Heme interplay between IlsA and IsdC: Two structurally different surface proteins from Bacillus cereus

Abi-Khalil, Elise; Segond, Diego; Terpstra, Tyson; André-Leroux, Gwénaëlle; Kallassy, Mireille; Lereclus, Didier; Bou-Abdallah, Fadi; Nielsen-LeRoux, Christina

doi:10.1016/j.bbagen.2015.06.006

Cited by 8 publications

(8 citation statements)

References 45 publications

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“…Then, the ABC system encoded within lsa1836-1840 genes was automatically annotated as involved in cobalamin transport, whilst it shows some levels of similarities with heme import systems described in Gram-positive bacteria (40–43). At first, we carried out a multiple alignment of all putative substrate-binding lipoproteins encoded in the L. sakei 23K genome and noticed that Lsa1839 protein was closely related to Lsa0399 from the Fhu system (data not shown) suggesting a possible link to iron/heme transport.…”

Section: Resultsmentioning

confidence: 99%

Heme uptake inLactobacillus sakeievidenced by a new ECF-like transport system

Verplaetse

André-Leroux

Duhutrel

et al. 2019

Preprint

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19Lactobacillus sakei is a non-pathogenic lactic acid bacterium and a natural inhabitant of meat 20 ecosystems. Although red meat is a heme-rich environment, L. sakei does not need iron or 21 heme for growth, while possessing a heme-dependent catalase. Iron incorporation into L. 22 sakei from myoglobin and hemoglobin was formerly shown by microscopy and the L. sakei 23 genome reveals a complete equipment for iron and heme transport. Here, we report the 24 characterization of a five-gene cluster (lsa1836-1840) encoding a putative metal iron ABC 25 transporter. Interestingly, this cluster, together with a heme dependent catalase gene, is also 26 conserved in other species from the meat ecosystem. Our bioinformatic analyses revealed 27 that the locus might refer to a complete machinery of an Energy Coupling Factor (ECF) 28 transport system. We quantified in vitro the intracellular heme in wild-type (WT) and in our 29Dlsa1836-1840 deletion mutant using an intracellular heme sensor and ICP-Mass 30 spectrometry for quantifying incorporated 57 Fe heme. We showed that in the WT L. sakei, 31 heme accumulation occurs fast and massively in the presence of hemin, while the deletion 32 mutant was impaired in heme uptake; this ability was restored by in trans complementation. 33 Our results establish the main role of the L. sakei Lsa1836-1840 ECF-like system in heme 34 uptake. This research outcome shed new light on other possible functions of ECF-like 35 systems. 36 37 38 Importance 39Lactobacillus sakei is a non-pathogenic bacterial species exhibiting high fitness in heme rich 40 environments such as meat products, although it does not need iron nor heme for growth. 41Heme capture and utilization capacities are often associated with pathogenic species and are 42 considered as virulence-associated factors in the infected hosts. For these reasons, iron 43 3 acquisition systems have been deeply studied in such species, while for non-pathogenic 44 bacteria the information is scarce. Genomic data revealed that several putative iron 45 transporters are present in the genome of the lactic acid bacterium L. sakei. In this study, we 46 demonstrate that one of them, is an ECF-like ABC transporter with a functional role in heme 47 transport. Such evidence has not yet been brought for an ECF, therefore our study reveals a 48 new class of heme transport system. 49 50 51 7 It was evidenced that ECF transporters constitute a novel family of conserved membrane 126 transporters in prokaryotes, while sharing a similar four domains organization as the ABC 127 transporters. Each ECF displays a pair of cytosolic nucleotide-binding ATPases (the A and A' 128 components also called EcfA and EcfA'), a membrane-embedded substrate-binding protein 129 (the S component or the EcfS), and a transmembrane energy-coupling component (The T 130 component or EcfT). The quadripartite organization has a 1:1:1:1 stoichiometry. Notably, the 131 S component renders ECF mechanistically distinct from ABC transport systems as it is 132 predicted to shuttle within the ...

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Section: Resultsmentioning

confidence: 99%

Heme uptake inLactobacillus sakeievidenced by a new ECF-like transport system

Verplaetse

André-Leroux

Duhutrel

et al. 2019

Preprint

Self Cite

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“…Then, the ABC system encoded within lsa1836-1840 genes was automatically annotated as involved in cobalamin transport, whilst it shows some levels of similarities with heme import systems described in Gram-positive bacteria (40)(41)(42)(43). At first, we carried out a multiple alignment of all putative substrate-binding lipoproteins encoded in the L. sakei 23K genome and noticed that Lsa1839 protein was closely related to Lsa0399 from the Fhu system (data not shown) suggesting a possible link to iron/heme transport.…”

Section: Putative Iron and Heme Transport Systems In Lactobacillus Sakeimentioning

confidence: 99%

Heme Uptake in Lactobacillus sakei Evidenced by a New Energy Coupling Factor (ECF)-Like Transport System

Verplaetse

André-Leroux

Duhutrel

et al. 2020

Appl Environ Microbiol

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Lactobacillus sakei is a non-pathogenic lactic acid bacterium and a natural inhabitant of meat ecosystems. Although red meat is a heme-rich environment, L. sakei does not need iron or heme for growth, while possessing a heme-dependent catalase. Iron incorporation into L. sakei from myoglobin and hemoglobin was formerly shown by microscopy and the L. sakei genome reveals the complete equipment for iron and heme transport. Here, we report the characterization of a five-gene cluster (lsa1836-1840) encoding a putative metal iron ABC transporter. Interestingly, this cluster, together with a heme dependent catalase gene, is also conserved in other species from the meat ecosystem. Our bioinformatic analyses revealed that the locus might correspond to a complete machinery of an Energy Coupling Factor (ECF) transport system. We quantified in vitro the intracellular heme in wild-type (WT) and in our Δlsa1836-1840 deletion mutant using an intracellular heme sensor and ICP-Mass spectrometry for quantifying incorporated 57Fe heme. We showed that in the WT L. sakei, heme accumulation occurs rapidly and massively in the presence of hemin, while the deletion mutant was impaired in heme uptake; this ability was restored by in trans complementation. Our results establish the main role of the L. sakei Lsa1836-1840 ECF-like system in heme uptake. Therefore, this research outcome sheds new light on other possible functions of ECF-like systems. Importance Lactobacillus sakei is a non-pathogenic bacterial species exhibiting high fitness in heme rich environments such as meat products, although it does not need iron nor heme for growth. Heme capture and utilization capacities are often associated with pathogenic species and are considered as virulence-associated factors in the infected hosts. For these reasons, iron acquisition systems have been deeply studied in such species, while for non-pathogenic bacteria the information is scarce. Genomic data revealed that several putative iron transporters are present in the genome of the lactic acid bacterium L. sakei. In this study, we demonstrate that one of them, is an ECF-like ABC transporter with a functional role in heme transport. Such evidence has not yet been brought for an ECF, therefore our study reveals a new class of heme transport system.

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“…B. cereus possesses different strategies to scavenge sequestered iron during host infection. This can be done through direct interactions using surface proteins [14][15][16], or indirect interactions using both surface proteins to bind the iron source and secreted siderophores, which are small molecules with high affinity for free Fe 3+ [17]. The siderophore bacillibactin, encoded by genes of the entA-dhbBCF cluster [18], has been shown to have a major impact on bacterial survival in a low iron environment.…”

Section: Introductionmentioning

confidence: 99%

“…G. mellonella was also proved to be an appropriate model for B. cereus, B. thuringiensis, Pseudomonas entomophila, and Listeria monocytogenes oral infection studies [35][36][37] and for a B. cereus IVET (in vivo expression technology) promoter trap screening [38]. The latter approach identified IlsA a unique surface protein involved in iron acquisition [14,15] and revealed expression of other iron regulated factors during infection [38]. To decipher interactions between microorganisms and the host during infection, we have chosen a challenging approach based on lasercapture microdissection (LCM) [39,40].…”

Section: Introductionmentioning

confidence: 99%

Laser capture microdissection to study Bacillus cereus iron homeostasis gene expression during Galleria mellonella in vivo gut colonization

et al. 2021

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Bacillus cereus is a Gram-positive opportunistic pathogen closely related to the entomopathogen, Bacillus thuringiensis , both of which are involved in intestinal infections. Iron is an essential micronutrient for full growth and virulence of pathogens during infection. However, little is known about iron homeostasis during gut infection. Therefore, we aimed to assess the expression of B. cereus genes related to bacterial iron homeostasis, virulence and oxidative stress. The hypothesis is that the expression of such genes would vary between early and later stage colonization in correlation to gut cell damage. To perform the study, a germ-free Galleria mellonella model was set up in order to adapt the use of Laser-capture microdissection (LCM), to select precise areas in the gut lumen from frozen whole larval cryo-sections. Analyses were performed from alive larvae and the expression of targeted genes was assessed byspecific pre-amplification of mRNA followed by quantitative PCR. Firstly, the results reinforce the reliability of LCM, despite a low amount of bacterial RNA recovered. Secondly, bacterial genes involved in iron homeostasis are expressed in the lumen at both 3 and 16 hours post force-feeding. Thirdly, iron gene expression is slightly modulated during gut infection, and lastly, the mRNA of G. mellonella encoding for ferritin and transferrin iron storage and transport are recovered too. Therefore, iron homeostasis should play a role in B. cereus gut colonization. Furthermore, we demonstrate for the first time the value of using LCM for specific in situ gene expression analysis of extracellular bacteria in a whole animal.

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Heme interplay between IlsA and IsdC: Two structurally different surface proteins from Bacillus cereus

Abstract: An important role for the complete Isd system in heme-associated bacterial growth is demonstrated and new insights into the interplay between an Isd NEAT surface protein and an IlsA-NEAT-LRR protein, both of which appear to be involved in heme-iron acquisition in B. cereus are revealed.

Cited by 8 publications

References 45 publications

Heme uptake inLactobacillus sakeievidenced by a new ECF-like transport system

Heme uptake inLactobacillus sakeievidenced by a new ECF-like transport system

Heme Uptake in Lactobacillus sakei Evidenced by a New Energy Coupling Factor (ECF)-Like Transport System

Laser capture microdissection to study Bacillus cereus iron homeostasis gene expression during Galleria mellonella in vivo gut colonization

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