Plasmodium falciparum leucine-rich repeat 5 disruption alters the transcription progression during asexual and sexual stage development.

How Plasmodium falciparum responds to stresses such as antimalarial drug pressure and uses them to trigger developmental decisions such as the production of sexual gametocytes remains poorly understood. Here, we investigated the function of the leucine-rich repeat protein LRR5 (PF3D7_1432400). Previous genome-wide piggyBac mutagenesis established that disruptions of LRR5 increase sensitivity to artemisinin and heat-shock and enhance gametocyte production. Using an optimized single-cell RNA sequencing approach applied to unsynchronized parasite cultures, we profiled transcriptional heterogeneity across the intraerythrocytic developmental cycle in wild-type NF54 parasites and an isogenic LRR5 piggyBac mutant (LRR5pB). Disruption of LRR5 reshaped the parasite transcriptome across multiple stages, with marked downregulation of pathways involved in protein folding, protein export, and host-cell remodeling, including Maurer's cleft components, the PTEX export machinery, and the exported chaperone HSP70-x. In contrast, cytoplasmic heat-shock proteins and translation-associated genes were upregulated, suggesting a compensatory stress response. Notably, LRR5pB parasites displayed premature activation of gametocyte-associated transcriptional programs, including elevated expression of the early gametocyte marker Pfs16, which was confirmed at the protein level. Pseudotime analysis further revealed accelerated transcriptional progression with mistimed expression of trophozoite-, merozoite-, and gametocyte-associated gene clusters. Together, these findings suggest LRR5 as a link between proteostasis, host-cell remodeling, and developmental timing in P. falciparum.IMPORTANCEMalaria parasites must survive environmental stress while deciding whether to continue asexual replication in the blood stream or instead commit to sexual gametocytes for transmission. Understanding how these processes are coordinated is critical for developing new strategies to block both disease and transmission. In this study, we used single-cell transcriptomics to examine how disruption of the P. falciparum protein LRR5 alters parasite development. By analyzing individual parasites from unsynchronized cultures, we uncovered hidden changes in stress-response pathways, host-cell remodeling, and early activation of genes linked to sexual development. Our findings show that loss of LRR5 weakens the parasite's ability to manage stress while prematurely priming parasites for gametocyte formation. These results suggest LRR5 as a link to stress adaptation to transmission potential.