Mutant/mutation
Mutant parasites lack the tert gene. The presence of tert- mutants in transfected populations of blood stages was shown by diagnostic PCR. The mutants could not be obtained by cloning (see 'Phenotype')

Protein (function)
Telomerase is an RNA-dependent DNA polymerase complex functioning in extension and/or maintenance of telomeres.
The enzyme responsible for synthesis of new repeats is the telomerase holoenzyme, which is an RNA-dependent DNA polymerase complex. This enzyme synthesises new tandem telomeric repeats de novo at the 3'chromosome strand end.
Telomerase consists of several subunits. The core subunit consists of Telomerase Reverse Transcriptase (TERT) together with its conserved RNA component (Telomerase RNA; TR), which acts as the template for the synthesis of telomeric repeats.
The gene encoding TERT has been deleted or mutated in a number of organisms which led to cell senescence and eventual cell death (‘‘delayed death’’).
The genome of malaria parasites is arranged into 14 linear chromosomes which contain telomeres consisting of 7-bp telomeric repeat sequences (GGGTTT/CA).
The average telomere length per species varies, ranging from 850 bp (estimated ~120 repeat copies) to 6700 bp (estimated ~955 repeat copies) in the human parasites P. falciparum and P. vivax, respectively. Plasmodium telomere length appears to be kept constant during the erythrocytic cycle. The genomes of different Plasmodium parasites contain a single copy tert gene.
The RNA component of telomerase (encoded by PF3D7_0918500 in P. falciparum; 2148 nt) has been identified in silico in several Plasmodium species based on structural comparisons of conserved domains in TR domains from other organisms.

Two adjacent incomplete gene models have been identified in the genome of P. berghei (www.GeneDB.org) that encode tert genes: PBANKA_141260 (5691 bp) and PBANKA_141270 (1305 bp), that both share homology with the single copy tert genes of P. falciparum and P. chabaudi (PCHAS_141450; 6753 bp). The P. chabaudi tert gene was subsequently used as a reference for assembling the complete Pbtert gene. Amplification of the predicted gap between the two Pbtert gene models using primers specific to the adjacent tremities of both gene models revealed a sequence duplication of 57 nt, which may explain the failure of correct assembly of Pbtert which has a total size of 6939 bp. The complete Pbtert gene ishared 77%, 83% and 43% identity with the predicted tert genes of P. yoelii (17X), P. chabaudi and P. falciparum, respectively.

Phenotype
The presence of tert- mutants in transfected populations of blood stages was shown by diganostic PCR. Cloning of tert- mutants has been attempted multiple times without success. Thorough analysis of the transfected parasite populations and the parasite obtained from extensive parasite cloning from these populations provide evidence for a so called delayed death phenotype as observed in different organisms lacking TERT. The findings indicate that TERT is essential for P. berghei.

Additional information
Expression of PbTERT in mixed blood stage parasites was shown by Western blot analysis using anti-TERT antibodies raised against P. falciparum TERT.
The presence of TR transcripts (Telomerase RNA; PBANKA_081945) was shown by Northern analysis of RNA of different blood stages.
Evidence is presented for an average telomere length of 950 bp of P. berghei chromosomes.

In order to check if tert-mutants were disappearing from populations that undergo asexual multiplication, 105 parasites of the uncloned parent populations containing tert- parasites were injected in two mice and analysed the presence of tert-deletion parasites by diagnostic PCR during a period of 1–2 weeks of asexual multiplication. In all three populations the tert- parasites could not be detected at the end of 1–2 week period. These results indicate that tert-deletion mutants have a growth disadvantage compared to parasites with a WT tert copy, possibly due to a delayed cell death phenotype of tert-deletion mutants.

Other mutants