Study of the biological importance of Toxoplasma gondii H2B.Z histone and function of its acetylated lysines

VANAGAS LAURA; MUÑOZ DANIELA; GANUZA AGUSTINA; NARDELLI SHEILA; GUZMÁN FANNY; KAMI KIM; SULLIVAN WILLIAM J.; ANGEL SERGIO O.

Quindío

Congreso; XVth International Toxoplasma Congress; 2019

Universidad de Quindio - Asociación Colombiana de Infectología (ACIN))

T. gondii and other apicomplexan parasites possess an unusual H2B variant called H2B.Z which is found in nucleosomes containing H2A.Z? another histone variant. The existence of a unique H2B variant in T. gondii offers an interesting matter of study to find new biological pathways and putative drug targets. H2A.Z and H2B.Z are positioned at transcription start sites (TSS) together with H3K4Me3 and both histone variants are highly acetylated at their N-terminal tails. Mutagenesis studies were performed to determine the biological importance of H2B.Z? particularly the role of the N-terminal lysines. We over-expressed myc-tagged H2B.Z in Pru strain in which the acetylatable lysine residues were maintained (c-Myc-WT) or mutated to either the neutral amino acid alanine (c-Myc-A) or the positively charged arginine (c-Myc-R). The resulting transgenic parasites did not show significant differences in replication rate? but we observe changes in the rate of bradyzoite differentiation. Parasites ectopically expressing c-Myc-R/H2B.Z differentiated more compared with parental? while c-Myc-A/H2B.Z had a lower differentiation rate (1.88 and 0.74? respectively? while c-Myc-wt/H2B.Z rate was 1.23 relative to parental taken as 1). Since the H2B.Z gene is essential? we generated new mutants by disruption of the endogenous H2B.Z gene using CRISPR/Cas9 in the lines overexpressing the mutant histones (c-Myc-A/KO and c-Myc-R/KO). Phenotypic analysis showed similar results to those obtained with the over-expressing lines: there is a higher differentiation rate in c-Myc-R/KO parasites? and lower in c-Myc-A/KO. These findings suggest that a correct acetylation of the Nterminal region of H2B.Z is relevant for the control of parasite development.