Department of Microbiology and Immunology, The Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire, USA.
Department of Microbiology and Immunology, The Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire, USA
mSphere. 2021 Apr 21;6(2):e00182-21. doi: 10.1128/mSphere.00182-21.
causes a chronic infection that renders the immunocompromised human host susceptible to toxoplasmic encephalitis triggered by cyst reactivation in the central nervous system. The dense granule protein GRA12 is a major parasite virulence factor required for parasite survival during acute infection. Here, we characterized the role of four GRA12-related genes in acute and chronic stages of infection. While GRA12A, GRA12B, and GRA12D were highly expressed in asexual stage tachyzoites and bradyzoites, expression of GRA12C appeared to be restricted to the sexual stages. In contrast to deletion of GRA12 (Δ), no major defects in acute virulence were observed in Δ, Δ, or Δg parasites, though Δ parasites exhibited an increased tachyzoite replication rate. Bradyzoites secreted GRA12A, GRA12B, and GRA12D and incorporated these molecules into the developing cyst wall, as well as the cyst matrix in distinct patterns. Similar to GRA12, GRA12A, GRA12B, and GRA12D colocalized with the dense granules in extracellular tachyzoites, with GRA2 and the intravacuolar network in the tachyzoite stage parasitophorous vacuole and with GRA2 in the cyst matrix and cyst wall. Chronic stage cyst burdens were decreased in mice infected with Δ parasites and were increased in mice infected with Δ parasites. However, Δ cysts were not efficiently maintained Δ, Δ, and Δ cysts displayed a reduced reactivation efficiency, and reactivation of Δ cysts was delayed. Collectively, our results suggest that a family of genes related to GRA12 play significant roles in the formation, maintenance, and reactivation of chronic stage cysts. If host immunity weakens, cysts recrudesce in the central nervous system and cause a severe toxoplasmic encephalitis. Current therapies target acute stage infection but do not eliminate chronic cysts. Parasite molecules that mediate the development and persistence of chronic infection are poorly characterized. Dense granule (GRA) proteins such as GRA12 are key virulence factors during acute infection. Here, we investigated four GRA12-related genes. GRA12-related genes were not major virulence factors during acute infection. Instead, GRA12-related proteins localized at the cyst wall and cyst matrix and played significant roles in cyst development, persistence, and reactivation during chronic infection. Similar to GRA12, the GRA12-related proteins selectively associated with the intravacuolar network of membranes inside the vacuole. Collectively, our results support the hypothesis that GRA12 proteins associated with the intravacuolar membrane system support parasite virulence during acute infection and cyst development, persistence, and reactivation during chronic infection.
弓形虫引起慢性感染，使免疫功能低下的人类宿主易患中枢神经系统中囊泡再激活引发的弓形体脑炎。致密颗粒蛋白 GRA12 是寄生虫在急性感染过程中存活所必需的主要寄生虫毒力因子。在这里，我们描述了四个与 GRA12 相关的基因在感染的急性和慢性阶段的作用。虽然 GRA12A、GRA12B 和 GRA12D 在无性阶段速殖子和缓殖子中高度表达，但 GRA12C 的表达似乎仅限于有性阶段。与 GRA12Δ（缺失）相比，Δ、Δ和Δg 寄生虫在急性毒力方面没有明显缺陷，尽管Δ 寄生虫的速殖子复制率增加。缓殖子分泌 GRA12A、GRA12B 和 GRA12D，并将这些分子整合到发育中的囊泡壁和囊泡基质中，呈现出不同的模式。与 GRA12 相似，GRA12A、GRA12B 和 GRA12D 与细胞外速殖子中的致密颗粒共定位，与速殖子阶段的囊泡内空泡网络中的 GRA2 以及囊泡基质和囊壁中的 GRA2 共定位。感染 GRA12Δ 寄生虫的小鼠慢性阶段囊泡负荷减少，感染 GRA12Δ 寄生虫的小鼠囊泡负荷增加。然而，Δ 囊泡不能有效地维持Δ、Δ和Δ 囊泡的再激活效率降低，并且 Δ 囊泡的再激活延迟。总的来说，我们的结果表明，与 GRA12 相关的一组基因在慢性阶段囊泡的形成、维持和再激活中发挥重要作用。如果宿主免疫力减弱，囊泡在中枢神经系统中重新出现，导致严重的弓形体脑炎。目前的治疗方法针对急性感染阶段，但不能消除慢性囊泡。介导慢性感染发展和持续的寄生虫分子特征描述较差。致密颗粒（GRA）蛋白，如 GRA12，是急性感染期间的关键毒力因子。在这里，我们研究了四个与 GRA12 相关的基因。与 GRA12 相关的基因在急性感染期间不是主要的毒力因子。相反，GRA12 相关蛋白定位于囊泡壁和囊泡基质，在慢性感染期间对囊泡发育、维持和再激活具有重要作用。与 GRA12 相似，GRA12 相关蛋白选择性地与囊泡内空泡网络中的膜结合。总的来说，我们的结果支持这样的假设，即与囊泡内膜系统相关的 GRA12 蛋白支持寄生虫在急性感染期间的毒力以及在慢性感染期间的囊泡发育、维持和再激活。
