tung Vu; Özge Karayel; Eleftheria C. Papadopoulou; Annette Gross; Kenji Schorpp; Susanne von Gronau; Kamyar Hadian; Peter J. Murray; Matthias Mann; Brenda A. Schulman; Arno F. Alpi

doi:10.1016/j.molcel.2024.04.014

Non-canonical substrate recognition by the human WDR26-CTLH E3 ligase regulates prodrug metabolism

Karthik V. Gottemukkala, Jakub Chrustowicz, Dawafuti Sherpa, Sara Sepic, Duc Tung Vu, Özge Karayel, Eleftheria C. Papadopoulou, Annette Gross, Kenji Schorpp, Susanne von Gronau, Kamyar Hadian, Peter J. Murray, Matthias Mann, Brenda A. Schulman, Arno F. Alpi^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

Abstract

The yeast glucose-induced degradation-deficient (GID) E3 ubiquitin ligase forms a suite of complexes with interchangeable receptors that selectively recruit N-terminal degron motifs of metabolic enzyme substrates. The orthologous higher eukaryotic C-terminal to LisH (CTLH) E3 complex has been proposed to also recognize substrates through an alternative subunit, WDR26, which promotes the formation of supramolecular CTLH E3 assemblies. Here, we discover that human WDR26 binds the metabolic enzyme nicotinamide/nicotinic-acid-mononucleotide-adenylyltransferase 1 (NMNAT1) and mediates its CTLH E3-dependent ubiquitylation independently of canonical GID/CTLH E3-family substrate receptors. The CTLH subunit YPEL5 inhibits NMNAT1 ubiquitylation and cellular turnover by WDR26-CTLH E3, thereby affecting NMNAT1-mediated metabolic activation and cytotoxicity of the prodrug tiazofurin. Cryoelectron microscopy (cryo-EM) structures of NMNAT1- and YPEL5-bound WDR26-CTLH E3 complexes reveal an internal basic degron motif of NMNAT1 essential for targeting by WDR26-CTLH E3 and degron mimicry by YPEL5’s N terminus antagonizing substrate binding. Thus, our data provide a mechanistic understanding of how YPEL5-WDR26-CTLH E3 acts as a modulator of NMNAT1-dependent metabolism.

Original language	English
Pages (from-to)	1948-1963.e11
Number of pages	28
Journal	Molecular Cell
Volume	84
Issue number	10
DOIs	https://doi.org/10.1016/j.molcel.2024.04.014
Publication status	Published - 16 May 2024
Externally published	Yes

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10.1016/j.molcel.2024.04.014

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Gottemukkala, K. V., Chrustowicz, J., Sherpa, D., Sepic, S., Vu, D. T., Karayel, Ö., Papadopoulou, E. C., Gross, A., Schorpp, K., Gronau, S. V., Hadian, K., Murray, P. J., Mann, M., Schulman, B. A., & Alpi, A. F. (2024). Non-canonical substrate recognition by the human WDR26-CTLH E3 ligase regulates prodrug metabolism. Molecular Cell, 84(10), 1948-1963.e11. https://doi.org/10.1016/j.molcel.2024.04.014

@article{3cb4c699f253412db8f0c4580bb4e06c,

title = "Non-canonical substrate recognition by the human WDR26-CTLH E3 ligase regulates prodrug metabolism",

abstract = "The yeast glucose-induced degradation-deficient (GID) E3 ubiquitin ligase forms a suite of complexes with interchangeable receptors that selectively recruit N-terminal degron motifs of metabolic enzyme substrates. The orthologous higher eukaryotic C-terminal to LisH (CTLH) E3 complex has been proposed to also recognize substrates through an alternative subunit, WDR26, which promotes the formation of supramolecular CTLH E3 assemblies. Here, we discover that human WDR26 binds the metabolic enzyme nicotinamide/nicotinic-acid-mononucleotide-adenylyltransferase 1 (NMNAT1) and mediates its CTLH E3-dependent ubiquitylation independently of canonical GID/CTLH E3-family substrate receptors. The CTLH subunit YPEL5 inhibits NMNAT1 ubiquitylation and cellular turnover by WDR26-CTLH E3, thereby affecting NMNAT1-mediated metabolic activation and cytotoxicity of the prodrug tiazofurin. Cryoelectron microscopy (cryo-EM) structures of NMNAT1- and YPEL5-bound WDR26-CTLH E3 complexes reveal an internal basic degron motif of NMNAT1 essential for targeting by WDR26-CTLH E3 and degron mimicry by YPEL5{\textquoteright}s N terminus antagonizing substrate binding. Thus, our data provide a mechanistic understanding of how YPEL5-WDR26-CTLH E3 acts as a modulator of NMNAT1-dependent metabolism.",

author = "Gottemukkala, \{Karthik V.\} and Jakub Chrustowicz and Dawafuti Sherpa and Sara Sepic and Vu, \{Duc Tung\} and {\"O}zge Karayel and Papadopoulou, \{Eleftheria C.\} and Annette Gross and Kenji Schorpp and Gronau, \{Susanne von\} and Kamyar Hadian and Murray, \{Peter J.\} and Matthias Mann and Schulman, \{Brenda A.\} and Alpi, \{Arno F.\}",

note = "We thank D. Bollschweiler and T. Sch{\"a}fer for assistance and maintaining the MPIB cryo-EM Core Facility, S. {\"U}bel and S. Pettera in the MPIB Bioorganic Chemistry \& Biophysics Core Facility for peptide synthesis, J. Rajan Prabu for guidance in structural analysis, Josef Kellermann for lab maintenance, members of the Schulman lab for helpful discussions, and L. Parada and Y-J. Chen for exchanging results prior to publication. The views and opinions expressed are, however, those of the authors only and do not necessarily reflect those of the European Union or the European Research Council. Neither the European Union nor the granting authority can be held responsible for them.",

year = "2024",

month = may,

day = "16",

doi = "10.1016/j.molcel.2024.04.014",

language = "English",

volume = "84",

pages = "1948--1963.e11",

journal = "Molecular Cell",

issn = "1097-2765",

publisher = "Cell Press",

number = "10",

}

Gottemukkala, KV, Chrustowicz, J, Sherpa, D, Sepic, S, Vu, DT, Karayel, Ö, Papadopoulou, EC, Gross, A, Schorpp, K, Gronau, SV, Hadian, K, Murray, PJ, Mann, M, Schulman, BA & Alpi, AF 2024, 'Non-canonical substrate recognition by the human WDR26-CTLH E3 ligase regulates prodrug metabolism', Molecular Cell, vol. 84, no. 10, pp. 1948-1963.e11. https://doi.org/10.1016/j.molcel.2024.04.014

TY - JOUR

T1 - Non-canonical substrate recognition by the human WDR26-CTLH E3 ligase regulates prodrug metabolism

AU - Gottemukkala, Karthik V.

AU - Chrustowicz, Jakub

AU - Sherpa, Dawafuti

AU - Sepic, Sara

AU - Vu, Duc Tung

AU - Karayel, Özge

AU - Papadopoulou, Eleftheria C.

AU - Gross, Annette

AU - Schorpp, Kenji

AU - Gronau, Susanne von

AU - Hadian, Kamyar

AU - Murray, Peter J.

AU - Mann, Matthias

AU - Schulman, Brenda A.

AU - Alpi, Arno F.

N1 - We thank D. Bollschweiler and T. Schäfer for assistance and maintaining the MPIB cryo-EM Core Facility, S. Übel and S. Pettera in the MPIB Bioorganic Chemistry & Biophysics Core Facility for peptide synthesis, J. Rajan Prabu for guidance in structural analysis, Josef Kellermann for lab maintenance, members of the Schulman lab for helpful discussions, and L. Parada and Y-J. Chen for exchanging results prior to publication. The views and opinions expressed are, however, those of the authors only and do not necessarily reflect those of the European Union or the European Research Council. Neither the European Union nor the granting authority can be held responsible for them.

PY - 2024/5/16

Y1 - 2024/5/16

N2 - The yeast glucose-induced degradation-deficient (GID) E3 ubiquitin ligase forms a suite of complexes with interchangeable receptors that selectively recruit N-terminal degron motifs of metabolic enzyme substrates. The orthologous higher eukaryotic C-terminal to LisH (CTLH) E3 complex has been proposed to also recognize substrates through an alternative subunit, WDR26, which promotes the formation of supramolecular CTLH E3 assemblies. Here, we discover that human WDR26 binds the metabolic enzyme nicotinamide/nicotinic-acid-mononucleotide-adenylyltransferase 1 (NMNAT1) and mediates its CTLH E3-dependent ubiquitylation independently of canonical GID/CTLH E3-family substrate receptors. The CTLH subunit YPEL5 inhibits NMNAT1 ubiquitylation and cellular turnover by WDR26-CTLH E3, thereby affecting NMNAT1-mediated metabolic activation and cytotoxicity of the prodrug tiazofurin. Cryoelectron microscopy (cryo-EM) structures of NMNAT1- and YPEL5-bound WDR26-CTLH E3 complexes reveal an internal basic degron motif of NMNAT1 essential for targeting by WDR26-CTLH E3 and degron mimicry by YPEL5’s N terminus antagonizing substrate binding. Thus, our data provide a mechanistic understanding of how YPEL5-WDR26-CTLH E3 acts as a modulator of NMNAT1-dependent metabolism.

AB - The yeast glucose-induced degradation-deficient (GID) E3 ubiquitin ligase forms a suite of complexes with interchangeable receptors that selectively recruit N-terminal degron motifs of metabolic enzyme substrates. The orthologous higher eukaryotic C-terminal to LisH (CTLH) E3 complex has been proposed to also recognize substrates through an alternative subunit, WDR26, which promotes the formation of supramolecular CTLH E3 assemblies. Here, we discover that human WDR26 binds the metabolic enzyme nicotinamide/nicotinic-acid-mononucleotide-adenylyltransferase 1 (NMNAT1) and mediates its CTLH E3-dependent ubiquitylation independently of canonical GID/CTLH E3-family substrate receptors. The CTLH subunit YPEL5 inhibits NMNAT1 ubiquitylation and cellular turnover by WDR26-CTLH E3, thereby affecting NMNAT1-mediated metabolic activation and cytotoxicity of the prodrug tiazofurin. Cryoelectron microscopy (cryo-EM) structures of NMNAT1- and YPEL5-bound WDR26-CTLH E3 complexes reveal an internal basic degron motif of NMNAT1 essential for targeting by WDR26-CTLH E3 and degron mimicry by YPEL5’s N terminus antagonizing substrate binding. Thus, our data provide a mechanistic understanding of how YPEL5-WDR26-CTLH E3 acts as a modulator of NMNAT1-dependent metabolism.

UR - http://dx.doi.org/10.1016/j.molcel.2024.04.014

U2 - 10.1016/j.molcel.2024.04.014

DO - 10.1016/j.molcel.2024.04.014

M3 - Article

SN - 1097-2765

VL - 84

SP - 1948-1963.e11

JO - Molecular Cell

JF - Molecular Cell

IS - 10

ER -

Non-canonical substrate recognition by the human WDR26-CTLH E3 ligase regulates prodrug metabolism

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