Mutant/mutation
The mutant expresses an N-terminal GFP-tagged version of P. berghei Sec61β. The transgene is integrated into the silent c/d-type rrna gene unit under control of the constitutive eef1a promoter. Sec61β is a known marker protein for endoplasmic reticulum (ER).

Two transgenes have been generated: one with Sec61β tagged with GFP and one with superfolder GFP (sfGFP). The sfGFP offers several advantages over GFP for the tagging of ER transmembrane proteins such as Sec61β. GFP has the tendency to weakly dimerize. When this occurs at the cytoplasmic face of the ER membrane, it can induce artificial reorganisation of the entire ER architecture, resulting in huge ER patches known as organised smooth ER. In contrast, sfGFP is a purely monomeric version of GFP that was designed to exhibit improved folding capacities and to be more resistant to denaturation. In this study no differences were observed in the localisation of the two fusion proteins.

Protein (function)
Sec61β is a known marker protein for endoplasmic reticulum (ER).
Sec61β, together with Sec61α and Sec61γ, builds the Sec61 complex, which forms a channel through the ER membrane and is central to co- and post-translational protein translocation. Membrane-bound ribosomes are known to be tightly associated with the Sec61 complex to facilitate co-translational protein transport.

Phenotype
Expression of GFP-Sec61β in cytoplasm of most life cycle stages. In this study expression of GFP-Sec61β in cytoplasm of liver stages is studied in detail.
Live cell imaging of PbcsfGFP-Sec61β parasites showed that during liver stage schizogony, the parasite ER closely surrounds each parasite nucleus and additionally builds a network of ER membranes that extend through the entire cytoplasm. The parasite ER seems to remain as a single interconnected organelle until very shortly before merozoite formation. In merozoites, only the perinuclear ER is present.
It was observed that the parasite ER, in addition to being perinuclear and existing as network-like structures, also forms extensive ER accumulations, which start to appear in trophozoites and become most prominent during the cytomere stage. Most of the ER accumulations disappear during the formation of individual merozoites and accumulations are completely absent at the end of liver stage development when merozoites have formed.

Additional information

Other mutants