Mutant/mutation
The mutant is a 'Flp/FRT conditional knock-out mutant' of Atg7. The mutant expresses the yeast FlpL recombinase under the control of the trap promoter  and contains a FRTed (part of the) ORF (open reading frame) of the Atg locus. 

This mutant has been generated by replacement of the endogenous Atg7 gene by an Atg7 gene with a FRTed ORF in mutant RMgm-4186 that expresses FlpL. This mutant expresses the yeast FlpL recombinase under the control of the promoter of trap and does not contain a selectable marker. The flpl gene is integrated into the silent 230p locus by double cross-over integration.

(Part of)The Atg7 ORF is removed from the genome by using the Flp/FRT site-specific recombination (SSR) system (see RMgm-269, RMgm-747). Removal of (part of) the FRTed ORF of Atg7-cKO has been achieved by  transmission of the mutant through mosquitoes, thereby activating expression of the FlpL recombinase in sporozoites that resulted in the excision of the ORF of Atg7 which was flanked by FRT sequences.

Published in: bioRxiv preprint doi: https://doi.org/10.1101/2023.08.16.553492

Protein (function)
Autophagy-related E1-like enzyme Atg7.
Atg8, a marker of active autophagy in yeast and mammals, is a small ubiquitin-like molecule, and its conjugation with phosphatidyl ethanolamine (PE) on the autophagosome membrane is essential for cargo selection, membrane tethering, hemifusion, expansion, and closure of autophagosomes. In apicomplexan parasites, Atg8 is conserved along with the intermediate genes required for its conjugation with PE and is specifically localized on the apicoplast membrane. The discrete localization of Atg8 on the apicoplast membrane throughout the parasite life cycle must have a distinct role from its role in autophagy.
In this study evidence is presented for the role of Atg7 in Atg8 conjugation, clearance of micronemes, organelle biogenesis, and expansion of parasites during liver stage development

Phenotype
Normal development of Atg7-cKO sporozoites.
All  mice inoculated with wildtype sporozoites became patent on day 3 p.i., whereas parasites lacking Atg7 (Atg7cKO) failed to initiate blood-stage infection.
Comparable parasite burden in livers in mice infected with Atg7 cKO and wildtype sporozoites until 38 hours post infection (hpi), which which was decreased in late time point samples harvested at 55 hpi. In culture Atg7 cKO sporozoites invaded hepatocytes normally. Immunofluorescence analysis of exo-erythrocytic forms (EEFs) with Hsp70 and UIS4 antibodies followed by quantification revealed normal development until 24 hpi. However, EEFs harvested at 40, 55 and 64 hpi showed arrested growth and a reduction in size and number. A strong defect in nuclear division in Atg7 cKO parasites.

Additional information
To check the excision efficiency of the Atg7 gene in sporozoites, genotyping was performed, which revealed successful excision of the flirted Atg7 locus transfected in the TRAP/FlpL line. These results suggest that Atg7 is not required for the progression of sporozoites from the midgut to the salivary gland.

In the paper evidence is presented that:
- Atg7 cKO parasites failed to mature into hepatic merozoites due to impaired organelle biogenesis
Extensive branching of the apicoplast and ER in TRAP/FlpL parasites, while this branching was lost in the Atg7 cKO parasites.
- Atg7-mediated conjugation of Atg8 is required for biogenesis of the apicoplast.
- Atg7 is essential for the exocytosis of unnecessary superfluous organelles during liver stage development.

Other mutants