Disruption of Plasmodium falciparum histidine-rich protein 2 may affect haem metabolism in the blood stage.
Disruption of Plasmodium falciparum histidine-rich protein 2 may affect haem metabolism in the blood stage.
复制标题
恶性疟原虫富含组氨酸的蛋白2的破坏可能会影响血液阶段的血红素代谢
DOI：
10.1186/s13071-020-04460-0
复制
发表时间：
2020-12-09
影响因子：
3.2
通讯作者：
Chen Q
中科院分区：
文献类型：
--
作者：
Yang Y;Tang T;Feng B;Li S;Hou N;Ma X;Jiang L;Xin X;Chen Q
背景
血红素是疟原虫生命周期中的一个关键代谢因子。在血液阶段，寄生虫获得宿主血红蛋白以产生用于蛋白质合成的氨基酸和用于代谢的副产物血红素。疟疾寄生虫也可以自行合成血红素。恶性疟原虫特异性富组氨酸蛋白2（PfHRP 2）具有血红素结合位点以介导血红素的形成，血红素是血红素聚集体的生物固化形式。值得注意的是，该基因调节机制的血红蛋白衍生血红素代谢和从头血红素生物合成途径在Pfhrp 2破坏的寄生虫线在红细胞内阶段。
方法
CRISPR/Cas9系统用于破坏Pfhrp 2的基因座。从转基因虫体中提取DNA，用PCR、Southern blotting和Western blotting等方法对转基因虫体进行鉴定。进行RNA测序和比较转录组分析以鉴定3D 7和Pfhrp 2 - 3D 7寄生虫之间的基因表达差异。
结果
通过CRISPR/Cas9系统成功建立Pfhrp 2转基因寄生虫。在6个对照组中分别鉴定了总共964、1261、3138、1064、2512和1778个差异表达基因（DEG），其中373、520、1499、353、1253和742个DEG上调，591、741、1639、711、分别有1259和1036例下调。在6个时间点（裂殖子侵入后0、8、16、24、32和40 h），在对照组中鉴定出5个与血红素代谢和合成相关的DEG。与血红素生物合成相关的δ-氨基乙酰丙酸合成酶和亚铁螯合酶的编码基因在对照组中被发现显著上调，而血红素加氧酶、基质加工肽酶和胆色素原脱氨酶的编码基因被发现显著下调。无GO术语在血液相关过程中显著富集（Q值= 1）。
结论
我们的数据揭示了Pfhrp 2 - 3D 7寄生虫在红细胞内阶段的转录组表达谱的变化。这些发现提供了基因转录水平的见解，将有助于进一步研究和开发抗疟疾药物。
图形摘要
Background
Haem is a key metabolic factor in the life cycle of the malaria parasite. In the blood stage, the parasite acquires host haemoglobin to generate amino acids for protein synthesis and the by-product haem for metabolic use. The malaria parasite can also synthesize haem de novo on its own. Plasmodium falciparum-specific histidine-rich protein 2 (PfHRP2) has a haem-binding site to mediate the formation of haemozoin, a biocrystallized form of haem aggregates. Notably, the gene regulates the mechanism of haemoglobin-derived haem metabolism and the de novo haem biosynthetic pathway in the Pfhrp2-disrupted parasite line during the intraerythrocytic stages.
Methods
The CRISPR/Cas9 system was used to disrupt the gene locus of Pfhrp2. DNA was extracted from the transgenic parasite, and PCR, Southern blotting and Western blotting were used to confirm the establishment of transgenic parasites. RNA-sequencing and comparative transcriptome analysis were performed to identify differences in gene expression between 3D7 and Pfhrp2--3D7 parasites.
Results
Pfhrp2- transgenic parasites were successfully established by the CRISPR/Cas9 system. A total of 964, 1261, 3138, 1064, 2512 and 1778 differentially expressed genes (DEGs) were identified in the six comparison groups, respectively, with 373, 520, 1499, 353, 1253 and 742 of these DEGs upregulated and 591, 741, 1639, 711, 1259 and 1036 of them downregulated, respectively. Five DEGs related to haem metabolism and synthesis were identified in the comparison groups at six time points (0, 8, 16, 24, 32, and 40 h after merozoite invasion). The genes encoding delta-aminolevulinic acid synthetase and ferrochelatase, both related to haem biosynthesis, were found to be significantly upregulated in the comparison groups, and those encoding haem oxygenase, stromal-processing peptidase and porphobilinogen deaminase were found to be significantly downregulated. No GO terms were significantly enriched in haem-related processes (Q value = 1).
Conclusion
Our data revealed changes in the transcriptome expression profile of the Pfhrp2--3D7 parasite during the intraerythrocytic stages. The findings provide insight at the gene transcript level that will facilitate further research on and development of anti-malaria drugs.
Graphical Abstract
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影响因子：
6.7
作者：
Nagaraj VA;Sundaram B;Varadarajan NM;Subramani PA;Kalappa DM;Ghosh SK;Padmanaban G
通讯作者：
Padmanaban G
影响因子：
48
作者：
Langmead, Ben;Salzberg, Steven L.
通讯作者：
Salzberg, Steven L.
影响因子：
46.9
作者：
Ghorbal, Mehdi;Gorman, Molly;Lopez-Rubio, Jose-Juan
通讯作者：
Lopez-Rubio, Jose-Juan
影响因子：
7.7
作者：
Sigala PA;Crowley JR;Henderson JP;Goldberg DE
通讯作者：
Goldberg DE
影响因子：
56.9
作者：
Jinek, Martin;Chylinski, Krzysztof;Charpentier, Emmanuelle
通讯作者：
Charpentier, Emmanuelle