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Bifidobacterium breve UCC2003 Employs Multiple Transcriptional Regulators To Control Metabolism of Particular Human Milk Oligosaccharides.

ABSTRACT: Bifidobacterial carbohydrate metabolism has been studied in considerable detail for a variety of both plant- and human-derived glycans, particularly involving the bifidobacterial prototype strain Bifidobacterium breve UCC2003. We recently elucidated the metabolic pathways by which the human milk oligosaccharide (HMO) constituents lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT) and lacto-N-biose (LNB) are utilized by B. breve UCC2003. However, to date, no work has been carried out on the regulatory mechanisms that control the expression of the genetic loci involved in these HMO metabolic pathways. In this study, we describe the characterization of three transcriptional regulators and the corresponding operator and associated (inducible) promoter sequences, with the latter governing the transcription of the genetic elements involved in LN(n)T/LNB metabolism. The activity of these regulators is dependent on the release of specific monosaccharides, which are believed to act as allosteric effectors and which are derived from the corresponding HMOs targeted by the particular locus.IMPORTANCE Human milk oligosaccharides (HMOs) are a key factor in the development of the breastfed-infant microbiota. They function as prebiotics, selecting for a specific range of microbes, including a number of infant-associated species of bifidobacteria, which are thought to provide a range of health benefits to the infant host. While much research has been carried out on elucidating the mechanisms of HMO metabolism in infant-associated bifidobacteria, to date there is very little understanding of the transcriptional regulation of these pathways. This study reveals a multicomponent transcriptional regulation system that controls the recently identified pathways of HMO metabolism in the infant-associated Bifidobacterium breve prototype strain UCC2003. This not only provides insight into the regulatory mechanisms present in other infant-associated bifidobacteria but also provides an example of a network of sequential steps regulating microbial carbohydrate metabolism.

SUBMITTER: James K

PROVIDER: S-EPMC5930313 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Bifidobacterium breve UCC2003 Employs Multiple Transcriptional Regulators To Control Metabolism of Particular Human Milk Oligosaccharides.

James Kieran K O'Connell Motherway Mary M Penno Christophe C O'Brien Rebecca Louise RL van Sinderen Douwe D

Applied and environmental microbiology 20180416 9

Bifidobacterial carbohydrate metabolism has been studied in considerable detail for a variety of both plant- and human-derived glycans, particularly involving the bifidobacterial prototype strain Bifidobacterium breve UCC2003. We recently elucidated the metabolic pathways by which the human milk oligosaccharide (HMO) constituents lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT) and lacto-N-biose (LNB) are utilized by B. breve UCC2003. How ...[more]

PMID: 29500268

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Project description:Members of the genus Bifidobacterium are common inhabitants of the gastrointestinal tract of humans and other mammals, where they ferment many diet-derived carbohydrates that cannot be digested by their host. To extend our understanding of bifidobacterial carbohydrate utilisation, we investigated the molecular mechanisms by which various strains of Bifidobacterium breve metabolize four distinct α-glucose and/or α-galactose-containing oligosaccharides, namely raffinose, stachyose, melibiose and melezitose. Here we demonstrate that all B. breve strains examined possess the ability to utilise raffinose, stachyose and melibiose. However, the ability to metabolize melezitose was not ubiquitous for all tested B. breve strains. Transcriptomic and functional genomic approaches identified a gene cluster dedicated to the metabolism of α-galactose-containing carbohydrates, while an adjacent gene cluster, dedicated to the metabolism of α-glucose-containing melezitose, was identified, yet being present only in those B. breve strains that were able to support growth on this carbohydrate. DNA-microarrays containing oligonucleotide primers representing each of the 1864 annotated genes on the genome of B. breve UCC2003 (O'Connell Motherway et al., 2011) were designed by and obtained from Agilent Technologies (Palo Alto, Ca., USA). Methods for cell disruption, RNA isolation, RNA quality control, complementary DNA synthesis and labeling were performed as described previously (Pokusaeva et al., 2009). Labeled cDNA was hybridized using the Agilent Gene Expression hybridization kit (part number 5188-5242) as described in the Agilent Two-Color Microarray-Based Gene Expression Analysis v4.0 manual (G4140-90050). Following hybridization, microarrays were washed in accordance with Agilent’s standard procedures and scanned using an Agilent DNA microarray scanner (model G2565A). Generated scans were converted to data files with Agilent's Feature Extraction software (Version 9.5). DNA-microarray data were processed as previously described (Garcia De La Nava et al., 2003). Differential expression tests were performed with the Cyber-T implementation of a variant of the t-test (Long et al., 2001). A gene was considered differentially expressed when p < 0.001 and an expression ratio of >3 or <0.33 relative to the control.

Dataset Information

Bifidobacterium breve UCC2003 Employs Multiple Transcriptional Regulators To Control Metabolism of Particular Human Milk Oligosaccharides.

Publications

Bifidobacterium breve UCC2003 Employs Multiple Transcriptional Regulators To Control Metabolism of Particular Human Milk Oligosaccharides.

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