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A tamB homolog is involved in maintenance of cell envelope integrity and stress resistance of Deinococcus radiodurans

View Article: PubMed Central - PubMed

ABSTRACT

The translocation and assembly module (TAM) in bacteria consists of TamA and TamB that form a complex to control the transport and secretion of outer membrane proteins. Herein, we demonstrated that the DR_1462-DR_1461-DR_1460 gene loci on chromosome 1 of Deinococcus radiodurans, which lacks tamA homologs, is a tamB homolog (DR_146T) with two tamB motifs and a DUF490 motif. Mutation of DR_146T resulted in cell envelope peeling and a decrease in resistance to shear stress and osmotic pressure, as well as an increase in oxidative stress resistance, consistent with the phenotype of a surface layer (S-layer) protein SlpA (DR_2577) mutant, demonstrating the involvement of DR_146T in maintenance of cell envelope integrity. The 123 kDa SlpA was absent and only its fragments were present in the cell envelope of DR_146T mutant, suggesting that DR_146T might be involved in maintenance of the S-layer. A mutant lacking the DUF490 motif displayed only a slight alteration in phenotype compared with the wild type, suggesting DUF490 is less important than tamB motif for the function of DR_146T. These findings enhance our understanding of the properties of the multilayered envelope in extremophilic D. radiodurans, as well as the diversity and functions of TAMs in bacteria.

No MeSH data available.


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SEM and TEM images of D. radiodurans wild type and mutant strains.The cell envelope of ∆DR_146T exhibited severe damage with parts of the outer layer peeling off, and the inner layer exposed to the environment compared with that of the wild type. Images represent the SEM and TEM results for D. radiodurans wild type (a1–a2), ∆DR_146T-DUF490 (b1–b2), ∆DR_146T-TamB2nd -DUF490 (c1–c2), ∆DR_146T (d1–d2) and ∆DR_2577 (e1–e2), respectively. The inset diagram in (b2) shows an amplified region of the cell envelope. Scale bars indicate the corresponding length.
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f3: SEM and TEM images of D. radiodurans wild type and mutant strains.The cell envelope of ∆DR_146T exhibited severe damage with parts of the outer layer peeling off, and the inner layer exposed to the environment compared with that of the wild type. Images represent the SEM and TEM results for D. radiodurans wild type (a1–a2), ∆DR_146T-DUF490 (b1–b2), ∆DR_146T-TamB2nd -DUF490 (c1–c2), ∆DR_146T (d1–d2) and ∆DR_2577 (e1–e2), respectively. The inset diagram in (b2) shows an amplified region of the cell envelope. Scale bars indicate the corresponding length.

Mentions: The wild type colonies were circular and smooth, while the colonies of DR_146T mutant were ring-shaped and rugose similar to those of SlpA mutant (Supplementary Fig. S3). In liquid media, mutant cells tended to aggregate and settle more easily than wild type cells (Supplementary Fig. S4). Scanning electron microscopy (SEM) images showed that wild type cells were elliptical and grouped into diplococci or tetracocci (Fig. 3a1), whereas mutant cells displayed surface variation and shedding of the outer layer (Fig. 3b1–e1). Meanwhile, the results of transmission electron microscopy (TEM) demonstrated that the ultrastructure of the cell envelope of the mutants was different from that of wild type cells (Fig. 3a2–e2). The cell envelope of ∆DR_146T exhibited the most severe damage with parts of the outer layer peeling off, and the inner layer exposed to the environment (Fig. 3d). Damage to the cell envelope of ∆DR_146T-TamB2nd-DUF490 that is mutated in the second TamB and DUF490 motifs (Fig. 3c) was similar to that of the ∆DR_146T cells (Fig. 3d), but more severe than that of ∆DR_146T-DUF490 cells (Fig. 3b), indicating that TamB motif appeared to be more important than the DUF490 motif. SEM and TEM images revealed that the outmost layer peeled in the SlpA mutant (∆DR_2577) (Fig. 3e). Together with the cell growth phenotype, these results indicated that DR_146T might play an important role in maintaining cell envelope integrity. The cytoplasmic membrane (inner membrane) and peptidoglycan layer could still be formed during cell division, but formation of the outer layers was clearly inhibited in the mutant.


A tamB homolog is involved in maintenance of cell envelope integrity and stress resistance of Deinococcus radiodurans
SEM and TEM images of D. radiodurans wild type and mutant strains.The cell envelope of ∆DR_146T exhibited severe damage with parts of the outer layer peeling off, and the inner layer exposed to the environment compared with that of the wild type. Images represent the SEM and TEM results for D. radiodurans wild type (a1–a2), ∆DR_146T-DUF490 (b1–b2), ∆DR_146T-TamB2nd -DUF490 (c1–c2), ∆DR_146T (d1–d2) and ∆DR_2577 (e1–e2), respectively. The inset diagram in (b2) shows an amplified region of the cell envelope. Scale bars indicate the corresponding length.
© Copyright Policy - open-access
Related In: Results  -  Collection

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f3: SEM and TEM images of D. radiodurans wild type and mutant strains.The cell envelope of ∆DR_146T exhibited severe damage with parts of the outer layer peeling off, and the inner layer exposed to the environment compared with that of the wild type. Images represent the SEM and TEM results for D. radiodurans wild type (a1–a2), ∆DR_146T-DUF490 (b1–b2), ∆DR_146T-TamB2nd -DUF490 (c1–c2), ∆DR_146T (d1–d2) and ∆DR_2577 (e1–e2), respectively. The inset diagram in (b2) shows an amplified region of the cell envelope. Scale bars indicate the corresponding length.
Mentions: The wild type colonies were circular and smooth, while the colonies of DR_146T mutant were ring-shaped and rugose similar to those of SlpA mutant (Supplementary Fig. S3). In liquid media, mutant cells tended to aggregate and settle more easily than wild type cells (Supplementary Fig. S4). Scanning electron microscopy (SEM) images showed that wild type cells were elliptical and grouped into diplococci or tetracocci (Fig. 3a1), whereas mutant cells displayed surface variation and shedding of the outer layer (Fig. 3b1–e1). Meanwhile, the results of transmission electron microscopy (TEM) demonstrated that the ultrastructure of the cell envelope of the mutants was different from that of wild type cells (Fig. 3a2–e2). The cell envelope of ∆DR_146T exhibited the most severe damage with parts of the outer layer peeling off, and the inner layer exposed to the environment (Fig. 3d). Damage to the cell envelope of ∆DR_146T-TamB2nd-DUF490 that is mutated in the second TamB and DUF490 motifs (Fig. 3c) was similar to that of the ∆DR_146T cells (Fig. 3d), but more severe than that of ∆DR_146T-DUF490 cells (Fig. 3b), indicating that TamB motif appeared to be more important than the DUF490 motif. SEM and TEM images revealed that the outmost layer peeled in the SlpA mutant (∆DR_2577) (Fig. 3e). Together with the cell growth phenotype, these results indicated that DR_146T might play an important role in maintaining cell envelope integrity. The cytoplasmic membrane (inner membrane) and peptidoglycan layer could still be formed during cell division, but formation of the outer layers was clearly inhibited in the mutant.

View Article: PubMed Central - PubMed

ABSTRACT

The translocation and assembly module (TAM) in bacteria consists of TamA and TamB that form a complex to control the transport and secretion of outer membrane proteins. Herein, we demonstrated that the DR_1462-DR_1461-DR_1460 gene loci on chromosome 1 of Deinococcus radiodurans, which lacks tamA homologs, is a tamB homolog (DR_146T) with two tamB motifs and a DUF490 motif. Mutation of DR_146T resulted in cell envelope peeling and a decrease in resistance to shear stress and osmotic pressure, as well as an increase in oxidative stress resistance, consistent with the phenotype of a surface layer (S-layer) protein SlpA (DR_2577) mutant, demonstrating the involvement of DR_146T in maintenance of cell envelope integrity. The 123 kDa SlpA was absent and only its fragments were present in the cell envelope of DR_146T mutant, suggesting that DR_146T might be involved in maintenance of the S-layer. A mutant lacking the DUF490 motif displayed only a slight alteration in phenotype compared with the wild type, suggesting DUF490 is less important than tamB motif for the function of DR_146T. These findings enhance our understanding of the properties of the multilayered envelope in extremophilic D. radiodurans, as well as the diversity and functions of TAMs in bacteria.

No MeSH data available.


Related in: MedlinePlus