Secretome profiling of Propionibacterium freudenreichii reveals highly variable responses even among the closely related strains

Abstract

SummaryThis study compared the secretomes (proteins exported out of the cell) of Propionibacterium freudenreichii of different origin to identify plausible adaptation factors. Phylosecretomics indicated strain‐specific variation in secretion of adhesins/invasins (SlpA, InlA), cell‐wall hydrolysing (NlpC60 peptidase, transglycosylase), protective (RpfB) and moonlighting (DnaK, GroEL, GaPDH, IDH, ENO, ClpB) enzymes and/or proteins. Detailed secretome comparison suggested that one of the cereal strains (JS14) released a tip fimbrillin (FimB) in to the extracellular milieu, which was in line with the electron microscopy and genomic analyses, indicating the lack of surface‐associated fimbrial‐like structures, predicting a mutated type‐2 fimbrial gene cluster (fimB‐fimA‐srtC2) and production of anchorless FimB. Instead, the cereal strain produced high amounts of SlpB that tentatively mediated adherent growth on hydrophilic surface and adherence to hydrophobic material. One of the dairy strains (JS22), producing non‐covalently bound surface‐proteins (LspA, ClpB, AraI) and releasing SlpA and InlA into the culture medium, was found to form clumps under physiological conditions. The JS22 strain lacked SlpB and displayed a non‐clumping and biofilm‐forming phenotype only under conditions of increased ionic strength (300 mM NaCl). However, this strain cultured under the same conditions was not adherent to hydrophobic support, which supports the contributory role of SlpB in mediating hydrophobic interactions. Thus, this study reports significant secretome variation in P. freudenreichii and suggests that strain‐specific differences in protein export, modification and protein–protein interactions have been the driving forces behind the adaptation of this bacterial species.

Extracellular Vesicles Produced by the Probiotic Propionibacterium freudenreichii CIRM-BIA 129 Mitigate Inflammation by Modulating the NF-κB Pathway

“…The predicted interactions mapped to several immunology-related KEGG terms, such as signal transduction, infectious diseases and the immune system, thus shedding light on a possible immunomodulatory role for P. freudenreichii EVs. These predictions corroborate some previous findings which associated the P. freudenreichii bacterium with immunomodulatory roles ( Foligne et al, 2010 ; Deutsch et al, 2017 ; Do Carmo et al, 2017 ; Frohnmeyer et al, 2018 ; Rabah et al, 2018a ). Interestingly, the predicted data also suggested that this immunomodulation could involve the NF-κB pathway, since the nuclear factor NF-κB p105 subunit (NFKB1, P19838) was the most frequent interacting human protein according to the intersppi predictions and also the interactions shared between the two methods.…”

“…That is in line with the growing recognition of the widespread occurrence and diverse functions of EVs ( Brown et al, 2015 ; Liu et al, 2018a , b ). In the case of P. freudenreichii , a Gram-positive dairy probiotic bacterium, a recent study indicated the presence of an extracellular structure resembling a potential EV, which bulged out from the membrane of P. freudenreichii strain JS22 ( Frohnmeyer et al, 2018 ), suggesting that this species might produce EVs. Nevertheless, ours is the first complete report to have identified the occurrence of EVs in P. freudenreichii strain CIRM-BIA 129, and included their physicochemical, biochemical and functional characterization.…”

Mutation of the Surface Layer Protein SlpB Has Pleiotropic Effects in the Probiotic Propionibacterium freudenreichii CIRM-BIA 129

“…In addition, differential proteome was functionally classified using COG, showing a functional implication of differential proteins in cellular processes such as signaling, information storage, processing, and metabolism. Specifically, this study showed that the moonlighting enolase and NlpC/P60 are both exported (Frohnmeyer et al, 2018 ), as it was recently observed in the cutaneous Propionibacterium acnes strain (Jeon et al, 2017 ). These moonlighting proteins were downregulated in CB129Δ slpB .…”

Partial catalytic Cys oxidation of human GAPDH to Cys-sulfonic acid.

“…GAPDH is mainly a cytoplasmic enzyme, but it has been reported to moonlight as a cell surface and/or extracellular enzyme in healthy human cells 49,50 , in human cells subjected to stresses such as micronutrient starvation, hypoxia, infection, and cancer [50][51][52][53][54] , and to act as a extracellular effector in fungi and bacteria [55][56][57][58][59][60][61][62] . For recent reviews on GAPDH inhibitors and their multiple roles in several pharmacological applications see 63,64. In our study, HsGAPDH was purified from the supernatant of HEK293F cells treated with the mannosidase inhibitors kifunensine and kept for 4 days at 37°C after transfection with a DNA plasmid for recombinant expression of an endoplasmic reticulum glycoprotein.…”