Expression of the Plasmodium berghei actin II gene is controlled by elements in a long genomic region

Plasmodium parasites have two actin isoforms. Actin I is ubiquitously expressed, while the second actin isoform is expressed in the sexual stages and ookinetes. Reverse genetic analysis revealed two phenotypes in parasites lacking the protein: a block in male gametogenesis (exflagellation) and a second phenotype in oocyst development, dependent upon the expression of the gene in female gametocytes. Here, we report that the genetic complementation of two independent mutants lacking actin II does not fully restore wild-type function. Constructs were integrated in the c-rrna locus, previously used for expression of transgenes, in order to determine the dependence of expression on actin II flanking genomic regions. Partial restoration of male gametogenesis was achieved when the transgene contained, in addition to the coding region, 1.2 kb upstream of the actin II open reading frame. Another transgene, which comprised 2.7 kb of actin II 5′ flanking regions and the cognate 3′ downstream sequence, fully restored exflagellation. However, in both complemented strains, oocyst development was severely impaired compared to the WT. These data suggest that male gametocyte expression of actin II is dependent upon extensive flanking regions, while female expression requires even longer genomic sequences for correct expression of the gene.