Godoy Pablo, Darlington Peter John, Whiteway Malcolm
Centre of Structural and Functional Genomics, Biology Department, Concordia University - Loyola Campus, Montreal, QC, Canada.
Department of Pathology and Microbiology, Faculty of Veterinary Medicine, University of Montreal, Saint-Hyacinthe, QC, Canada.
Front Microbiol. 2022 Apr 5;13:833655. doi: 10.3389/fmicb.2022.833655. eCollection 2022.
, an important fungal pathogen of humans, displays different morphologies, such as yeast, pseudo-hyphae and hyphae, which are recognized unequally by phagocytic cells of the innate immune response. Once cells invade host tissues, immune cells such as macrophages are attracted to the site of infection and activated to recognize, engulf and kill the pathogen. We have investigated this fungal cell-macrophage interface by using high-throughput screening of the GRACE library to identify genes that can influence this interaction and modify the kinetics of engulfment. Compared with the wild-type (WT) strain, we identified generally faster rates of engulfment for those fungal strains with constitutive pseudo-hyphal and hyphal phenotypes, whereas yeast-form-locked strains showed a reduced and delayed recognition and internalization by macrophages. We identified a number of GRACE strains that showed normal morphological development but exhibited different recognition and engulfment kinetics by cultured macrophages and characterized two mutants that modified interactions with the murine and human-derived macrophages. One mutant inactivated an uncharacterized open reading frame that is the ortholog of , the other inactivated . The modified interaction was monitored during a 4 h co-culture. Early in the interaction, both and mutant strains showed reduced recognition and engulfment rates by macrophages when compared with WT cells. At fungal germ tube initiation, the engulfment kinetics increased for both mutants and WT cells, however the WT cells still showed a higher internalization by macrophages up to 2 h of interaction. Subsequently, between 2 and 4 h of the interaction, when most macrophages contain engulfed fungal cells, the engulfment kinetics increased for the mutant and further decreased for the mutant compared with Ca-WT. It appears that fungal morphology influences macrophage association with cells and that both and play roles in the interaction of the fungal cells with phagocytes.
是一种重要的人类真菌病原体，具有不同的形态，如酵母、假菌丝和菌丝，天然免疫反应的吞噬细胞对它们的识别程度不同。一旦细胞侵入宿主组织，巨噬细胞等免疫细胞就会被吸引到感染部位并被激活，以识别、吞噬和杀死病原体。我们通过对GRACE文库进行高通量筛选来研究这种真菌细胞与巨噬细胞的界面，以鉴定能够影响这种相互作用并改变吞噬动力学的基因。与野生型（WT）菌株相比，我们发现那些具有组成型假菌丝和菌丝表型的真菌菌株的吞噬速度通常更快，而锁定酵母形态的菌株则表现出巨噬细胞对其识别和内化的减少和延迟。我们鉴定出一些GRACE菌株，它们显示出正常的形态发育，但在培养的巨噬细胞中表现出不同的识别和吞噬动力学，并对两个与小鼠和人源巨噬细胞相互作用发生改变的突变体进行了表征。一个突变体使一个未表征的开放阅读框失活，该开放阅读框是 的直系同源物，另一个突变体使 失活。在4小时的共培养过程中监测改变后的相互作用。在相互作用早期，与WT细胞相比， 和 突变株均显示出巨噬细胞对其识别和吞噬率降低。在真菌芽管起始时，两个突变体和WT细胞的吞噬动力学均增加，然而在长达2小时的相互作用中，WT细胞仍显示出被巨噬细胞更高的内化率。随后，在相互作用进行到2至4小时之间，当大多数巨噬细胞含有吞噬的真菌细胞时，与Ca-WT相比， 突变体的吞噬动力学增加，而 突变体的吞噬动力学进一步降低。看来真菌形态会影响巨噬细胞与细胞的结合，并且 和 在真菌细胞与吞噬细胞的相互作用中都发挥作用。
