Mutant/mutation
The mutant lacks expression of CITH (CAR-I/Trailer Hitch Homolog) and expresses GFP under control of a male and RFP under control of a female gametocyte specific promoter.

Protein (function)
CITH shows homology with proteins that co-locolaize with germ cell and P granules in other organisms such as worm CAR-I and fly Trailer Hitch but also Xenopus Rap55. The Plasmodium protein contains both the conserved LSM14 domain and the extended FDF motif, known to compete with the enhancer of mRNA decapping EDC3 for binding to DDX6 helicases.

Phenotype
The phenotype analyses indicate an essential role of CITH in the development of fertilized zygotes. Fertilisation and meiosis (meiotic DNA replication) is comparable to these processes in wild type parasites but the development of zygotes into ookinetes is completely blocked.
Through RNA-immunoprecipitation, and global expression profiling of CITH-KO mutants evidence is presented that CITH is a crucial repressor of maternally supplied mRNAs and plays an important role in translational repression of mRNAs in gametocytes.

Additional information
In Plasmodium CITH was identified in immunoprecipitation experiments with the protein DOZI (protein development of zygote inhibited; ATP-dependent RNA helicase, putative). DOZI is an RNA helicase that is highly up-regulated in female gametocytes and shows high sequence homology to the DDX6 family of RNA helicases.  It contains domains involved in RNA-binding and RNA-unwinding activity.  In other eukaryotes the DDX6 family of DEAD-box RNA helicases is tightly linked both to storage of mRNAs encoding proteins associated with progression through meiosis into translationally silent mRNPs and with the transport of mRNA to degradation centers in the cell (P-bodies). These helicases are found in organisms as diverse as yeast (e.g., Dhh1p) and humans (e.g., RCK/p54).
RNA immunoprecipitation experiments indicate that CITH together with DOZI resides in a stable, translationally quiescent P-body-like structure (containing translationally repressed mRNA's).

The cith gene has been disrupted in a reporter line (RMgm-164) expressing GFP under control of a male and RFP under control of a female gametocyte specific promoter. This reporter line mutant can used to determine fertilisation of female gametes and meiosis in zygotes by FACS analysis. Female gametes/zygotes express RFP allowing for FACS selection of the RFP-positive females to be analysed. Before FACS analysis live cells are stained with the DNA specific dye Hoechst 32258 to measure DNA content and determine the ploïdy of female cells by FACS analysis (Ponzi, M. et al., 2009, Cell. Microbiol 11, 1272-88).

Other mutants
RMgm-358: A mutant that expresses a c-terminally GFP-tagged version of CITH
RMgm-359: A mutant lacking expression of CITH which expresses GFP under the constitutive eef1a promoter