Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific Functions
ATR is a PI3K-like kinase protein, regulating checkpoint responses to DNA damage and replication stress. Apart from its checkpoint function in the nucleus, ATR actively engages in an antiapoptotic role at mitochondria following DNA damage. The different functions of ATR in the nucleus and cytoplasm...
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Frontiers Media S.A.
2022-06-01
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| Series: | Frontiers in Cell and Developmental Biology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fcell.2022.826576/full |
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| author | Himadri Biswas Shu-Jun Zhao Shu-Jun Zhao Yetunde Makinwa James S. Bassett Phillip R. Musich Jing-Yuan Liu Jing-Yuan Liu Yue Zou |
| author_facet | Himadri Biswas Shu-Jun Zhao Shu-Jun Zhao Yetunde Makinwa James S. Bassett Phillip R. Musich Jing-Yuan Liu Jing-Yuan Liu Yue Zou |
| author_sort | Himadri Biswas |
| collection | DOAJ |
| description | ATR is a PI3K-like kinase protein, regulating checkpoint responses to DNA damage and replication stress. Apart from its checkpoint function in the nucleus, ATR actively engages in an antiapoptotic role at mitochondria following DNA damage. The different functions of ATR in the nucleus and cytoplasm are carried out by two prolyl isomeric forms of ATR: trans- and cis-ATR, respectively. The isomerization occurs at the Pin1 Ser428-Pro429 motif of ATR. Here, we investigated the structural basis of the subcellular location-specific functions of human ATR. Using a mass spectrometry-based footprinting approach, the surface accessibility of ATR lysine residues to sulfo-NHS-LC-biotin modification was monitored and compared between the cis- and the trans-isomers. We have identified two biotin-modified lysine residues, K459 and K469, within the BH3-like domain of cis-ATR that were not accessible in trans-ATR, indicating a conformational change around the BH3 domain between cis- and trans-ATR. The conformational alteration also involved the N-terminal domain and the middle HEAT domain. Moreover, experimental results from an array of complementary assays show that cis-ATR with the accessible BH3 domain was able to bind to tBid while trans-ATR could not. In addition, both cis- and trans-ATR can directly form homodimers via their C-terminal domains without ATRIP, while nuclear (trans-ATR) in the presence of ATRIP forms dimer–dimer complexes involving both N- and C-termini of ATR and ATRIP after UV. Structural characteristics around the Ser428-Pro429 motif and the BH3 domain region are also analyzed by molecular modeling and dynamics simulation. In support, cis conformation was found to be significantly more energetically favorable than trans at the Ser428-Pro429 bond in a 20-aa wild-type ATR peptide. Taken together, our results suggest that the isomerization-induced structural changes of ATR define both its subcellular location and compartment-specific functions and play an essential role in promoting cell survival and DNA damage responses. |
| first_indexed | 2024-04-12T17:31:46Z |
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| id | doaj.art-1e74f91378c245a0a0d73db857a2765c |
| institution | Directory Open Access Journal |
| issn | 2296-634X |
| language | English |
| last_indexed | 2024-04-12T17:31:46Z |
| publishDate | 2022-06-01 |
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| series | Frontiers in Cell and Developmental Biology |
| spelling | doaj.art-1e74f91378c245a0a0d73db857a2765c2022-12-22T03:23:07ZengFrontiers Media S.A.Frontiers in Cell and Developmental Biology2296-634X2022-06-011010.3389/fcell.2022.826576826576Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific FunctionsHimadri Biswas0Shu-Jun Zhao1Shu-Jun Zhao2Yetunde Makinwa3James S. Bassett4Phillip R. Musich5Jing-Yuan Liu6Jing-Yuan Liu7Yue Zou8Department of Cancer Biology, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United StatesDepartment of Medicine, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United StatesDepartment of Bioengineering, University of Toledo College of Engineering, Toledo, OH, United StatesDepartment of Cancer Biology, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United StatesDepartment of Medicine, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United StatesDepartment of Biomedical Sciences, Quillen College of Medicine, East Tennessee State University, Johnson City, TN, United StatesDepartment of Medicine, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United StatesDepartment of Bioengineering, University of Toledo College of Engineering, Toledo, OH, United StatesDepartment of Cancer Biology, University of Toledo College of Medicine and Life Sciences, Toledo, OH, United StatesATR is a PI3K-like kinase protein, regulating checkpoint responses to DNA damage and replication stress. Apart from its checkpoint function in the nucleus, ATR actively engages in an antiapoptotic role at mitochondria following DNA damage. The different functions of ATR in the nucleus and cytoplasm are carried out by two prolyl isomeric forms of ATR: trans- and cis-ATR, respectively. The isomerization occurs at the Pin1 Ser428-Pro429 motif of ATR. Here, we investigated the structural basis of the subcellular location-specific functions of human ATR. Using a mass spectrometry-based footprinting approach, the surface accessibility of ATR lysine residues to sulfo-NHS-LC-biotin modification was monitored and compared between the cis- and the trans-isomers. We have identified two biotin-modified lysine residues, K459 and K469, within the BH3-like domain of cis-ATR that were not accessible in trans-ATR, indicating a conformational change around the BH3 domain between cis- and trans-ATR. The conformational alteration also involved the N-terminal domain and the middle HEAT domain. Moreover, experimental results from an array of complementary assays show that cis-ATR with the accessible BH3 domain was able to bind to tBid while trans-ATR could not. In addition, both cis- and trans-ATR can directly form homodimers via their C-terminal domains without ATRIP, while nuclear (trans-ATR) in the presence of ATRIP forms dimer–dimer complexes involving both N- and C-termini of ATR and ATRIP after UV. Structural characteristics around the Ser428-Pro429 motif and the BH3 domain region are also analyzed by molecular modeling and dynamics simulation. In support, cis conformation was found to be significantly more energetically favorable than trans at the Ser428-Pro429 bond in a 20-aa wild-type ATR peptide. Taken together, our results suggest that the isomerization-induced structural changes of ATR define both its subcellular location and compartment-specific functions and play an essential role in promoting cell survival and DNA damage responses.https://www.frontiersin.org/articles/10.3389/fcell.2022.826576/fullATRcis/trans prolyl isomerizationmass spectrometric protein footprintingstructure-function of ATRmitochondrial ATR-tBid interactionantiapoptosis |
| spellingShingle | Himadri Biswas Shu-Jun Zhao Shu-Jun Zhao Yetunde Makinwa James S. Bassett Phillip R. Musich Jing-Yuan Liu Jing-Yuan Liu Yue Zou Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific Functions Frontiers in Cell and Developmental Biology ATR cis/trans prolyl isomerization mass spectrometric protein footprinting structure-function of ATR mitochondrial ATR-tBid interaction antiapoptosis |
| title | Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific Functions |
| title_full | Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific Functions |
| title_fullStr | Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific Functions |
| title_full_unstemmed | Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific Functions |
| title_short | Prolyl Isomerization-Mediated Conformational Changes Define ATR Subcellular Compartment-Specific Functions |
| title_sort | prolyl isomerization mediated conformational changes define atr subcellular compartment specific functions |
| topic | ATR cis/trans prolyl isomerization mass spectrometric protein footprinting structure-function of ATR mitochondrial ATR-tBid interaction antiapoptosis |
| url | https://www.frontiersin.org/articles/10.3389/fcell.2022.826576/full |
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