The possibility for either opposite or synergistic effects on gene regulation must be considered at this point, and future investigations will be necessary to increase our knowledge of the interactions between HMGB1 and histones in T. gondii. Istradefylline however, there is no Histone H1 in T. gondii but in addition to having a single copy of canonical histones, T. gondii encodes five variant histones, H3.3, H2A. X, H2A. Z, and the parasite-specific H2Bv. Such studies might provide exciting new insights to advance understanding of the transcription-related functions of TgHMGBs. More direct and very important evidence for the involvement of TgHMGB1a in transcription regulation was generated by quantitative RT-PCR studies, which indicated that the transcription levels of many genes were elevated in the TgHMGB1a overexpression parasites, but were not significantly reduced in the TgHMGB1a B box-deficient strain. Nevertheless, it��s more like that TgHMGB1a is negative regulated for ROP16 but promotes transcription of profilin, however, it needs to be further investigated. Furthermore, promoter binding assays with ChIP-qPCR analysis showed that TgHMGB1a prefers bind to promoter regions and maybe in non-gene specific promoter manner. Collectively, we suggest that TgHMGB1a plays an Torsemide activator role in gene transcription regulation in T. gondii, as in mammals, and we supposed that the replication of T. gondii was controlled by cell cycle regulatory genes, which were upregulated in the TgHMGB1a overexpression strain, enhancing the role of inhibition for replication, and then the parasites showed slower growth compare with their parental strain. The DNA-related functions of TgHMGB1a are consistent with its nuclear location. Interestingly, no canonical NLS was found in TgHMGB1a by bioinformatic predictions, and there were no signal sites identified in the B box, even though B box deficient TgHMGB1a did not concentrate in the nucleus but dispersed throughout the parasite cells. In other species, the acetylation status of HMGB proteins can alter both their DNA-binding properties and their subcellular localizations; N-myristoylation and palmitoylation sites have also been shown to contribute to protein localization.