Novobiocin inhibits membrane synthesis and vacuole formation of Enterococcus faecalis protoplasts.

Rintaro Tsuchikado,Satoshi Kami,Sawako Takahashi,Hiromi Nishida

doi:10.15698/mic2020.11.735

Rintaro Tsuchikado, Satoshi Kami + Show 2 more

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https://doi.org/10.15698/mic2020.11.735

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Abstract

We demonstrate that plasma membrane biosynthesis and vacuole formation require DNA replication in Enterococcus faecalis protoplasts. The replication inhibitor novobiocin inhibited not only DNA replication but also cell enlargement (plasma membrane biosynthesis) and vacuole formation during the enlargement of the E. faecalis protoplasts. After novobiocin treatment prior to vacuole formation, the cell size of E. faecalis protoplasts was limited to 6 μm in diameter and the cells lacked vacuoles. When novobiocin was added after vacuole formation, E. faecalis protoplasts grew with vacuole enlargement; after novobiocin removal, protoplasts were enlarged again. Although cell size distribution of the protoplasts was similar following the 24 h and 48 h novobiocin treatments, after 72 h of novobiocin treatment there was a greater number of smaller sized protoplasts, suggesting that extended novobiocin treatment may inhibit the re-enlargement of E. faecalis protoplasts after novobiocin removal. Our findings demonstrate that novobiocin can control the enlargement of E. faecalis protoplasts due to inhibition of DNA replication.

Highlights

DNA replication is essential for cell division in normally dividing bacterial cells [1] and affects both cell morphology and cell division [2,3,4]
Replication, membrane, and vacuole of protoplasts tion cycle (Cq) value decreases with increasing DNA concentration. quantitative PCR (qPCR) showed that the quantification cycle (Cq) values decreased from 0 h to 120 h of incubation of E. faecalis protoplasts in Difco Marine Broth (DMB) plus penicillin (Fig. 1A, Table 1 upper part)
This study demonstrates that DNA replication and cell enlargement of E. faecalis protoplasts stopped at the same time, at 120 h of incubation in DMB containing penicillin (Fig. 1), and suggests that plasma membrane biosynthesis requires

Summary

Introduction

DNA replication is essential for cell division in normally dividing bacterial cells (native forms) [1] and affects both cell morphology and cell division [2,3,4]. It has been reported that the bacterial chromosome DNA attaches to the plasma membrane [5,6,7,8,9,10] cell division does not occur in bacterial protoplasts or spheroplasts in the presence of an inhibitor of peptidoglycan biosynthesis [11,12,13,14]; removal of the peptidoglycan synthetic inhibitor can induce recovery from Escherichia coli spheroplasts to their native forms [15,16,17]. Spheroplasts can enlarge in Difco Marine Broth (DMB) containing a peptidoglycan biosynthesis inhibitor, for example, penicillin [18,19,20,21], and interestingly, DNA replication has been reported during protoplast or spheroplast enlargement [13, 14, 18, 19, 23]. Direct injection of DNA into spheroplasts that are larger than 15 μm in diameter leads to vacuole formation [13, 14, 22, 23]

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