Mutant/mutation
In the mutant the uis4 gene (up-regulated in infective sporozoites 4) is replaced with the P. falciparum gene PF10_0164 (ETRAMP10.3; early transcribed membrane protein 10.3), which is C-terminally myc-tagged. PF10_0164 is under the control of the endogenous uis4 promoter

Protein (function)
UIS4 has been identified by transcription-profiling of P. berghei sporozoites (Kaiser et al., (2001). Mol. Microbiol. 51, 1221-32). The protein is expressed in sporozoites and throughout liver stage development and localizes to the parasitophorous vacuole membrane. Mutant parasites lacking expression of UIS4 (RMgm-35, RMgm-38) produce normal numbers of sporozoites that are able to infect liver cells. These parasites are however unable to develop into mature liver stages

P. falciparum gene PF10_0164 (ETRAMP10.3; early transcribed membrane protein 10.3) is a member of the 'early transcribed membrane protein (ETRAMP)' family. This is a Plasmodium conserved family with greater than ten members in P. falciparum. ETRAMPs are abundantly expressed early in the intraerythrocytic cycle and are small (frequently less than 200 aa) integral membrane proteins that are localized within the parasitophorous vacuolar membrane (PVM). All members have signal peptides plus a transmembrane domain. PF10_0164 is expressed in sporozoites and blood stages, where it localizes to the parasitophorous vacuole, and is also exported to the host erythrocyte. In the paper evidence is presented that PF10_0164 is refractory to disruption in asexual blood stages. PF10_0164 is syntenic with UIS4 and it has been proposed that this P. falciparum gene is the functional ortholog of P.berghei/P.yoelii UIS4

Phenotype
In the P. yoelii mutant PF10_0164myc localized to the parasitophorous vacuole membrane of liver stages, but the mutant parasites are unable to complete liver stage development. These data indicate that PF10_0164 is a parasitophorous vacuole protein that  does not complement P. yoelii UIS4, and is thus likely not a functional ortholog of UIS4. Mutant sporozoites are able to invade liver cells but liver stage development is strongly impaired. This phenotype is comparable to the phenotype of mutant parasites lacking expression of UIS4 (see RMgm-35, RMgm-38).

Additional information
The amino acid sequence identity between the PF10_0164 and UIS4 is 9% (17% conserved residues), and the predicted PF10_0164 protein is considerably shorter. Topology prediction tools suggest that the transmembrane domain is located at a position in the protein that makes the length of the N-terminus comparable between the proteins, whereas the C-terminus of PF10_0164 is considerably shorter. Both, however, possess a high density of charged residues in their C-termini (51% and 64% of residues in PF10_0164 and PyUIS4, respectively), with a net negative charge for each.

As controls, two additional mutant P. yoelii lines have been generated in which the endogenous uis4 has been replaced with either PF10_0164 without myc-tag or with the P. yoelii uis4 gene with a myc tag (PyUIS4myc). Liver stages of the first line were not able to develop into mature liver schizonts whereas liver stages of the second line showed normal liver stage development (RMgm-395).
Since the C-terminal domain of PF10_0164 is significantly shorter than that of UIS4, it can be argued that the introduction of a myc epitope tag to the C-terminus of either protein may affect their functions differently, thus compromising the use of PyUIS4myc parasites (RMgm-395) as a control. Therefore, as an additional control to ensure that the failure of PF10_0164 to complement the function of UIS4was not due to the presence of the epitope tag, the mutant line was generated in which uis4  was replaced with PF10_0164 without a tag. These parasites formed liver stages in vitro that expressed PF10_0164 without a myc tag, and the protein localized in a similar manner as seen in the Pyuis4(-)PF10_0164myc parasites. However, infecting mice by intravenous injection of sporozoites of this line did not result in blood stage infections  indicating that that PF10_0164 does not complement UIS4 function in the liver, regardless of the presence or absence of an epitope tag.

Other mutants
RMgm-35, RMgm-38: P. berghei/P. yoelii mutants lacking expression of UIS4
RMgm-395: A P. yoelii mutant expressing a c-terminal myc tagged version of UIS4