RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcanii
ABSTRACT Nucleases are strictly regulated and often localized in the cell to avoid the uncontrolled degradation of DNA and RNA. Here, a new type of nuclease complex, composed of RecJ3, RecJ4, and aRNase J, was identified through its ATP-dependent association with the ubiquitin-like SAMP1 and AAA-ATP...
| Main Authors: | , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
American Society for Microbiology
2023-08-01
|
| Series: | mBio |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/mbio.00852-23 |
| _version_ | 1797730243510796288 |
|---|---|
| author | Huiyong Jia Swathi Dantuluri Shae Margulies Victoria Smith Rebecca Lever Thorsten Allers Jin Koh Sixue Chen Julie A. Maupin-Furlow |
| author_facet | Huiyong Jia Swathi Dantuluri Shae Margulies Victoria Smith Rebecca Lever Thorsten Allers Jin Koh Sixue Chen Julie A. Maupin-Furlow |
| author_sort | Huiyong Jia |
| collection | DOAJ |
| description | ABSTRACT Nucleases are strictly regulated and often localized in the cell to avoid the uncontrolled degradation of DNA and RNA. Here, a new type of nuclease complex, composed of RecJ3, RecJ4, and aRNase J, was identified through its ATP-dependent association with the ubiquitin-like SAMP1 and AAA-ATPase Cdc48a. The complex was discovered in Haloferax volcanii, an archaeon lacking an RNA exosome. Genetic analysis revealed aRNase J to be essential and RecJ3, RecJ4, and Cdc48a to function in the recovery from DNA damage including genotoxic agents that generate double-strand breaks. The RecJ3:RecJ4:aRNase J complex (isolated in 2:2:1 stoichiometry) functioned primarily as a 3′−5′ exonuclease in hydrolyzing RNA and ssDNA, with the mechanism non-processive for ssDNA. aRNase J could also be purified as a homodimer that catalyzed endoribonuclease activity and, thus, was not restricted to the 5′−3′ exonuclease activity typical of aRNase J homologs. Moreover, RecJ3 and RecJ4 could be purified as a 560-kDa subcomplex in equimolar subunit ratio with nuclease activities mirroring the full RecJ3/4-aRNase J complex. These findings prompted reconstitution assays that suggested RecJ3/4 could suppress, alter, and/or outcompete the nuclease activities of aRNase J. Based on the phenotypic results, this control mechanism of aRNase J by RecJ3/4 is not necessary for cell growth but instead appears important for DNA repair. IMPORTANCE Nucleases are critical for various cellular processes including DNA replication and repair. Here, a dynamic type of nuclease complex is newly identified in the archaeon Haloferax volcanii, which is missing the canonical RNA exosome. The complex, composed of RecJ3, RecJ4, and aRNase J, functions primarily as a 3′–5′ exonuclease and was discovered through its ATP-dependent association with the ubiquitin-like SAMP1 and Cdc48a. aRNase J alone forms a homodimer that has endonuclease function and, thus, is not restricted to 5′–3′ exonuclease activity typical of other aRNase J enzymes. RecJ3/4 appears to suppress, alter, and/or outcompete the nuclease activities of aRNase J. While aRNase J is essential for growth, RecJ3/4, Cdc48a, and SAMPs are important for recovery against DNA damage. These biological distinctions may correlate with the regulated nuclease activity of aRNase J in the RecJ3/4-aRNaseJ complex. |
| first_indexed | 2024-03-12T11:41:27Z |
| format | Article |
| id | doaj.art-3fe972fd54944158bffd9fd1aefa96f5 |
| institution | Directory Open Access Journal |
| issn | 2150-7511 |
| language | English |
| last_indexed | 2024-03-12T11:41:27Z |
| publishDate | 2023-08-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mBio |
| spelling | doaj.art-3fe972fd54944158bffd9fd1aefa96f52023-08-31T15:04:20ZengAmerican Society for MicrobiologymBio2150-75112023-08-0114410.1128/mbio.00852-23RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcaniiHuiyong Jia0Swathi Dantuluri1Shae Margulies2Victoria Smith3Rebecca Lever4Thorsten Allers5Jin Koh6Sixue Chen7Julie A. Maupin-Furlow8Department of Microbiology and Cell Science, Institute of Food and Agricultural Science, University of Florida , Gainesville, Florida, USADepartment of Microbiology and Cell Science, Institute of Food and Agricultural Science, University of Florida , Gainesville, Florida, USADepartment of Microbiology and Cell Science, Institute of Food and Agricultural Science, University of Florida , Gainesville, Florida, USASchool of Life Sciences, University of Nottingham , Nottingham, United KingdomSchool of Life Sciences, University of Nottingham , Nottingham, United KingdomSchool of Life Sciences, University of Nottingham , Nottingham, United KingdomProteomics and Mass Spectrometry, Interdisciplinary Center for Biotechnology Research, University of Florida , Gainesville, Florida, USAProteomics and Mass Spectrometry, Interdisciplinary Center for Biotechnology Research, University of Florida , Gainesville, Florida, USADepartment of Microbiology and Cell Science, Institute of Food and Agricultural Science, University of Florida , Gainesville, Florida, USAABSTRACT Nucleases are strictly regulated and often localized in the cell to avoid the uncontrolled degradation of DNA and RNA. Here, a new type of nuclease complex, composed of RecJ3, RecJ4, and aRNase J, was identified through its ATP-dependent association with the ubiquitin-like SAMP1 and AAA-ATPase Cdc48a. The complex was discovered in Haloferax volcanii, an archaeon lacking an RNA exosome. Genetic analysis revealed aRNase J to be essential and RecJ3, RecJ4, and Cdc48a to function in the recovery from DNA damage including genotoxic agents that generate double-strand breaks. The RecJ3:RecJ4:aRNase J complex (isolated in 2:2:1 stoichiometry) functioned primarily as a 3′−5′ exonuclease in hydrolyzing RNA and ssDNA, with the mechanism non-processive for ssDNA. aRNase J could also be purified as a homodimer that catalyzed endoribonuclease activity and, thus, was not restricted to the 5′−3′ exonuclease activity typical of aRNase J homologs. Moreover, RecJ3 and RecJ4 could be purified as a 560-kDa subcomplex in equimolar subunit ratio with nuclease activities mirroring the full RecJ3/4-aRNase J complex. These findings prompted reconstitution assays that suggested RecJ3/4 could suppress, alter, and/or outcompete the nuclease activities of aRNase J. Based on the phenotypic results, this control mechanism of aRNase J by RecJ3/4 is not necessary for cell growth but instead appears important for DNA repair. IMPORTANCE Nucleases are critical for various cellular processes including DNA replication and repair. Here, a dynamic type of nuclease complex is newly identified in the archaeon Haloferax volcanii, which is missing the canonical RNA exosome. The complex, composed of RecJ3, RecJ4, and aRNase J, functions primarily as a 3′–5′ exonuclease and was discovered through its ATP-dependent association with the ubiquitin-like SAMP1 and Cdc48a. aRNase J alone forms a homodimer that has endonuclease function and, thus, is not restricted to 5′–3′ exonuclease activity typical of other aRNase J enzymes. RecJ3/4 appears to suppress, alter, and/or outcompete the nuclease activities of aRNase J. While aRNase J is essential for growth, RecJ3/4, Cdc48a, and SAMPs are important for recovery against DNA damage. These biological distinctions may correlate with the regulated nuclease activity of aRNase J in the RecJ3/4-aRNaseJ complex.https://journals.asm.org/doi/10.1128/mbio.00852-23post-translational modificationubiquitinproteasomesDNA repairarchaea |
| spellingShingle | Huiyong Jia Swathi Dantuluri Shae Margulies Victoria Smith Rebecca Lever Thorsten Allers Jin Koh Sixue Chen Julie A. Maupin-Furlow RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcanii mBio post-translational modification ubiquitin proteasomes DNA repair archaea |
| title | RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcanii |
| title_full | RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcanii |
| title_fullStr | RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcanii |
| title_full_unstemmed | RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcanii |
| title_short | RecJ3/4-aRNase J form a Ubl-associated nuclease complex functioning in survival against DNA damage in Haloferax volcanii |
| title_sort | recj3 4 arnase j form a ubl associated nuclease complex functioning in survival against dna damage in haloferax volcanii |
| topic | post-translational modification ubiquitin proteasomes DNA repair archaea |
| url | https://journals.asm.org/doi/10.1128/mbio.00852-23 |
| work_keys_str_mv | AT huiyongjia recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT swathidantuluri recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT shaemargulies recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT victoriasmith recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT rebeccalever recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT thorstenallers recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT jinkoh recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT sixuechen recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii AT julieamaupinfurlow recj34arnasejformaublassociatednucleasecomplexfunctioninginsurvivalagainstdnadamageinhaloferaxvolcanii |