RESUMO
When Escherichia coli grows in the presence of DNA-damaging agents such as methyl methanesulphonate (MMS), absence of the full-length form of Translation Initiation Factor 2 (IF2-1) or deficiency in helicase activity of replication restart protein PriA leads to a considerable loss of viability. MMS sensitivity of these mutants was contingent on the stringent response alarmone (p)ppGpp being at low levels. While zero levels (ppGpp°) greatly aggravated sensitivity, high levels promoted resistance. Moreover, M+ mutations, which suppress amino acid auxotrophy of ppGpp° strains and which have been found to map to RNA polymerase subunits, largely restored resistance to IF2-1- and PriA helicase-deficient mutants. The truncated forms IF2-2/3 played a key part in inducing especially severe negative effects in ppGpp° cells when restart function priB was knocked out, causing loss of viability and severe cell filamentation, indicative of SOS induction. Even a strain with the wild-type infB allele exhibited significant filamentation and MMS sensitivity in this background whereas mutations that prevent expression of IF2-2/3 essentially eliminated filamentation and largely restored MMS resistance. The results suggest different influences of IF2-1 and IF2-2/3 on the replication restart system depending on (p)ppGpp levels, each having the capacity to maximize survival under differing growth conditions.
Assuntos
DNA Helicases/metabolismo , Escherichia coli K12/genética , Proteínas de Escherichia coli/metabolismo , Guanosina Tetrafosfato/farmacologia , Fator de Iniciação 2 em Procariotos/metabolismo , Bacteriófago mu/genética , Bacteriófago mu/fisiologia , Dano ao DNA/efeitos dos fármacos , DNA Helicases/genética , Replicação do DNA/efeitos dos fármacos , Escherichia coli K12/efeitos dos fármacos , Escherichia coli K12/crescimento & desenvolvimento , Proteínas de Escherichia coli/genética , Metanossulfonato de Metila/farmacologia , Fator de Iniciação 2 em Procariotos/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismoRESUMO
Escherichia coli translation initiation factor 2 (IF2) performs the unexpected function of promoting transition from recombination to replication during bacteriophage Mu transposition in vitro, leading to initiation by replication restart proteins. This function has suggested a role of IF2 in engaging cellular restart mechanisms and regulating the maintenance of genome integrity. To examine the potential effect of IF2 on restart mechanisms, we characterized its influence on cellular recovery following DNA damage by methyl methanesulfonate (MMS) and UV damage. Mutations that prevent expression of full-length IF2-1 or truncated IF2-2 and IF2-3 isoforms affected cellular growth or recovery following DNA damage differently, influencing different restart mechanisms. A deletion mutant (del1) expressing only IF2-2/3 was severely sensitive to growth in the presence of DNA-damaging agent MMS. Proficient as wild type in repairing DNA lesions and promoting replication restart upon removal of MMS, this mutant was nevertheless unable to sustain cell growth in the presence of MMS; however, growth in MMS could be partly restored by disruption of sulA, which encodes a cell division inhibitor induced during replication fork arrest. Moreover, such characteristics of del1 MMS sensitivity were shared by restart mutant priA300, which encodes a helicase-deficient restart protein. Epistasis analysis indicated that del1 in combination with priA300 had no further effects on cellular recovery from MMS and UV treatment; however, the del2/3 mutation, which allows expression of only IF2-1, synergistically increased UV sensitivity in combination with priA300. The results indicate that full-length IF2, in a function distinct from truncated forms, influences the engagement or activity of restart functions dependent on PriA helicase, allowing cellular growth when a DNA-damaging agent is present.
Assuntos
Bacteriófago mu/genética , Reparo do DNA , Replicação do DNA , Escherichia coli , Fator de Iniciação 2 em Procariotos , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/efeitos da radiação , DNA Helicases/genética , DNA Helicases/metabolismo , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/genética , Replicação do DNA/efeitos dos fármacos , Replicação do DNA/genética , Escherichia coli/genética , Escherichia coli/crescimento & desenvolvimento , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Instabilidade Genômica , Metanossulfonato de Metila/farmacologia , Fator de Iniciação 2 em Procariotos/genética , Fator de Iniciação 2 em Procariotos/metabolismo , Deleção de Sequência , Raios UltravioletaRESUMO
Background: Isotretinoin is frequently used for treatment of severe nodulocystic and papulopustular acne, however use is limited by mucocutaneous, ocular, and systemic side effects.Objective: (1) provide a systematic meta-analysis of ocular side effects during isotretinoin use and their corresponding incidences; (2) provide a narrative summary of ocular side effects during isotretinoin use reported in case reports.Methods: A systematic database search using predefined search terms was performed in PubMed, EMBASE, and Scopus from inception to 5 March, 2021. Predetermined inclusion and exclusion criteria were used to select included studies. In total, 53 original studies qualified for meta-analysis, and 41 case reports/series qualified for narrative results.Results: The studies included in the meta-analysis reported incidences of various ocular side effects including dry eye, eye sensitivity, vision changes, and ocular inflammatory conditions. Incidences across studies did vary, leading to considerable heterogeneity. The narrative results summarize more uncommon, but equally important, ocular side effects.Conclusions: Dry eye is the most commonly reported ocular side effect. Other less common, but more serious, ocular side effects including vision changes can occur. We recommend that isotretinoin prescribers monitor for dry eye. Limitations include the heterogeneity of reported incidences of ocular side effects between studies.