SUMMARY Cytosine DNA methylation is an epigenetic mark in most eukaryotic cells that regulates numerous processes, including gene expression and stress responses. We performed a genome-wide analysis of DNA methylation in the human malaria parasite Plasmodium falciparum. We mapped the positions of methylated cytosines and identified a single functional DNA methyltransferase, PfDNMT, that may mediate these genomic modifications. These analyses revealed that the malaria genome is asymmetrically methylated, in which only one DNA strand is methylated, and shares common features with undifferentiated plant and mammalian cells. Notably, core promoters are hypomethylated and transcript levels correlate with intra-exonic methylation. Additionally, there are sharp methylation transitions at nucleosome and exon-intron boundaries. These data suggest that DNA methylation could regulate virulence gene expression and transcription elongation. Furthermore, the broad range of action of DNA methylation and uniqueness of PfDNMT suggest that the methylation pathway is a potential target for anti-malarial strategies.

All materials had suitable biocompatibility and bioactivity. The MTA P, BIO and CSCR ZrO groups had the highest viability rates and velocity of proliferation whilst the CSCR Nb O group produced more mineralized nodules.

Apicomplexan parasites are responsible for numerous important human diseases including toxoplasmosis, cryptosporidiosis, and most importantly malaria. There is a constant need for new antimalarials, and one of most keenly pursued drug targets is an ancient algal endosymbiont, the apicoplast. The apicoplast is essential for parasite survival, and several aspects of its metabolism and maintenance have been validated as targets of anti-parasitic drug treatment. Most apicoplast proteins are nuclear encoded and have to be imported into the organelle. Recently, a protein translocon typically required for endoplasmic reticulum associated protein degradation (ERAD) has been proposed to act in apicoplast protein import. Here, we show ubiquitylation to be a conserved and essential component of this process. We identify apicoplast localized ubiquitin activating, conjugating and ligating enzymes in Toxoplasma gondii and Plasmodium falciparum and observe biochemical activity by in vitro reconstitution. Using conditional gene ablation and complementation analysis we link this activity to apicoplast protein import and parasite survival. Our studies suggest ubiquitylation to be a mechanistic requirement of apicoplast protein import independent to the proteasomal degradation pathway.

Aim To evaluate the biocompatibility and mineralized nodule formation of an experimental tricalcium silicate cement with tantalum oxide (TSC/Ta 2 O 5 ) as radiopacifier, Neo MTA Plus (Avalon Biomed Inc., Bradenton, FL, USA) and MTA (Angelus, Londrina, PR, Brazil) on human osteoblast-like cells (Saos-2). Methodology Biocompatibility was evaluated by 3-(4,5-dimethyl-thiazoyl)-2,5-diphenyl-tetrazolium bromide (MTT) and neutral red (NR) assays, after exposure of Saos-2 to cement extracts at 1 : 1, 1 : 2, 1 : 4 and 1 : 8 dilutions for 24 h. Bioactivity was evaluated by alkaline phosphatase (ALP) activity, and calcium deposits were detected with alizarin red staining (ARS). Statistical analysis was performed with analysis of variance and Bonferroni or Tukey post-test (a = 0.05). Results The MTT assay revealed lower cytotoxicity for NEO and MTA (P < 0.05), and higher for TSC/ Ta 2 O 5 at 1 : 1 and 1 : 2 dilutions when compared to serum-free medium -control (P > 0.05). At 1 : 4 dilution, the TSC/Ta 2 O 5 cytotoxicity was similar to the control (P > 0.05). At 1 : 8 dilution, cell viability was significantly greater than the control (P < 0.05). Saos-2 cell viability performed using the NR assay at all dilutions revealed no cytotoxic effect of MTA, NEO and TSC/Ta 2 O 5 . ALP activity at 1 and 3 days was similar to the control (P > 0.05). TSC/Ta 2 O 5 had significantly greater ALP activity at 7 days when compared with the control (P < 0.05). All materials induced the production of mineralized nodules, and NEO produced significantly more mineralized nodules than MTA and TSC/Ta 2 O 5 (P < 0.05). Conclusions Neo MTA Plus and TSC/Ta 2 O 5 were biocompatible and induced ALP activity in Saos-2 cells. Both materials induced mineralized nodule formation by Saos-2 with Neo MTA Plus producing significantly more.