Sexton Danielle L, Burgold Steffen, Schertel Andreas, Tocheva Elitza I
Department of Microbiology and Immunology, University of British Columbia, Vancouver, Canada.
ZEISS Microscopy Customer Center, Oberkochen, Germany.
Curr Res Struct Biol. 2021 Dec 13;4:1-9. doi: 10.1016/j.crstbi.2021.12.001. eCollection 2022.
Studying bacterial cell envelope architecture with electron microscopy is challenging due to the poor preservation of microbial ultrastructure with traditional methods. Here, we established and validated a super-resolution cryo-correlative light and electron microscopy (cryo-CLEM) method, and combined it with cryo-focused ion beam (cryo-FIB) milling and scanning electron microscopy (SEM) volume imaging to structurally characterize the bacterium . Subsequent cryo-electron tomography (cryo-ET) revealed an unusual diderm cell envelope architecture with a thick layer of peptidoglycan (PG) between the inner and outer membranes, an additional periplasmic layer, and a proteinaceous surface S-layer. Cells grew in tetrads, and division septa were formed by invagination of the inner membrane (IM), followed by a thick layer of PG. Cytoskeletal filaments, FtsA and FtsZ, were observed at the leading edges of constricting septa. Numerous macromolecular complexes were found associated with the cytoplasmic side of the IM. Altogether, our study revealed several unique ultrastructural features of cells, opening new lines of investigation into the physiology and evolution of the bacterium.
由于传统方法对微生物超微结构的保存不佳，利用电子显微镜研究细菌细胞包膜结构具有挑战性。在此，我们建立并验证了一种超分辨率冷冻关联光电子显微镜（cryo-CLEM）方法，并将其与冷冻聚焦离子束（cryo-FIB）铣削和扫描电子显微镜（SEM）体积成像相结合，以对该细菌进行结构表征。随后的冷冻电子断层扫描（cryo-ET）揭示了一种不寻常的双膜细胞包膜结构，在内膜和外膜之间有一层厚厚的肽聚糖（PG）、一个额外的周质层和一个蛋白质表面S层。细胞以四联形式生长，分裂隔膜由内膜（IM）内陷形成，随后是一层厚厚的PG。在收缩隔膜的前沿观察到细胞骨架丝、FtsA和FtsZ。发现许多大分子复合物与IM的细胞质侧相关联。总之，我们的研究揭示了该细胞的几个独特超微结构特征，为该细菌的生理学和进化研究开辟了新的途径。
