The transcriptional regulator Fur modulates the expression of uge, a gene essential for the core lipopolysaccharide biosynthesis in Klebsiella pneumoniae

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Muner, Jose Julio ; de Oliveira, Paloma Aparecida Alves ; Baboghlian, Juliana ; Moura, Stefany Casarin ; de Andrade, Abissair Gabriel ; de Oliveira, Michelly Macedo ; Campos, Yasmin Ferreira de ; Mancano, Alquiandra Stefani Ferreira ; Siqueira, Nathalia Maria Goncalves ; Pacheco, Thaisy ; Ferraz, Lucio Fabio Caldas

• Tipo de documento: Artigo Científico
• Fonte: BMC Microbiology; v. 24, n. 1, p. 9-pg., 2024-07-27.

Resumo

Background Klebsiella pneumoniae is a Gram-negative pathogen that has become a threat to public health worldwide due to the emergence of hypervirulent and multidrug-resistant strains. Cell-surface components, such as polysaccharide capsules, fimbriae, and lipopolysaccharides (LPS), are among the major virulence factors for K. pneumoniae. One of the genes involved in LPS biosynthesis is the uge gene, which encodes the uridine diphosphate galacturonate 4-epimerase enzyme. Although essential for the LPS formation in K. pneumoniae, little is known about the mechanisms that regulate the expression of uge. Ferric uptake regulator (Fur) is an iron-responsive transcription factor that modulates the expression of capsular and fimbrial genes, but its role in LPS expression has not yet been identified. This work aimed to investigate the role of the Fur regulator in the expression of the K. pneumoniae uge gene and to determine whether the production of LPS by K. pneumoniae is modulated by the iron levels available to the bacterium. Results Using bioinformatic analyses, a Fur-binding site was identified on the promoter region of the uge gene; this binding site was validated experimentally through Fur Titration Assay (FURTA) and DNA Electrophoretic Mobility Shift Assay (EMSA) techniques. RT-qPCR analyses were used to evaluate the expression of uge according to the iron levels available to the bacterium. The iron-rich condition led to a down-regulation of uge, while the iron-restricted condition resulted in up-regulation. In addition, LPS was extracted and quantified on K. pneumoniae cells subjected to iron-replete and iron-limited conditions. The iron-limited condition increased the amount of LPS produced by K. pneumoniae. Finally, the expression levels of uge and the amount of the LPS were evaluated on a K. pneumoniae strain mutant for the fur gene. Compared to the wild-type, the strain with the fur gene knocked out presented a lower LPS amount and an unchanged expression of uge, regardless of the iron levels. Conclusions Here, we show that iron deprivation led the K. pneumoniae cells to produce higher amount of LPS and that the Fur regulator modulates the expression of uge, a gene essential for LPS biosynthesis. Thus, our results indicate that iron availability modulates the LPS biosynthesis in K. pneumoniae through a Fur-dependent mechanism. (AU)

• Processo FAPESP: 08/11365-1 - Estudo do mecanismo de regulação da expressão gênica pelo regulador Fur em Klebsiella pneumoniae: implicações na expressão de fatores de virulência mediada por quorum-sensing
• Beneficiário: Lúcio Fábio Caldas Ferraz
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 13/13949-9 - Caracterização genotípica e fenotípica de Klebsiella pneumoniae proveniente de infecções do trato urinário e análise in vitro dos processos de adesão e invasão de células hospedeiras
• Beneficiário: Thaisy Eliza Pacheco dos Santos
• Modalidade de apoio: Bolsas no Brasil - Mestrado
• Processo FAPESP: 13/06042-7 - Estudo in vivo e in vitro dos processos de adesão e invasão de células hospedeiras por linhagens de Klebsiella pneumoniae mutantes para a expressão de fimbrias
• Beneficiário: Lúcio Fábio Caldas Ferraz
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 18/26203-9 - Estudo da contribuição das vias de síntese e de transporte de poliaminas para a patogenicidade de Klebsiella pneumoniae
• Beneficiário: Lúcio Fábio Caldas Ferraz
• Modalidade de apoio: Auxílio à Pesquisa - Regular