Abstract
Twin-arginine protein translocation (Tat) pathways are required for transport of folded proteins across bacterial, archaeal and chloroplast membranes. Recent studies indicate that Tat has evolved into a mainstream pathway for protein secretion in certain halophilic archaea, which thrive in highly saline environments. Here, we investigated the effects of environmental salinity on Tat-dependent protein secretion by the Gram-positive soil bacterium Bacillus subtilis, which encounters widely differing salt concentrations in its natural habitats. The results show that environmental salinity determines the specificity and need for Tat-dependent secretion of the Dyp-type peroxidase YwbN in B. subtilis. Under high salinity growth conditions, at least three Tat translocase subunits, namely TatAd, TatAy and TatCy, are involved in the secretion of YwbN. Yet, a significant level of Tat-independent YwbN secretion is also observed under these conditions. When B. subtilis is grown in medium with 1% NaCl or without NaCl, the secretion of YwbN depends strictly on the previously described “minimal Tat translocase” consisting of the TatAy and TatCy subunits. Notably, in medium without NaCl, both tatAyCy and ywbN mutants display significantly reduced exponential growth rates and severe cell lysis. This is due to a critical role of secreted YwbN in the acquisition of iron under these conditions. Taken together, our findings show that environmental conditions, such as salinity, can determine the specificity and need for the secretion of a bacterial Tat substrate.
Highlights
The transport of proteins across biological membranes and their subsequent secretion into external milieus are vital processes for all known microorganisms
TatAyCy-independent secretion of YwbN at high salinity To investigate whether high salinity might impact on Tatdependent secretion, we investigated the secretion of YwbN in LB medium with a final salt concentration of 6%, which is comparable to that of highly saline environments or brine
Secreted LipA was readily detectable (Figure 1), and Sypro Ruby-staining of the respective gels did not reveal any major differences in the total amounts of protein secreted by the parental and tat mutant strains grown in LB with 6% salt (Figure S2). These findings show that the TatAyCy translocase is not exclusively involved in YwbN secretion under high salinity growth conditions, and that TatAd and/or TatCd are involved in this process
Summary
The transport of proteins across biological membranes and their subsequent secretion into external milieus are vital processes for all known microorganisms. These processes depend on the activity of dedicated molecular machines. A first critical step in protein secretion is the passage of transported proteins through the cytoplasmic membrane, which can occur either in an unfolded state via the general secretion (Sec) machinery, or in a folded state via the twin-arginine translocation (Tat) machinery [1,2,3,4,5]. Tat-dependent proteins fold prior to translocation, while the folding of Sec-dependent proteins occurs post-translocationally [7]
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