生物
无乳链球菌
基因
基因敲除
CRISPR干扰
清脆的
调解人
微生物学
细胞生物学
遗传学
链球菌
Cas9
细菌
作者
Kyle Firestone,Kathyayini P. Gopalakrishna,Lisa M. Rogers,Anju T. Peters,Jennifer A. Gaddy,Christine Nichols,Martica H. Hall,H Varela,Stephen M. Carlin,Gideon H. Hillebrand,EJ Giacobe,David M. Aronoff,Thomas A. Hooven
标识
DOI:10.1101/2024.12.06.627252
摘要
Abstract Group B Streptococcus (GBS; Streptococcus agalactiae ) is an important pathobiont capable of colonizing various host environments, contributing to severe perinatal infections. Surface proteins play critical roles in GBS-host interactions, yet comprehensive studies of these proteins’ functions have been limited by genetic manipulation challenges. This study leveraged a CRISPR interference (CRISPRi) library to target genes encoding surface-trafficked proteins in GBS, identifying their roles in modulating macrophage cytokine responses. Bioinformatic analysis of 654 GBS genomes revealed 66 conserved surface protein genes. Using a GBS strain expressing chromosomally integrated dCas9, we generated and validated CRISPRi strains targeting these genes. THP-1 macrophage-like cells were exposed to ethanol-killed GBS variants, and pro-inflammatory cytokines TNF-α and IL-1β were measured. Notably, knockdown of the sip gene, encoding the Surface Immunogenic Protein (Sip), significantly increased IL-1β secretion, implicating Sip in caspase-1-dependent regulation. Further, Δ sip mutants demonstrated impaired biofilm formation, reduced adherence to human fetal membranes, and diminished uterine persistence in a mouse colonization model. These findings suggest Sip modulates GBS- host interactions critical for pathogenesis, underscoring its potential as a therapeutic target or vaccine component.
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