Malaria parasites have a complex life cycle, during which they undergo significant biological changes to adapt to different hosts and changing environments. Plasmodium falciparum, the species responsible for the deadliest form of human malaria, maintains this complex life cycle with a relatively small number of genes. Alternative splicing (AS) is an important post-transcriptional mechanisms that enables eukaryotic organisms to expand their protein repertoire out of relatively small number of genes. SR proteins are major regulators of AS in higher eukaryotes. Nevertheless, the regulation of splicing as well as the AS machinery in Plasmodium spp. are still elusive. Here, we show that PfSR1, a putative P. falciparum SR protein, can mediate RNA splicing in vitro. In addition, we show that PfSR1 functions as an AS factor in mini-gene in vivo systems similar to the mammalian SR protein SRSF1. Expression of PfSR1-myc in P. falciparum shows distinct patterns of cellular localization during intra erythrocytic development. Furthermore, we determine that the predicted RS domain of PfSR1 is essential for its localization to the nucleus. Finally, we demonstrate that proper regulation of pfsr1 is required for parasite proliferation in human RBCs and over-expression of pfsr1 influences AS activity of P. falciparum genes in vivo.