scholarly journals Direct Conjugation of NEDD8 to the N-Terminus of a Model Protein Can Induce Degradation

Cells ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 854
Author(s):  
Kartikeya Vijayasimha ◽  
Marilyn Vo Tran ◽  
Amy L. Leestemaker-Palmer ◽  
Brian P. Dolan
Keyword(s):  
Protein Degradation ◽  
Model System ◽  
Precursor Cell ◽  
Neural Precursor Cell ◽  
Conjugation System ◽  
Covalent Linkage ◽  
N Terminus ◽  
Model Protein ◽  
Ubiquitin Conjugation

While the role of ubiquitin in protein degradation is well established, the role of other ubiquitin-like proteins (UBLs) in protein degradation is less clear. Neural precursor cell expressed developmentally down-regulated protein 8 (NEDD8) is the UBL with the highest level of amino acids identified when compared to ubiquitin. Here we tested if the N-terminal addition of NEDD8 to a protein of interest could lead to degradation. Mutation of critical glycine residues required for normal NEDD8 processing resulted in a non-cleavable fusion protein that was rapidly degraded within the cells by both the proteasome and autophagy. Both degradation pathways were dependent on a functional ubiquitin-conjugation system as treatment with MLN7243 increased levels of non-cleavable NEDD8-GFP. The degradation of non-cleavable, N-terminal NEDD8-GFP was not due to a failure of GFP folding as different NEDD8-GFP constructs with differing abilities to fold and fluoresce were similarly degraded. Though the fusion of NEDD8 to a protein resulted in degradation, treatment of cells with MLN4924, an inhibitor of the E1 activating enzyme for NEDD8, failed to prevent degradation of other destabilized substrates. Taken together these data suggest that under certain conditions, such as the model system described here, the covalent linkage of NEDD8 to a protein substrate may result in the target proteins degradation.

scholarly journals The Role of Mcl-1 in Embryonic Neural Precursor Cell Apoptosis

2021 ◽  
Vol 9 ◽  
Author(s):  
Robert T. Flemmer ◽  
Sarah P. Connolly ◽  
Brittany A. Geizer ◽  
Joseph T. Opferman ◽  
Jacqueline L. Vanderluit
Keyword(s):  
Spinal Cord ◽  
Nervous System ◽  
Precursor Cell ◽  
Embryonic Lethality ◽  
Conditional Knockout ◽  
Neural Precursor ◽  
Null Mouse ◽  
Neural Precursor Cell ◽  
Thoracic Spinal Cord

Myeloid cell leukemia-1 (Mcl-1), an anti-apoptotic Bcl-2 protein, regulates neural precursor cell (NPC) survival in both the developing and adult mammalian nervous system. It is unclear when during the neurogenic period Mcl-1 becomes necessary for NPC survival and whether Bax is the sole pro-apoptotic target of Mcl-1. To address these questions, we used the nervous system-specific Nestin-Cre Mcl-1 conditional knockout mouse line (Mcl-1 CKO) to assess the anti-apoptotic role of Mcl-1 in developmental neurogenesis. Loss of Mcl-1 resulted in a wave of apoptosis beginning in the brainstem and cervical spinal cord at embryonic day 9.5 (E9.5) and in the forebrain at E10.5. Apoptosis was first observed ventrally in each region and spread dorsally over time. Within the spinal cord, apoptosis also spread in a rostral to caudal direction following the path of differentiation. Breeding the Mcl-1 CKO mouse with the Bax null mouse rescued the majority of NPC from apoptosis except in the dorsomedial brainstem and ventral thoracic spinal cord where only 50% were rescued. This demonstrates that Mcl-1 promotes NPC survival primarily by inhibiting the activation of Bax, but that Bax is not the sole pro-apoptotic target of Mcl-1 during embryonic neurogenesis. Interestingly, although co-deletion of Bax rescued the majority of NPC apoptosis, it resulted in embryonic lethality at E13, whereas conditional deletion of both Mcl-1 and Bax rescued embryonic lethality. In summary, this study demonstrates the widespread dependency on Mcl-1 during nervous system development.

scholarly journals The Role of Selective Protein Degradation in the Regulation of Iron and Sulfur Homeostasis in Plants

2020 ◽  
Vol 21 (8) ◽  
pp. 2771 ◽  
Author(s):  
Anna Wawrzyńska ◽  
Agnieszka Sirko
Keyword(s):  
Protein Degradation ◽  
Plant Development ◽  
Sulfur Metabolism ◽  
High Energy ◽  
26S Proteasome ◽  
Control Mechanisms ◽  
Wide Range ◽  
Ubiquitin Conjugation ◽  
Cellular Components

Plants are able to synthesize all essential metabolites from minerals, water, and light to complete their life cycle. This plasticity comes at a high energy cost, and therefore, plants need to tightly allocate resources in order to control their economy. Being sessile, plants can only adapt to fluctuating environmental conditions, relying on quality control mechanisms. The remodeling of cellular components plays a crucial role, not only in response to stress, but also in normal plant development. Dynamic protein turnover is ensured through regulated protein synthesis and degradation processes. To effectively target a wide range of proteins for degradation, plants utilize two mechanistically-distinct, but largely complementary systems: the 26S proteasome and the autophagy. As both proteasomal- and autophagy-mediated protein degradation use ubiquitin as an essential signal of substrate recognition, they share ubiquitin conjugation machinery and downstream ubiquitin recognition modules. Recent progress has been made in understanding the cellular homeostasis of iron and sulfur metabolisms individually, and growing evidence indicates that complex crosstalk exists between iron and sulfur networks. In this review, we highlight the latest publications elucidating the role of selective protein degradation in the control of iron and sulfur metabolism during plant development, as well as environmental stresses.

scholarly journals Insights Into the Biological Role of NEDD4L E3 Ubiquitin Ligase in Human Cancers

2021 ◽  
Vol 11 ◽  
Author(s):  
Shangdan Xie ◽  
Lu Xia ◽  
Yizuo Song ◽  
Hejing Liu ◽  
Zhi-wei Wang ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Therapeutic Strategy ◽  
Potential Role ◽  
E3 Ubiquitin Ligase ◽  
Precursor Cell ◽  
Neural Precursor ◽  
Neural Precursor Cell ◽  
Cancer Types ◽  
Clinical Drugs

Neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) is an E3 ubiquitin ligase that has been reported to participate in multiple cellular procedures by regulating of substrate ubiquitination and subsequent protein degradation. A great amount of evidence has demonstrated that NEDD4L mainly functions as a tumor suppressor in most cancer types, while it also acts as an oncogene in a few cancers. In this review, we summarize the potential role of NEDD4L in carcinogenesis and the related underlying molecular mechanism to improve our understanding of its functions in the tumorigenesis of human malignancies. Developing clinical drugs targeting NEDD4L could be a potential therapeutic strategy for cancer therapy in the future.

Role of the Ubiquitin Conjugation System in the Maintenance of Mitochondrial Homeostasis

2008 ◽  
Vol 1147 (1) ◽  
pp. 242-253 ◽  
Author(s):  
Albert Neutzner ◽  
Giovanni Benard ◽  
Richard J. Youle ◽  
Mariusz Karbowski
Keyword(s):  
Conjugation System ◽  
Mitochondrial Homeostasis ◽  
Ubiquitin Conjugation

scholarly journals Regulation of cancer-related pathways by protein NEDDylation and strategies for the use of NEDD8 inhibitors in the clinic

2014 ◽  
Vol 22 (1) ◽  
pp. T55-T70 ◽  
Author(s):  
Naima Abidi ◽  
Dimitris P Xirodimas
Keyword(s):  
Therapeutic Intervention ◽  
Biological Process ◽  
Precursor Cell ◽  
Neural Precursor ◽  
Neural Precursor Cell ◽  
Post Translational Modification ◽  
Recent Advances

Post-translational modification of proteins with ubiquitin and ubiquitin-like molecules (UBLs) controls a vast if not every biological process in the cell. It is not surprising that deregulation in ubiquitin and UBL signalling has been implicated in the pathogenesis of many diseases and that these pathways are considered as major targets for therapeutic intervention. In this review, we summarise recent advances in our understanding of the role of the UBL neural precursor cell expressed developmentally downregulated-8 (NEDD8) in cancer-related processes and potential strategies for the use of NEDD8 inhibitors as chemotherapeutics.

scholarly journals Effect of directional pulling on mechanical protein degradation by ATP-dependent proteolytic machines

2017 ◽  
Vol 114 (31) ◽  
pp. E6306-E6313 ◽  
Author(s):  
Adrian O. Olivares ◽  
Hema Chandra Kotamarthi ◽  
Benjamin J. Stein ◽  
Robert T. Sauer ◽  
Tania A. Baker
Keyword(s):  
Escherichia Coli ◽  
Protein Degradation ◽  
Single Molecule ◽  
Wild Type ◽  
N Terminus ◽  
Multiple Copies ◽  
Rate Limiting ◽  
Hydrolysis Of ◽  
Translocation Speed

AAA+ proteases and remodeling machines couple hydrolysis of ATP to mechanical unfolding and translocation of proteins following recognition of sequence tags called degrons. Here, we use single-molecule optical trapping to determine the mechanochemistry of two AAA+ proteases, Escherichia coli ClpXP and ClpAP, as they unfold and translocate substrates containing multiple copies of the titinI27 domain during degradation initiated from the N terminus. Previous studies characterized degradation of related substrates with C-terminal degrons. We find that ClpXP and ClpAP unfold the wild-type titinI27 domain and a destabilized variant far more rapidly when pulling from the N terminus, whereas translocation speed is reduced only modestly in the N-to-C direction. These measurements establish the role of directionality in mechanical protein degradation, show that degron placement can change whether unfolding or translocation is rate limiting, and establish that one or a few power strokes are sufficient to unfold some protein domains.

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