PfSWIB, a potential chromatin regulator for var gene regulation and parasite development in Plasmodium falciparum

Wei-Feng Wang, Yi-Long Zhang

Various transcription factors are involved in the process of mutually exclusive expression and clonal variation of the Plasmodium multigene (var) family. Recent studies revealed that a P. falciparum SWI/SNF-related matrix-associated actin-dependent regulator of chromatin (PfSWIB) might trigger stage-specific programmed cell death (PCD), and was not only crucial for the survival and development of parasite, but also had profound effects on the parasite by interacting with other unknown proteins. However, it remains unclear whether PfSIWB is involved in transcriptional regulation of this virulence gene and its functional properties. A conditional knockdown system "PfSWIB-FKBP-LID" was introduced to the parasite clone 3D7, and an integrated parasite line "PfSWIB-HA-FKBP-LID" was obtained by drug cycling and clone screening. Growth curve analysis (GCA) was performed to investigate the growth and development of different parasite lines during 96 h in vitro culturing, by assessing parasitemia. Finally, we performed qPCR assays to detect var gene expression profiling in various comparison groups, as well as the mutually exclusive expression pattern of the var genes within a single 48 h life-cycle of P. falciparum in different parasite lines. In addition, RNA-seq was applied to analyze the var gene expression in different lines. GCA revealed that conditional knockdown of PfSWIB could interfere with the growth and development of P. falciparum. The parasitemia of PfSWIB∆ showed a significant decline at 96 h during in vitro culture compared with the PfSWIB and 3D7 lines (P < 0.0001). qPCR and RNA-seq analysis confirmed that depletion of PfSWIB not only silences upsA, upsC and partial upsB var genes, as well as removes the silencing of partial upsB var genes at the ring stage in PfSWIB∆ line, but also leads to aberrant expression of upsA and partial upsB/upsC var genes at the mature stage of P. falciparum, during a single 48-h life-cycle. We demonstrated that PfSWIB was involved in the process of clonal variation in var gene expression, and crucial for the survival and development of Plasmodium parasite. These findings could provide better understanding of the mechanism and function of PfSWIB contributing to the pathogenesis in malaria parasites.