{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,10]],"date-time":"2025-11-10T14:01:48Z","timestamp":1762783308816,"version":"build-2065373602"},"reference-count":49,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2025,7,9]],"date-time":"2025-07-09T00:00:00Z","timestamp":1752019200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Microorganisms"],"abstract":"<jats:p>Neisseria gonorrhoeae is the causative agent of the sexually transmitted infection gonorrhea. Preventative vaccines or novel treatments based on a better understanding of the molecular basis of N. gonorrhoeae infection are required as resistance to current antibiotics is widespread. Toxin\u2013antitoxin (TA) systems modulate bacterial physiology by interfering with vital cellular processes; type II TA systems, where both toxin and antitoxin are proteins, are the best-studied. Bioinformatics analysis revealed genes encoding an uncharacterized type II HicAB TA system in the N. gonorrhoeae strain FA1090 chromosome, which were also present in &gt;83% of the other gonococcal genome sequences examined. Gonococcal HicA overproduction inhibited bacterial growth in Escherichia coli, an effect that could be counteracted by the co-expression of HicB. Kill\/rescue assays showed that this effect was bacteriostatic rather than bactericidal. The site-directed mutagenesis of key histidine and glycine residues (Gly22, His24, His29) abolished HicA-mediated growth arrest. N. gonorrhoeae FA1090\u2206hicAB and complemented derivatives that expressed IPTG-inducible hicA, hicB, or hicAB, respectively, grew as wild type, except for IPTG-induced FA1090\u2206hicAB::hicA. RT-PCR demonstrated that hicAB are transcribed in vitro under the culture conditions used. The deletion of hicAB had no effect on biofilm formation. Our study describes the first characterization of a HicAB TA system in N. gonorrhoeae.<\/jats:p>","DOI":"10.3390\/microorganisms13071619","type":"journal-article","created":{"date-parts":[[2025,7,10]],"date-time":"2025-07-10T07:38:27Z","timestamp":1752133107000},"page":"1619","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["The Characterization of a Gonococcal HicAB Toxin\u2013Antitoxin System Capable of Causing Bacteriostatic Growth Arrest"],"prefix":"10.3390","volume":"13","author":[{"given":"Salwa S.","family":"Bagabas","sequence":"first","affiliation":[{"name":"Department of Biological Sciences, Faculty of Science, University of Jeddah, Jeddah 23445, Saudi Arabia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0004-4438-1957","authenticated-orcid":false,"given":"Jorge","family":"Trujillo-Mendoza","sequence":"additional","affiliation":[{"name":"School of Life Sciences, University of Nottingham, Nottingham NG7 2RD, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3046-137X","authenticated-orcid":false,"given":"Michael J.","family":"Stocks","sequence":"additional","affiliation":[{"name":"School of Pharmacy, University of Nottingham, Nottingham NG7 2RD, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"David P. J.","family":"Turner","sequence":"additional","affiliation":[{"name":"School of Life Sciences, University of Nottingham, Nottingham NG7 2RD, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7423-8378","authenticated-orcid":false,"given":"Neil J.","family":"Oldfield","sequence":"additional","affiliation":[{"name":"School of Life Sciences, University of Nottingham, Nottingham NG7 2RD, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,7,9]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"401","DOI":"10.1071\/SH19061","article-title":"Epidemiology of gonorrhoea: A global perspective","volume":"16","author":"Kirkcaldy","year":"2019","journal-title":"Sex. Health"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"79","DOI":"10.1038\/s41572-019-0128-6","article-title":"Gonorrhoea","volume":"5","author":"Unemo","year":"2019","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"258","DOI":"10.1053\/j.spid.2005.06.006","article-title":"Gonococcal infections in neonates and young children","volume":"16","author":"Woods","year":"2005","journal-title":"Semin. Pediatr. Infect. Dis."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"554","DOI":"10.1097\/OLQ.0000000000000340","article-title":"Chlamydia and gonorrhea in HIV-infected pregnant women and infant HIV transmission","volume":"42","author":"Adachi","year":"2015","journal-title":"Sex. Transm. Dis."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"e532","DOI":"10.1016\/S1473-3099(24)00001-X","article-title":"Managing treatment failure in Neisseria gonorrhoeae infection: Current guidelines and future directions","volume":"24","author":"Fifer","year":"2024","journal-title":"Lancet Infect. Dis."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Raccagni, A.R., Ranzenigo, M., Bruzzesi, E., Maci, C., Castagna, A., and Nozza, S. (2023). Neisseria gonorrhoeae antimicrobial resistance: The future of antibiotic therapy. J. Clin. Med., 12.","DOI":"10.3390\/jcm12247767"},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Qiu, J., Zhai, Y., Wei, M., Zheng, C., and Jiao, X. (2022). Toxin-antitoxin systems: Classification, biological roles, and applications. Microbiol. Res., 264.","DOI":"10.1016\/j.micres.2022.127159"},{"key":"ref_8","first-page":"100204","article-title":"Bacterial toxin-antitoxin systems: Novel insights on toxin activation across populations and experimental shortcomings","volume":"5","author":"Spira","year":"2023","journal-title":"Curr. Res. Microb. Sci."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"e14220","DOI":"10.1016\/j.heliyon.2023.e14220","article-title":"Toxin-antitoxin systems in bacterial pathogenesis","volume":"9","author":"Sonika","year":"2023","journal-title":"Heliyon"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Kamruzzaman, M., Wu, A.Y., and Iredell, J.R. (2021). Biological functions of type II toxin-antitoxin systems in bacteria. Microorganisms, 9.","DOI":"10.3390\/microorganisms9061276"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"335","DOI":"10.1038\/s41579-021-00661-1","article-title":"Biology and evolution of bacterial toxin-antitoxin systems","volume":"20","author":"Fraikin","year":"2022","journal-title":"Nat. Rev. Microbiol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"5513","DOI":"10.1093\/nar\/gkr131","article-title":"Diversity of bacterial type II toxin-antitoxin systems: A comprehensive search and functional analysis of novel families","volume":"39","author":"Leplae","year":"2011","journal-title":"Nucleic Acids Res."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"68","DOI":"10.1007\/s41048-020-00109-8","article-title":"Type II toxin\u2013antitoxin system in bacteria: Activation, function, and mode of action","volume":"6","author":"Zhang","year":"2020","journal-title":"Biophys. Rep."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"996","DOI":"10.1016\/j.str.2010.04.018","article-title":"Crystal structures of Phd-Doc, HigA, and YeeU establish multiple evolutionary links between microbial growth-regulating toxin-antitoxin systems","volume":"18","author":"Arbing","year":"2010","journal-title":"Structure"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"283","DOI":"10.4161\/mge.18477","article-title":"Bacterial toxin-antitoxin systems: Translation inhibitors everywhere","volume":"1","author":"Guglielmini","year":"2011","journal-title":"Mob. Genet. Elem."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"2581","DOI":"10.1093\/bioinformatics\/btl418","article-title":"The HicAB cassette, a putative novel, RNA-targeting toxin-antitoxin system in archaea and bacteria","volume":"22","author":"Makarova","year":"2006","journal-title":"Bioinformatics"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1191","DOI":"10.1128\/JB.01013-08","article-title":"HicA of Escherichia coli defines a novel family of translation-independent mRNA interferases in bacteria and archaea","volume":"191","author":"Pandey","year":"2009","journal-title":"J. Bacteriol."},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Encina-Robles, J., Perez-Villalobos, V., and Bustamante, P. (2024). The HicAB system: Characteristics and biological roles of an underappreciated toxin-antitoxin system. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms252212165"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Hou, B., Wang, C.-Y., Li, S.-W., Zhou, L.-J., Che, Y.-L., and Chen, Q.-Y. (2023). Effects of toxin-antitoxin system HicAB on biofilm formation by extraintestinal pathogenic E. coli. Curr. Microbiol., 80.","DOI":"10.1007\/s00284-022-03138-3"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"86","DOI":"10.1111\/1574-6968.12032","article-title":"Identification of type II toxin\u2013antitoxin modules in Burkholderia pseudomallei","volume":"338","author":"Butt","year":"2013","journal-title":"FEMS Microbiol. Lett."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"333","DOI":"10.1042\/BJ20140073","article-title":"The HicA toxin from Burkholderia pseudomallei has a role in persister cell formation","volume":"459","author":"Butt","year":"2014","journal-title":"Biochem. J."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"3712","DOI":"10.1128\/JB.01932-14","article-title":"Functional and structural analysis of HicA3-HicB3, a novel toxin-antitoxin system of Yersinia pestis","volume":"196","author":"Lazar","year":"2014","journal-title":"J. Bacteriol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"6371","DOI":"10.1093\/nar\/gky469","article-title":"Functional insights into the Streptococcus pneumoniae HicBA toxin-antitoxin system based on a structural study","volume":"46","author":"Kim","year":"2018","journal-title":"Nucleic Acids Res."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Chan, W.T., Espinosa, M., and Yeo, C.C. (2016). Keeping the wolves at bay: Antitoxins of prokaryotic type II toxin-antitoxin systems. Front. Mol. Biosci., 3.","DOI":"10.3389\/fmolb.2016.00009"},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Gerdes, K. (2024). Diverse genetic contexts of HicA toxin domains propose a role in anti-phage defense. mBio, 15.","DOI":"10.1128\/mbio.03293-23"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"D749","DOI":"10.1093\/nar\/gkx1033","article-title":"TADB 2.0: An updated database of bacterial type II toxin-antitoxin loci","volume":"46","author":"Xie","year":"2018","journal-title":"Nucleic Acids Res."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"4121","DOI":"10.1128\/jb.177.14.4121-4130.1995","article-title":"Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter","volume":"177","author":"Guzman","year":"1995","journal-title":"J. Bacteriol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"232","DOI":"10.1094\/MPMI.2000.13.2.232","article-title":"Small, stable shuttle vectors based on the minimal pVS1 replicon for use in gram-negative, plant-associated bacteria","volume":"13","author":"Heeb","year":"2000","journal-title":"Mol. Plant-Microbe Interact."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"5291","DOI":"10.1128\/JB.180.20.5291-5298.1998","article-title":"Iron-responsive gene regulation in a Campylobacter jejuni fur mutant","volume":"180","author":"Wooldridge","year":"1998","journal-title":"J. Bacteriol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2077","DOI":"10.1128\/JB.01408-06","article-title":"New functional identity for the DNA uptake sequence in transformation and its presence in transcriptional terminators","volume":"189","author":"Ambur","year":"2007","journal-title":"J. Bacteriol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1007\/978-1-4939-9496-0_10","article-title":"Transformation in Neisseria gonorrhoeae","volume":"1997","author":"Callaghan","year":"2019","journal-title":"Methods Mol. Biol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"3068","DOI":"10.1128\/AEM.07871-11","article-title":"New complementation constructs for inducible and constitutive gene expression in Neisseria gonorrhoeae and Neisseria meningitidis","volume":"78","author":"Ramsey","year":"2012","journal-title":"Appl. Environ. Microbiol."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Mitchev, N., Singh, R., Ramsuran, V., Ismail, A., Allam, M., Kwenda, S., Mnyameni, F., Garrett, N., Swe Swe-Han, K., and Niehaus, A.J. (2022). Assessment of antibiotic resistance and efflux pump gene expression in Neisseria gonorrhoeae isolates from South Africa by quantitative real-time PCR and regression analysis. Int. J. Microbiol., 2022.","DOI":"10.1155\/2022\/7318325"},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Piekarowicz, A., K\u0142yz, A., Majchrzak, M., Adamczyk-Pop\u0142awska, M., Maugel, T.K., and Stein, D.C. (2007). Characterization of the dsDNA prophage sequences in the genome of Neisseria gonorrhoeae and visualization of productive bacteriophage. BMC Microbiol., 7.","DOI":"10.1186\/1471-2180-7-66"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"124","DOI":"10.12688\/wellcomeopenres.14826.1","article-title":"Open-access bacterial population genomics: BIGSdb software, the PubMLST.org website and their applications","volume":"3","author":"Jolley","year":"2018","journal-title":"Wellcome Open Res."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1675","DOI":"10.1016\/j.str.2019.08.008","article-title":"The E. coli HicB antitoxin contains a structurally stable helix-turn-helix DNA binding domain","volume":"27","author":"Manav","year":"2019","journal-title":"Structure"},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Ross, B.N., Micheva-Viteva, S., Hong-Geller, E., and Torres, A.G. (2019). Evaluating the role of Burkholderia pseudomallei K96243 toxins BPSS0390, BPSS0395, and BPSS1584 in persistent infection. Cell Microbiol., 21.","DOI":"10.1111\/cmi.13096"},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Li, G., Shen, M., Lu, S., Le, S., Tan, Y., Wang, J., Zhao, X., Shen, W., Guo, K., and Yang, Y. (2016). Identification and characterization of the HicAB toxin-antitoxin system in the opportunistic pathogen Pseudomonas aeruginosa. Toxins, 8.","DOI":"10.3390\/toxins8040113"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Yarahmadi, A., Najafiyan, H., Yousefi, M.H., Khosravi, E., Shabani, E., Afkhami, H., and Aghaei, S.S. (2025). Beyond antibiotics: Exploring multifaceted approaches to combat bacterial resistance in the modern era: A comprehensive review. Front. Cell Infect. Microbiol., 15.","DOI":"10.3389\/fcimb.2025.1493915"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"7505","DOI":"10.1093\/emboj\/17.24.7505","article-title":"Molecular basis of double-stranded RNA-protein interactions: Structure of a dsRNA-binding domain complexed with dsRNA","volume":"17","author":"Ryter","year":"1998","journal-title":"EMBO J."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"e0004925","DOI":"10.1128\/jb.00049-25","article-title":"CRISPRi-mediated repression of three cI repressors induces the expression of three related Neisseria gonorrhoeae bacteriophages","volume":"207","author":"Geslewitz","year":"2025","journal-title":"J. Bacteriol."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"2228","DOI":"10.1128\/JB.00165-16","article-title":"Seminal plasma promotes Neisseria gonorrhoeae aggregation and biofilm formation","volume":"198","author":"Anderson","year":"2016","journal-title":"J. Bacteriol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1856","DOI":"10.1086\/593336","article-title":"Gonococcal cervicitis: A role for biofilm in pathogenesis","volume":"198","author":"Steichen","year":"2008","journal-title":"J. Infect. Dis."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"896","DOI":"10.1128\/IAI.68.2.896-905.2000","article-title":"Isolation of Neisseria gonorrhoeae mutants that show enhanced trafficking across polarized T84 epithelial monolayers","volume":"68","author":"Hopper","year":"2000","journal-title":"Infect. Immun."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"12515","DOI":"10.1021\/bi0511080","article-title":"Neisseria gonorrhoeae FitA interacts with FitB to bind DNA through its ribbon-helix-helix motif","volume":"44","author":"Wilbur","year":"2005","journal-title":"Biochemistry"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"37942","DOI":"10.1074\/jbc.M605198200","article-title":"Structure of FitAB from Neisseria gonorrhoeae bound to DNA reveals a tetramer of toxin-antitoxin heterodimers containing PIN domains and ribbon-helix-helix motifs","volume":"281","author":"Mattison","year":"2006","journal-title":"J. Biol. Chem."},{"key":"ref_47","doi-asserted-by":"crossref","unstructured":"Pachulec, E., and van der Does, C. (2010). Conjugative plasmids of Neisseria gonorrhoeae. PLoS ONE, 5.","DOI":"10.1371\/journal.pone.0009962"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"1686","DOI":"10.1038\/s41467-018-03652-8","article-title":"The ng_\u03b61 toxin of the gonococcal epsilon\/zeta toxin\/antitoxin system drains precursors for cell wall synthesis","volume":"9","author":"Rocker","year":"2018","journal-title":"Nat. Commun."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"939","DOI":"10.1038\/s41564-025-01968-y","article-title":"A quinolone N-oxide antibiotic selectively targets Neisseria gonorrhoeae via its toxin-antitoxin system","volume":"10","author":"Mix","year":"2025","journal-title":"Nat. Microbiol."}],"container-title":["Microorganisms"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2607\/13\/7\/1619\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T18:07:14Z","timestamp":1760033234000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2607\/13\/7\/1619"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,7,9]]},"references-count":49,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2025,7]]}},"alternative-id":["microorganisms13071619"],"URL":"https:\/\/doi.org\/10.3390\/microorganisms13071619","relation":{},"ISSN":["2076-2607"],"issn-type":[{"type":"electronic","value":"2076-2607"}],"subject":[],"published":{"date-parts":[[2025,7,9]]}}}