scholarly journals Discovery of a molecular glue promoting CDK12-DDB1 interaction to trigger cyclin K degradation

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Lu Lv ◽  
Peihao Chen ◽  
Longzhi Cao ◽  
Yamei Li ◽  
Zhi Zeng ◽  
...  
Keyword(s):  
Protein Degradation ◽  
High Throughput Screening ◽  
Human Cancer ◽  
Ubiquitin Proteasome System ◽  
Loss Of Function ◽  
Interacting Protein ◽  
Human Cancer Cells ◽  
Relationship Analysis ◽  
New Strategy ◽  
Ubiquitin Proteasome

Molecular-glue degraders mediate interactions between target proteins and components of the ubiquitin-proteasome system to cause selective protein degradation. Here, we report a new molecular glue HQ461 discovered by high-throughput screening. Using loss-of-function and gain-of-function genetic screening in human cancer cells followed by biochemical reconstitution, we show that HQ461 acts by promoting an interaction between CDK12 and DDB1-CUL4-RBX1 E3 ubiquitin ligase, leading to polyubiquitination and degradation of CDK12-interacting protein Cyclin K (CCNK). Degradation of CCNK mediated by HQ461 compromised CDK12 function, leading to reduced phosphorylation of a CDK12 substrate, downregulation of DNA damage response genes, and cell death. Structure-activity relationship analysis of HQ461 revealed the importance of a 5-methylthiazol-2-amine pharmacophore and resulted in an HQ461 derivate with improved potency. Our studies reveal a new molecular glue that recruits its target protein directly to DDB1 to bypass the requirement of a substrate-specific receptor, presenting a new strategy for targeted protein degradation.

scholarly journals Discovery of a molecular glue promoting CDK12-DDB1 interaction to trigger Cyclin K degradation

2020 ◽  
Author(s):  
Lu Lv ◽  
Peihao Chen ◽  
Longzhi Cao ◽  
Yamei Li ◽  
Zhi Zeng ◽  
...  
Keyword(s):  
Protein Degradation ◽  
Small Molecules ◽  
High Throughput Screening ◽  
Ubiquitin Proteasome System ◽  
Great Promise ◽  
Loss Of Function ◽  
Gain Of Function ◽  
Interacting Protein ◽  
Relationship Analysis ◽  
New Strategy

AbstractMolecular glues are small molecules that exert their biologic or therapeutic activities by inducing gain-of-function interactions between pairs of proteins. In particular, molecular-glue degraders, which mediate interactions between target proteins and components of the ubiquitin proteasome system to cause targeted protein degradation, hold great promise as a unique modality for therapeutic targeting of proteins that are currently intractable. Here, we report a new molecular glue HQ461 discovered by high-throughput screening of small molecules that inhibited NRF2 activity. Using unbiased loss-of-function and gain-of-function genetic screening followed by biochemical reconstitution, we show that HQ461 acts by promoting interaction between CDK12 and DDB1-CUL4-RBX1 E3 ubiquitin ligase, leading to polyubiquitination and proteasomal degradation of CDK12’s interacting protein Cyclin K (CCNK). Degradation of CCNK mediated by HQ461 compromised CDK12 function, leading to reduced phosphorylation of CDK12 substrate, downregulation of DNA damage response genes, and cell death. Structure-activity relationship analysis of HQ461 revealed the importance of a 5-methylthiazol-2-amine pharmacophore and resulted in an HQ461 derivate with improved potency. Our studies reveal a new molecular glue that engages its target protein directly with DDB1 to bypass the requirement of a substrate-specific receptor, presenting a new strategy for targeted protein degradation.

scholarly journals Light-induced control of protein destruction by opto-PROTAC

2020 ◽  
Vol 6 (8) ◽  
pp. eaay5154 ◽  
Author(s):  
Jing Liu ◽  
He Chen ◽  
Leina Ma ◽  
Zhixiang He ◽  
Dong Wang ◽  
...  
Keyword(s):  
Protein Degradation ◽  
Precision Medicine ◽  
E3 Ligase ◽  
Ubiquitin Proteasome System ◽  
Specific Time ◽  
Ultraviolet A ◽  
Protein Targets ◽  
New Strategy ◽  
Ubiquitin Proteasome ◽  
Inducible Protein

By hijacking endogenous E3 ligase to degrade protein targets via the ubiquitin-proteasome system, PROTACs (PRoteolysis TArgeting Chimeras) provide a new strategy to inhibit protein targets that were regarded as undruggable before. However, the catalytic nature of PROTAC potentially leads to uncontrolled degradation that causes systemic toxicity issues, limiting the application of PROTAC in the clinic. Here, we introduce a light-inducible switch on PROTACs, thereafter termed as opto-PROTAC, to enable the degradation of protein targets in a spatiotemporal manner. By adding a photolabile caging group on pomalidomide as a parental compound and two additional PROTACs, dBET1 and dALK, we demonstrated light-inducible protein degradation. These opto-PROTACs display no activities in the dark, while the restricted degradation can be induced at a specific time and rate by ultraviolet A irradiation. Our approach provides a generalizable platform for the development of light-controlled PROTACs and enables PROTAC to be a precision medicine.

scholarly journals ATP13A2 Regulates Cellular α-Synuclein Multimerization, Membrane Association, and Externalization

2021 ◽  
Vol 22 (5) ◽  
pp. 2689
Author(s):  
Jianmin Si ◽  
Chris Van den Haute ◽  
Evy Lobbestael ◽  
Shaun Martin ◽  
Sarah van Veen ◽  
...  
Keyword(s):  
Membrane Integrity ◽  
Ubiquitin Proteasome System ◽  
Regulatory Function ◽  
Membrane Association ◽  
Loss Of Function ◽  
Atypical Form ◽  
Polyamine Transport ◽  
Independent Functions ◽  
Ubiquitin Proteasome ◽  
The Impact

ATP13A2, a late endo-/lysosomal polyamine transporter, is implicated in a variety of neurodegenerative diseases, including Parkinson’s disease and Kufor–Rakeb syndrome, an early-onset atypical form of parkinsonism. Loss-of-function mutations in ATP13A2 result in lysosomal deficiency as a consequence of impaired lysosomal export of the polyamines spermine/spermidine. Furthermore, accumulating evidence suggests the involvement of ATP13A2 in regulating the fate of α-synuclein, such as cytoplasmic accumulation and external release. However, no consensus has yet been reached on the mechanisms underlying these effects. Here, we aimed to gain more insight into how ATP13A2 is linked to α-synuclein biology in cell models with modified ATP13A2 activity. We found that loss of ATP13A2 impairs lysosomal membrane integrity and induces α-synuclein multimerization at the membrane, which is enhanced in conditions of oxidative stress or exposure to spermine. In contrast, overexpression of ATP13A2 wildtype (WT) had a protective effect on α-synuclein multimerization, which corresponded with reduced αsyn membrane association and stimulation of the ubiquitin-proteasome system. We also found that ATP13A2 promoted the secretion of α-synuclein through nanovesicles. Interestingly, the catalytically inactive ATP13A2 D508N mutant also affected polyubiquitination and externalization of α-synuclein multimers, suggesting a regulatory function independent of the ATPase and transport activity. In conclusion, our study demonstrates the impact of ATP13A2 on α-synuclein multimerization via polyamine transport dependent and independent functions.

scholarly journals Interaction between Parkin and α-Synuclein in PARK2-Mediated Parkinson’s Disease

Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 283
Author(s):  
Daniel Aghaie Madsen ◽  
Sissel Ida Schmidt ◽  
Morten Blaabjerg ◽  
Morten Meyer
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Ubiquitin Ligase ◽  
Current Knowledge ◽  
Lewy Bodies ◽  
Ubiquitin Proteasome System ◽  
Loss Of Function ◽  
Functional Interaction ◽  
Future Studies ◽  
Ubiquitin Proteasome

Parkin and α-synuclein are two key proteins involved in the pathophysiology of Parkinson’s disease (PD). Neurotoxic alterations of α-synuclein that lead to the formation of toxic oligomers and fibrils contribute to PD through synaptic dysfunction, mitochondrial impairment, defective endoplasmic reticulum and Golgi function, and nuclear dysfunction. In half of the cases, the recessively inherited early-onset PD is caused by loss of function mutations in the PARK2 gene that encodes the E3-ubiquitin ligase, parkin. Parkin is involved in the clearance of misfolded and aggregated proteins by the ubiquitin-proteasome system and regulates mitophagy and mitochondrial biogenesis. PARK2-related PD is generally thought not to be associated with Lewy body formation although it is a neuropathological hallmark of PD. In this review article, we provide an overview of post-mortem neuropathological examinations of PARK2 patients and present the current knowledge of a functional interaction between parkin and α-synuclein in the regulation of protein aggregates including Lewy bodies. Furthermore, we describe prevailing hypotheses about the formation of intracellular micro-aggregates (synuclein inclusions) that might be more likely than Lewy bodies to occur in PARK2-related PD. This information may inform future studies aiming to unveil primary signaling processes involved in PD and related neurodegenerative disorders.

scholarly journals Ccr4 is a novel shuttle factor required for ubiquitin-dependent proteolysis by the 26S proteasome

2020 ◽  
Author(s):  
Ganapathi Kandasamy ◽  
Ashis Kumar Pradhan ◽  
R Palanimurugan
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Degradation ◽  
Ubiquitin Proteasome System ◽  
Proteasomal Degradation ◽  
Rna Degradation ◽  
26S Proteasome ◽  
Cellular Proteins ◽  
Cellular Homeostasis ◽  
Substrate Degradation ◽  
Ubiquitin Proteasome

AbstractDegradation of short-lived and abnormal proteins are essential for normal cellular homeostasis. In eukaryotes, such unstable cellular proteins are selectively degraded by the ubiquitin proteasome system (UPS). Furthermore, abnormalities in protein degradation by the UPS have been linked to several human diseases. Ccr4 protein is a known component of the Ccr4-Not complex, which has established roles in transcription, mRNA de-adenylation and RNA degradation etc. Excitingly in this study, we show that Ccr4 protein has a novel function as a shuttle factor that promotes ubiquitin-dependent degradation of short-lived proteins by the 26S proteasome. Using a substrate of the well-studied ubiquitin fusion degradation (UFD) pathway, we found that its UPS-mediated degradation was severely impaired upon deletion of CCR4 in Saccharomyces cerevisiae. Additionally, we show that Ccr4 binds to cellular ubiquitin conjugates and the proteasome. In contrast to Ccr4, most other subunits of the Ccr4-Not complex proteins are dispensable for UFD substrate degradation. From our findings we conclude that Ccr4 functions in the UPS as a shuttle factor targeting ubiquitylated substrates for proteasomal degradation.

scholarly journals E2A protein degradation by the ubiquitin-proteasome system is stage-dependent during muscle differentiation

Oncogene ◽  
2006 ◽  
Vol 26 (3) ◽  
pp. 441-448 ◽  
Author(s):  
L Sun ◽  
J S Trausch-Azar ◽  
A Ciechanover ◽  
A L Schwartz
Keyword(s):  
Protein Degradation ◽  
Ubiquitin Proteasome System ◽  
Muscle Differentiation ◽  
Ubiquitin Proteasome

IRF6 and TAK1 coordinately promote the activation of HIPK2 to stimulate apoptosis during palate fusion

2019 ◽  
Vol 12 (593) ◽  
pp. eaav7666 ◽  
Author(s):  
Chen-Yeh Ke ◽  
Hua-Hsuan Mei ◽  
Fen-Hwa Wong ◽  
Lun-Jou Lo
Keyword(s):  
Transcription Factor ◽  
Cell Apoptosis ◽  
Transforming Growth Factor ◽  
Kinase Inhibitor ◽  
Ex Vivo ◽  
Human Cancer ◽  
Ectopic Expression ◽  
Interacting Protein ◽  
Human Cancer Cells

Cleft palate is a common craniofacial defect caused by a failure in palate fusion. The palatal shelves migrate toward one another and meet at the embryonic midline, creating a seam. Transforming growth factor–β3 (TGF-β3)–induced apoptosis of the medial edge epithelium (MEE), the cells located along the seam, is required for completion of palate fusion. The transcription factor interferon regulatory factor 6 (IRF6) promotes TGF-β3–induced MEE cell apoptosis by stimulating the degradation of the transcription factor ΔNp63 and promoting the expression of the gene encoding the cyclin-dependent kinase inhibitor p21. Because homeodomain-interacting protein kinase 2 (HIPK2) functions downstream of IRF6 in human cancer cells and is required for ΔNp63 protein degradation in keratinocytes, we investigated whether HIPK2 played a role in IRF6-induced ΔNp63 degradation in palate fusion. HIPK2 was present in the MEE cells of mouse palatal shelves during seam formation in vivo, and ectopic expression of IRF6 in palatal shelves cultured ex vivo stimulated the expression of Hipk2 and the accumulation of phosphorylated HIPK2. Knockdown and ectopic expression experiments in organ culture demonstrated that p21 was required for HIPK2- and IRF6-dependent activation of caspase 3, MEE apoptosis, and palate fusion. Contact between palatal shelves enhanced the phosphorylation of TGF-β–activated kinase 1 (TAK1), which promoted the phosphorylation of HIPK2 and palate fusion. Our findings demonstrate that HIPK2 promotes seam cell apoptosis and palate fusion downstream of IRF6 and that IRF6 and TAK1 appear to coordinately enhance the abundance and activation of HIPK2 during palate fusion.

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