The protozoan parasite Plasmodium falciparum causes the most severe form of malaria in humans and is responsible for 200 million clinical cases and 600’000 deaths annually. Proliferating blood stage parasites, which are responsible for all malaria-related morbidity and mortality, use epigenetic control of gene expression as their prime strategy to secure their own survival and transmission. We have shown that the parasite ortholog of heterochromatin protein 1 (HP1), an evolutionary conserved regulator of heterochromatin formation and heritable silencing, plays a fundamental role in this strategy; PfHP1 provides the epigenetic framework for clonally variant expression of hundreds of genes implicated in parasite adaptation to changing environmental conditions. The hallmark example is antigenic variation of the major virulence factor PfEMP1, which is encoded by 60 var gene paralogs per parasite genome. PfHP1 silences transcription of all var genes except for one locus, which is activated in dedicated perinuclear zone believed to constitute a singular var expression site (VES). Switches in singular var activation lead to antigenic variation of PfEMP1. Through this sophisticated system, P. falciparum prevents premature exposure of the antigenic repertoire and secures continuous immune evasion and chronic infection. Our research focuses on elucidating the molecular mechanisms that determine and modulate the function of PfHP1 in heterochromatin formation and reversible gene silencing and activation. Whereas PfHP1’s role in heritable silencing is well established, the processes responsible for recruitment and local eviction of PfHP1, as well as the existence and molecular composition of the proposed VES remain unknown.

The research proposed here pursues two main objectives. Firstly, we will identify and characterise PfHP1 interaction partners and posttranslational modifications in vivo in order to understand the mechanisms responsible for the dynamic regulation of PfHP1 function. Secondly, we will gain novel insight into the elusive mechanism of mutually exclusive var gene transcription/singular var gene choice. To achieve this we will use complementary approaches and combine molecular biology, cell biology and biochemical techniques with state-of-the-art reverse genetics tools as well as high throughput approaches. Specifically, we will (1) experimentally confirm the interaction between PfHP1 and four potential PfHP1-interacting factors recently identified in our lab and characterise their role in heterochromatin formation and heritable silencing; (2) will investigate the in vivo role of PfHP1 phosphorylation and sumoylation in modulating PfHP1’s role in heterochromatin formation and heritable silencing; (3) we will test the hypothesis that the active state of var gene transcription is heritable via association with the VES in a PfHP1-independent manner; (4) we will attempt to identify and confirm protein components of the VES; and (5) we will identify and characterise a DNA-binding protein that was previously identified in our lab and has a potential role in singular var gene choice. The anticipated results will deliver an in-depth understanding of the epigenetic mechanisms responsible for phenotypic variation in P. falciparum, and in particular for antigenic variation of PfEMP1.