Critchlow Jeanette M, Rocchio Joseph S, McKell Melanie C, Campbell Courtney J, Barraza Juan P, Krystofiak Evan S, Green Erin R, Akizuki Tae, Chazin Walter J, VanNieuwenhze Michael S, Stemmler Timothy L, Giedroc David P, Skaar Eric P
Department of Pathology, Microbiology, and Immunology, and Vanderbilt Institute for Infection, Immunology, and Inflammation, Vanderbilt University Medical Center, Nashville, Tennessee, United States of America.
Department of Chemistry, Indiana University, Bloomington, Indiana, United States of America.
PLoS Pathog. 2025 Jun 16;21(6):e1013209. doi: 10.1371/journal.ppat.1013209. eCollection 2025 Jun.
To colonize and survive in the host, bacterial pathogens like Acinetobacter baumannii must acquire zinc (Zn). To maintain Zn homeostasis, A. baumannii synthesizes proteins of the COG0523 family which are predicted to chaperone Zn to metalloproteins. Bioinformatic tools identified A. baumannii A1S_0934 as a COG0523 protein, and yeast two-hybrid screening revealed that MurD, an essential muramyl ligase, interacts with A1S_0934. As such, we have named A1S_0934 MurD interacting GTPase COG0523 (MigC). Here we show that MigC is a GTPase whose activity is stimulated upon Zn coordination to a characteristic CxCC (C = Cys; x = Leu/Ile/Met) motif to form a S3(N/O) complex. MigC-deficient strains (ΔmigC) display sensitivity to Zn depletion and exhibit altered cell wall architecture in vitro. Biochemical and functional assays confirm the MigC-MurD interaction, which inhibits the catalytic activity of MurD. CRISPRi knockdowns of murD reduce A. baumannii fitness and increase filamentation during Zn depletion, a phenotype reversed in ΔmigC strains, suggesting that MigC also inhibits MurD activity in cells. ΔmigC cells are elongated and sensitized to ceftriaxone, a cephalosporin antibiotic, consistent with decreased cell wall integrity. The ΔmigC strain has reduced ability to colonize in a murine model of pneumonia highlighting the importance of the MigC-MurD interaction induced by A. baumannii infection. Together these data suggest that MigC impacts cell wall biogenesis, in part through interactions with MurD, emphasizing the importance of MigC and MurD to the survival and pathogenicity of A. baumannii while expanding the potential functions of the COG0523 family of enzymes.
为了在宿主体内定殖并存活，像鲍曼不动杆菌这样的细菌病原体必须获取锌（Zn）。为维持锌稳态，鲍曼不动杆菌会合成COG0523家族的蛋白质，预计这些蛋白质会将锌转运至金属蛋白。生物信息学工具将鲍曼不动杆菌A1S_0934鉴定为一种COG0523蛋白，酵母双杂交筛选显示，必需的胞壁酰连接酶MurD与A1S_0934相互作用。因此，我们将A1S_0934命名为MurD相互作用GTP酶COG0523（MigC）。在此我们表明，MigC是一种GTP酶，当锌与特征性的CxCC（C = 半胱氨酸；x = 亮氨酸/异亮氨酸/甲硫氨酸）基序配位形成S3（N/O）复合物时，其活性会受到刺激。MigC缺陷菌株（ΔmigC）对锌耗竭敏感，并且在体外表现出细胞壁结构改变。生化和功能分析证实了MigC与MurD的相互作用，这种相互作用会抑制MurD的催化活性。murD的CRISPRi敲低会降低鲍曼不动杆菌的适应性，并在锌耗竭期间增加丝状化，这一表型在ΔmigC菌株中得到逆转，表明MigC在细胞中也会抑制MurD的活性。ΔmigC细胞会伸长并对头孢曲松（一种头孢菌素类抗生素）敏感，这与细胞壁完整性降低一致。ΔmigC菌株在肺炎小鼠模型中的定殖能力降低，突出了鲍曼不动杆菌感染诱导的MigC-MurD相互作用的重要性。这些数据共同表明，MigC部分通过与MurD相互作用影响细胞壁生物合成，强调了MigC和MurD对鲍曼不动杆菌存活和致病性的重要性，同时扩展了COG0523酶家族的潜在功能。
