Division of Infectious Diseases, Boston Children's Hospital/Harvard Medical School, Boston, Massachusetts, United States of America.
Center for Genomic Science Innovation, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
PLoS Genet. 2024 Aug 13;20(8):e1011156. doi: 10.1371/journal.pgen.1011156. eCollection 2024 Aug.
Phosphorus is essential in all cells' structural, metabolic and regulatory functions. For fungal cells that import inorganic phosphate (Pi) up a steep concentration gradient, surface Pi transporters are critical capacitators of growth. Fungi must deploy Pi transporters that enable optimal Pi uptake in pH and Pi concentration ranges prevalent in their environments. Single, triple and quadruple mutants were used to characterize the four Pi transporters we identified for the human fungal pathogen Candida albicans, which must adapt to alkaline conditions during invasion of the host bloodstream and deep organs. A high-affinity Pi transporter, Pho84, was most efficient across the widest pH range while another, Pho89, showed high-affinity characteristics only within one pH unit of neutral. Two low-affinity Pi transporters, Pho87 and Fgr2, were active only in acidic conditions. Only Pho84 among the Pi transporters was clearly required in previously identified Pi-related functions including Target of Rapamycin Complex 1 signaling, oxidative stress resistance and hyphal growth. We used in vitro evolution and whole genome sequencing as an unbiased forward genetic approach to probe adaptation to prolonged Pi scarcity of two quadruple mutant lineages lacking all 4 Pi transporters. Lineage-specific genomic changes corresponded to divergent success of the two lineages in fitness recovery during Pi limitation. Initial, large-scale genomic alterations like aneuploidies and loss of heterozygosity eventually resolved, as populations gained small-scale mutations. Severity of some phenotypes linked to Pi starvation, like cell wall stress hypersensitivity, decreased in parallel to evolving populations' fitness recovery in Pi scarcity, while severity of others like membrane stress responses diverged from Pi scarcity fitness. Among preliminary candidate genes for contributors to fitness recovery, those with links to TORC1 were overrepresented. Since Pi homeostasis differs substantially between fungi and humans, adaptive processes to Pi deprivation may harbor small-molecule targets that impact fungal growth, stress resistance and virulence.
磷是所有细胞结构、代谢和调节功能所必需的。对于从无机磷酸盐（Pi）中导入细胞内的真菌细胞，表面 Pi 转运蛋白是生长的关键能力者。真菌必须部署能够在其环境中普遍存在的 pH 值和 Pi 浓度范围内实现最佳 Pi 摄取的 Pi 转运蛋白。我们使用单突变体、三突变体和四突变体来表征我们为人类真菌病原体白色念珠菌鉴定的四种 Pi 转运蛋白，这些蛋白必须适应宿主血液和深部器官入侵期间的碱性条件。一种高亲和力 Pi 转运蛋白 Pho84 在最宽的 pH 范围内效率最高，而另一种 Pho89 仅在中性 pH 单位内具有高亲和力特征。两种低亲和力 Pi 转运蛋白 Pho87 和 Fgr2 仅在酸性条件下活跃。在先前确定的与 Pi 相关的功能中，只有 Pi 转运蛋白 Pho84 明显是必需的，包括雷帕霉素靶蛋白复合物 1 信号、氧化应激抗性和菌丝生长。我们使用体外进化和全基因组测序作为一种无偏见的正向遗传方法，来探测两种缺乏所有 4 种 Pi 转运蛋白的四突变体系对长时间 Pi 缺乏的适应性。谱系特异性基因组变化与在 Pi 限制条件下两种谱系适应性恢复的不同成功相对应。最初，像非整倍体和杂合性丧失这样的大规模基因组改变最终得到解决，因为种群获得了小规模的突变。与 Pi 饥饿相关的一些表型的严重程度，如细胞壁应激超敏性，与在 Pi 匮乏时进化种群的适应性恢复平行降低，而其他表型如膜应激反应与 Pi 匮乏适应性不同。在适应 Pi 剥夺的初步候选基因中，与 TORC1 相关的基因数量过多。由于真菌和人类之间的 Pi 动态平衡有很大差异，因此 Pi 剥夺的适应过程可能具有影响真菌生长、应激抗性和毒力的小分子靶标。
