Functional analysis of N-terminally tagged Rab5a. (a–c) Cell lines comparing KS (a) and KS in the presence of a complementing functional copy of Rab5a (b) and after gene excision (c). Left, representative fluorescence microscopy images. Three different GFP-2×FKBP-Rab5a parasite lines were generated to perform KS with a nuclear mislocalizer (cell line 1), KS in the presence of a complementary copy of Rab5a (cell line 2) or excision of the functional gene with inducible Cre recombinase9–11 (cell line 3). After addition of rapalog to cell line 1, GFP-2×FKBP-Rab5a was efficiently mislocalized into the nucleus, and no detectable protein remained in the cytoplasm (a). In contrast, the second cell line, which expressed a complementary copy of Rab5a in addition to GFP-2×FKBP-Rab5a, grew normally after rapalog addition (b). In the third cell line, rapalog addition to dimerize split Cre led to excision of the functional copy of rab5a and to a phenotype comparable (c).
Birnbaum J, Flemming S, Reichard N, Soares AB, Mesén-Ramírez P, Jonscher E, Bergmann B, Spielmann T. A genetic system to study Plasmodium falciparum protein function. Nat Methods. 2017 Mar 13. PMID: 28288121