Analysis of a bac operon-silenced strain suggests pleiotropic effects of bacilysin in Bacillus subtilis

ERTEKİN O., Kutnu M., Taskin A. A., Demir M., Karataş A., ÖZCENGİZ G.

JOURNAL OF MICROBIOLOGY, cilt.58, sa.4, ss.297-313, 2020 (SCI-Expanded) identifier identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 58 Sayı: 4
  • Basım Tarihi: 2020
  • Doi Numarası: 10.1007/s12275-020-9064-0
  • Dergi Adı: JOURNAL OF MICROBIOLOGY
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus, Agricultural & Environmental Science Database, Aquatic Science & Fisheries Abstracts (ASFA), BIOSIS, Biotechnology Research Abstracts, CAB Abstracts, EMBASE, MEDLINE, Veterinary Science Database
  • Sayfa Sayıları: ss.297-313
  • Anahtar Kelimeler: Bacillus subtilis, bacilysin, bac operon, sporulation, chemical complementation, pleiotropy, comparative proteomics, RT-qPCR, GENERAL STRESS-RESPONSE, GENOME-WIDE ANALYSIS, TRANSCRIPTIONAL RESPONSES, GLOBAL CHARACTERIZATION, FUNCTIONAL GENOMICS, COAT PROTEIN, BIOSYNTHESIS, EXPRESSION, SPORULATION, REGULATOR
  • Orta Doğu Teknik Üniversitesi Adresli: Evet

Özet

Bacilysin, as the simplest peptide antibiotic made up of only L-alanine and L-anticapsin, is produced and excreted by Bacillus subtilis under the control of quorum sensing. We analyzed bacilysin-nonproducing strain OGU1 which was obtained by bacA-targeted pMutin T3 insertion into the parental strain genome resulting in a genomic organization (bacA '::lacZ::erm::bacABCDEF) to form an IPTG-inducible bac operon. Although IPTG induction provided 3- to 5-fold increment in the transcription of bac operon genes, no bacilysin activity was detectable in bioassays and inability of the OGU1 to form bacilysin was confirmed by UPLC-mass spectrometry analysis. Phenotypic analyses revealed the deficiencies in OGU1 with respect to colony pigmentation, spore coat proteins, spore resistance and germination, which could be rescued by external addition of bacilysin concentrate into its cultures. 2DE MALDI-TOF/MS and nanoLC-MS/MS were used as complementary approaches to compare cytosolic proteomes of OGU1. 2-DE identified 159 differentially expressed proteins corresponding to 121 distinct ORFs. In nanoLC-MS/MS, 76 proteins were differentially expressed in OGU1. Quantitative transcript analyses of selected genes validated the proteomic findings. Overall, the results pointed to the impact of bacilysin on expression of certain proteins of sporulation and morphogenesis; the members of mother cell compartment-specific sigma(E) and sigma(K) regulons in particular, quorum sensing and two component-global regulatory systems, peptide transport, stress response as well as CodY- and ScoC-regulated proteins.