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Zn2+ Uptake in Streptococcus pyogenes: Characterization of adcA and lmb Null Mutants.

Tedde V, Rosini R, Galeotti CL - PLoS ONE (2016)

Bottom Line: An effective regulation of metal ion homeostasis is essential for the growth of microorganisms in any environment and in pathogenic bacteria is strongly associated with their ability to invade and colonise their hosts.In contrast, Lmb and HtpA were expressed at detectable levels only during growth in the presence of low zinc concentrations or in the adcA mutant, when expression of lmb is required to compensate for the lack of adcA expression.In the latter case, Lmb and HtpA were overexpressed by several fold, thus indicating that also in GAS AdcA is a zinc-specific importer and, although it shares this function with Lmb, the two substrate-binding proteins do not show fully overlapping roles in zinc homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Research Centre, GlaxoSmithKline Vaccines S.r.l., Siena, Italy.

ABSTRACT
An effective regulation of metal ion homeostasis is essential for the growth of microorganisms in any environment and in pathogenic bacteria is strongly associated with their ability to invade and colonise their hosts. To gain a better insight into zinc acquisition in Group A Streptococcus (GAS) we characterized deletion mutants of the adcA and lmb genes of Streptococcus pyogenes strain MGAS5005 encoding the orthologues of AdcA and AdcAII, the two surface lipoproteins with partly redundant roles in zinc homeostasis in Streptococcus pneumoniae. Null adcA and lmb mutants were analysed for their capability to grow in zinc-depleted conditions and were found to be more susceptible to zinc starvation, a phenotype that could be rescued by the addition of Zn2+ ions to the growth medium. Expression of AdcA, Lmb and HtpA, the polyhistidine triad protein encoded by the gene adjacent to lmb, during growth under conditions of limited zinc availability was examined by Western blot analysis in wild type and mutant strains. In the wild type strain, AdcA was always present with little variation in expression levels between conditions of excess or limited zinc availability. In contrast, Lmb and HtpA were expressed at detectable levels only during growth in the presence of low zinc concentrations or in the adcA mutant, when expression of lmb is required to compensate for the lack of adcA expression. In the latter case, Lmb and HtpA were overexpressed by several fold, thus indicating that also in GAS AdcA is a zinc-specific importer and, although it shares this function with Lmb, the two substrate-binding proteins do not show fully overlapping roles in zinc homeostasis.

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Related in: MedlinePlus

Expression of AdcA, Lmb and HtpA during growth in zinc-depleted medium.Western blot analysis of total cell extracts from S. pyogenes MGAS5005 wild type, ΔadcA and Δlmb  mutants grown in complete medium (THY) or in zinc-depleted medium (THY + 35 μM TPEN) containing increasing amounts of ZnCl2. (A) Western blot using anti-AdcA specific antibodies. (B) Western blot using anti-Lmb specific antibodies. (C) Western blot using anti-HtpA specific antibodies.
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pone.0152835.g006: Expression of AdcA, Lmb and HtpA during growth in zinc-depleted medium.Western blot analysis of total cell extracts from S. pyogenes MGAS5005 wild type, ΔadcA and Δlmb mutants grown in complete medium (THY) or in zinc-depleted medium (THY + 35 μM TPEN) containing increasing amounts of ZnCl2. (A) Western blot using anti-AdcA specific antibodies. (B) Western blot using anti-Lmb specific antibodies. (C) Western blot using anti-HtpA specific antibodies.

Mentions: Wild type S. pyogenes MGAS5005 and isogenic mutant strains lacking adcA or lmb were analysed for expression of AdcA, Lmb and HtpA during growth in complete medium (THY) or in zinc-depleted medium (THY + 35 μM TPEN) containing increasing amounts of ZnCl2. The immunoblot analyses presented in Fig 6 confirm the results obtained for the ΔadcA mutant (Fig 3). Lmb and HtpA were both expressed at higher levels in the ΔadcA mutant, strongly supporting the assumption that also in S. pyogenes AdcA and Lmb have overlapping roles in zinc acquisition [8] (Fig 6B and 6C).


Zn2+ Uptake in Streptococcus pyogenes: Characterization of adcA and lmb Null Mutants.

Tedde V, Rosini R, Galeotti CL - PLoS ONE (2016)

Expression of AdcA, Lmb and HtpA during growth in zinc-depleted medium.Western blot analysis of total cell extracts from S. pyogenes MGAS5005 wild type, ΔadcA and Δlmb  mutants grown in complete medium (THY) or in zinc-depleted medium (THY + 35 μM TPEN) containing increasing amounts of ZnCl2. (A) Western blot using anti-AdcA specific antibodies. (B) Western blot using anti-Lmb specific antibodies. (C) Western blot using anti-HtpA specific antibodies.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4816340&req=5

pone.0152835.g006: Expression of AdcA, Lmb and HtpA during growth in zinc-depleted medium.Western blot analysis of total cell extracts from S. pyogenes MGAS5005 wild type, ΔadcA and Δlmb mutants grown in complete medium (THY) or in zinc-depleted medium (THY + 35 μM TPEN) containing increasing amounts of ZnCl2. (A) Western blot using anti-AdcA specific antibodies. (B) Western blot using anti-Lmb specific antibodies. (C) Western blot using anti-HtpA specific antibodies.
Mentions: Wild type S. pyogenes MGAS5005 and isogenic mutant strains lacking adcA or lmb were analysed for expression of AdcA, Lmb and HtpA during growth in complete medium (THY) or in zinc-depleted medium (THY + 35 μM TPEN) containing increasing amounts of ZnCl2. The immunoblot analyses presented in Fig 6 confirm the results obtained for the ΔadcA mutant (Fig 3). Lmb and HtpA were both expressed at higher levels in the ΔadcA mutant, strongly supporting the assumption that also in S. pyogenes AdcA and Lmb have overlapping roles in zinc acquisition [8] (Fig 6B and 6C).

Bottom Line: An effective regulation of metal ion homeostasis is essential for the growth of microorganisms in any environment and in pathogenic bacteria is strongly associated with their ability to invade and colonise their hosts.In contrast, Lmb and HtpA were expressed at detectable levels only during growth in the presence of low zinc concentrations or in the adcA mutant, when expression of lmb is required to compensate for the lack of adcA expression.In the latter case, Lmb and HtpA were overexpressed by several fold, thus indicating that also in GAS AdcA is a zinc-specific importer and, although it shares this function with Lmb, the two substrate-binding proteins do not show fully overlapping roles in zinc homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Research Centre, GlaxoSmithKline Vaccines S.r.l., Siena, Italy.

ABSTRACT
An effective regulation of metal ion homeostasis is essential for the growth of microorganisms in any environment and in pathogenic bacteria is strongly associated with their ability to invade and colonise their hosts. To gain a better insight into zinc acquisition in Group A Streptococcus (GAS) we characterized deletion mutants of the adcA and lmb genes of Streptococcus pyogenes strain MGAS5005 encoding the orthologues of AdcA and AdcAII, the two surface lipoproteins with partly redundant roles in zinc homeostasis in Streptococcus pneumoniae. Null adcA and lmb mutants were analysed for their capability to grow in zinc-depleted conditions and were found to be more susceptible to zinc starvation, a phenotype that could be rescued by the addition of Zn2+ ions to the growth medium. Expression of AdcA, Lmb and HtpA, the polyhistidine triad protein encoded by the gene adjacent to lmb, during growth under conditions of limited zinc availability was examined by Western blot analysis in wild type and mutant strains. In the wild type strain, AdcA was always present with little variation in expression levels between conditions of excess or limited zinc availability. In contrast, Lmb and HtpA were expressed at detectable levels only during growth in the presence of low zinc concentrations or in the adcA mutant, when expression of lmb is required to compensate for the lack of adcA expression. In the latter case, Lmb and HtpA were overexpressed by several fold, thus indicating that also in GAS AdcA is a zinc-specific importer and, although it shares this function with Lmb, the two substrate-binding proteins do not show fully overlapping roles in zinc homeostasis.

Show MeSH
Related in: MedlinePlus