scholarly journals DELTEX2 C-terminal domain recognizes and recruits ADP-ribosylated proteins for ubiquitination

2020 ◽  
Vol 6 (34) ◽  
pp. eabc0629
Author(s):  
Syed Feroj Ahmed ◽  
Lori Buetow ◽  
Mads Gabrielsen ◽  
Sergio Lilla ◽  
Chatrin Chatrin ◽  
...  
Keyword(s):  
Binding Pocket ◽  
Binding Property ◽  
Interacting Proteins ◽  
Label Free ◽  
Repair Pathway ◽  
Adp Ribosylation ◽  
Terminal Domain ◽  
Damage Repair ◽  
Key Players ◽  
Modified Proteins

Cross-talk between ubiquitination and ADP-ribosylation regulates spatiotemporal recruitment of key players in many signaling pathways. The DELTEX family ubiquitin ligases (DTX1 to DTX4 and DTX3L) are characterized by a RING domain followed by a C-terminal domain (DTC) of hitherto unknown function. Here, we use two label-free mass spectrometry techniques to investigate the interactome and ubiquitinated substrates of human DTX2 and identify a large proportion of proteins associated with the DNA damage repair pathway. We show that DTX2-catalyzed ubiquitination of these interacting proteins requires PARP1/2-mediated ADP-ribosylation and depends on the DTC domain. Using a combination of structural, biochemical, and cell-based techniques, we show that the DTX2 DTC domain harbors an ADP-ribose–binding pocket and recruits poly-ADP-ribose (PAR)–modified proteins for ubiquitination. This PAR-binding property of DTC domain is conserved across the DELTEX family E3s. These findings uncover a new ADP-ribose–binding domain that facilitates PAR-dependent ubiquitination.

scholarly journals Alteration of protein expression and spliceosome pathway activity during Barrett’s carcinogenesis

2021 ◽  
Author(s):  
Christoph Stingl ◽  
Angela Bureo Gonzalez ◽  
Coşkun Güzel ◽  
Kai Yi Nadine Phoa ◽  
Michail Doukas ◽  
...  
Keyword(s):  
Micro Rna ◽  
Differentially Expressed ◽  
Epithelial Tissue ◽  
Label Free ◽  
Tissue Samples ◽  
Damage Repair ◽  
Inter Observer Variability ◽  
Stromal Tissue ◽  
The Face ◽  
Abundance And Diversity

Abstract Background Barrett’s esophagus (BE) is a known precursor lesion and the strongest risk factor for esophageal adenocarcinoma (EAC), a common and lethal type of cancer. Prediction of risk, the basis for efficient intervention, is commonly solely based on histologic examination. This approach is challenged by problems such as inter-observer variability in the face of the high heterogeneity of dysplastic tissue. Molecular markers might offer an additional way to understand the carcinogenesis and improve the diagnosis—and eventually treatment. In this study, we probed significant proteomic changes during dysplastic progression from BE into EAC. Methods During endoscopic mucosa resection, epithelial and stromal tissue samples were collected by laser capture microdissection from 10 patients with normal BE and 13 patients with high-grade dysplastic/EAC. Samples were analyzed by mass spectrometry-based proteomic analysis. Expressed proteins were determined by label-free quantitation, and gene set enrichment was used to find differentially expressed pathways. The results were validated by immunohistochemistry for two selected key proteins (MSH6 and XPO5). Results Comparing dysplastic/EAC to non-dysplastic BE, we found in equal volumes of epithelial tissue an overall up-regulation in terms of protein abundance and diversity, and determined a set of 226 differentially expressed proteins. Significantly higher expressions of MSH6 and XPO5 were validated orthogonally and confirmed by immunohistochemistry. Conclusions Our results demonstrate that disease-related proteomic alterations can be determined by analyzing minute amounts of cell-type-specific collected tissue. Further analysis indicated that alterations of certain pathways associated with carcinogenesis, such as micro-RNA trafficking, DNA damage repair, and spliceosome activity, exist in dysplastic/EAC.

scholarly journals Determinants of the Endosomal Localization of Sorting Nexin 1

2005 ◽  
Vol 16 (4) ◽  
pp. 2049-2057 ◽  
Author(s):  
Qi Zhong ◽  
Martin J. Watson ◽  
Cheri S. Lazar ◽  
Andrea M. Hounslow ◽  
Jonathan P. Waltho ◽  
...  
Keyword(s):  
Binding Pocket ◽  
Early Endosome ◽  
Nmr Structure ◽  
Sorting Nexin ◽  
High Affinity ◽  
Terminal Domain ◽  
Px Domain ◽  
Phox Homology ◽  
Phosphatidylinositol 3

The sorting nexin (SNX) family of proteins is characterized by sequence-related phox homology (PX) domains. A minority of PX domains bind with high affinity to phosphatidylinositol 3-phosphate [PI(3)P], whereas the majority of PX domains exhibit low affinity that is insufficient to target them to vesicles. SNX1 is located on endosomes, but its low affinity PX domain fails to localize in vivo. The NMR structure of the PX domain of SNX1 reveals an overall fold that is similar to high-affinity PX domains. However, the phosphatidylinositol (PI) binding pocket of the SNX1 PX domain is incomplete; regions of the pocket that are well defined in high-affinity PX domains are highly mobile in SNX1. Some of this mobility is lost upon binding PI(3)P. The C-terminal domain of SNX1 is a long helical dimer that localizes to vesicles but not to the early endosome antigen-1–containing vesicles where endogenous SNX1 resides. Thus, the obligate dimerization of SNX1 that is driven by the C-terminal domain creates a high-affinity PI binding species that properly targets the holo protein to endosomes.

scholarly journals The Fanconi anemia DNA damage repair pathway in the spotlight for germline predisposition to colorectal cancer

2016 ◽  
Vol 24 (10) ◽  
pp. 1501-1505 ◽  
Author(s):  
Clara Esteban-Jurado ◽  
◽  
Sebastià Franch-Expósito ◽  
Jenifer Muñoz ◽  
Teresa Ocaña ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Dna Damage ◽  
Fanconi Anemia ◽  
Dna Damage Repair ◽  
Repair Pathway ◽  
Damage Repair

scholarly journals MicroRNA-146a-5p enhances radiosensitivity in hepatocellular carcinoma through replication protein A3-induced activation of the DNA repair pathway

2019 ◽  
Vol 316 (3) ◽  
pp. C299-C311 ◽  
Author(s):  
Jing Luo ◽  
Zhong-Zhou Si ◽  
Ting Li ◽  
Jie-Qun Li ◽  
Zhong-Qiang Zhang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Proliferation ◽  
Dna Damage ◽  
Dna Repair ◽  
Dna Damage Repair ◽  
Replication Protein ◽  
Repair Pathway ◽  
Related Factors ◽  
Damage Repair ◽  
Hcc Cells

Hepatocellular carcinoma (HCC) is known for its high mortality rate worldwide. Based on intensive studies, microRNA (miRNA) expression functions in tumor suppression. Therefore, we aimed to evaluate the contribution of miR-146a-5p to radiosensitivity in HCC through the activation of the DNA damage repair pathway by binding to replication protein A3 (RPA3). First, the limma package of R was performed to differentially analyze HCC expression chip, and regulative miRNA of RPA3 was predicted. Expression of miR-146a-5p, RPA3, and DNA damage repair pathway-related factors in tissues and cells was determined. The effects of radiotherapy on the expression of miR-146a-5p and RPA3 as well as on cell radiosensitivity, proliferation, cell cycle, and apoptosis were also assessed. The results showed that there exists a close correlation between miR-146a and the radiotherapy effect on HCC progression through regulation of RPA3 and the DNA repair pathway. The positive rate of ATM, pCHK2, and Rad51 in HCC tissues was higher when compared with that of the paracancerous tissues. SMMC-7721 and HepG2 cell proliferation were significantly inhibited following 8 Gy 6Mv dose. MiR-146a-5p restrained the expression of RPA3 and promoted the expression of relative genes associated with the DNA repair pathway. In addition, miR-146a-5p overexpression suppresses cell proliferation and enhances radiosensitivity and cell apoptosis in HCC cells. In conclusion, the present study revealed that miR-146a-5p could lead to the restriction of proliferation and the promotion of radiosensitivity and apoptosis in HCC cells through activation of DNA repair pathway and inhibition of RPA3.

scholarly journals Characterization of the ATPase activity of topoisomerase II from Leishmania donovani and identification of residues conferring resistance to etoposide

2005 ◽  
Vol 390 (2) ◽  
pp. 419-426 ◽  
Author(s):  
Tanushri Sengupta ◽  
Mandira Mukherjee ◽  
Aditi Das ◽  
Chhabinath Mandal ◽  
Rakhee Das ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Topoisomerase Ii ◽  
Leishmania Donovani ◽  
Binding Pocket ◽  
Dna Topoisomerase ◽  
Terminal Domain ◽  
Specific Selectivity ◽  
Eukaryotic Dna ◽  
First Time

We have cloned and expressed the 43 kDa N-terminal domain of Leishmania donovani topoisomerase II. This protein has an intrinsic ATPase activity and obeys Michaelis–Menten kinetics. Cross-linking studies indicate that the N-terminal domain exists as a dimer both in the presence and absence of nucleotides. Etoposide, an effective antitumour drug, traps eukaryotic DNA topoisomerase II in a covalent complex with DNA. In the present study, we report for the first time that etoposide inhibits the ATPase activity of the recombinant N-terminal domain of L. donovani topoisomerase II. We have modelled the structure of this 43 kDa protein and performed molecular docking analysis with the drug. Mutagenesis of critical amino acids in the vicinity of the ligand-binding pocket reveals less efficient inhibition of the ATPase activity of the enzyme by etoposide. Taken together, these results provide an insight for the development of newer therapeutic agents with specific selectivity.

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