Acanthamoeba castellanii exhibits intron retention during encystment

Abstract

Intron retention (IR) refers to the mechanism of alternative splicing in which an intron is not excised from the mature transcript. IR in the cosmopolitan free-living amoeba Acanthamoeba castellanii has not been studied.We performed an analysis of RNA sequencing data during encystment to identify genes that presented differentially retained introns during this process. We show that IR increases during cyst formation, indicating a potential mechanism of gene regulation that could help downregulate metabolism. We identify 69 introns from 67 genes that are differentially retained comparing the trophozoite stage and encystment after 24 and 48 hours. These genes include several hypothetical proteins. We show different patterns of IR during encystment taking as examples a lipase, a peroxin-3 protein, an Fbox domain containing protein, a proteasome subunit, a polynucleotide adenylyltransferase and a tetratricopeptide domain containing protein. A better understanding of IR in Acanthamoeba, and even other protists, could help elucidate changes in life cycle and combat disease such as Acanthamoeba keratitis in which the cyst is key for its persistence.

Identification of an Antimicrobial Protease from Acanthamoeba via a Novel Zymogram

“…Alcian blue (incubation for 3 h in 18 mL of H 2 O, 2 mL of 95% ethanol, and 0.06 g Alcian blue while destaining with H 2 O) and methylene blue (90 min with 20 mL of H 2 O with 0.02 g of methylene blue, 0.002 g NaOH, and 0.002 g KCl while destaining with H 2 O) did not destain properly. Coomassie blue showed proper destaining and contrast to observe the band of enzymatic activity [20]. The zymogram showed similar results using E. coli and A. butzleri, which are both Gram-negative bacteria (Figure 1A).…”

“…Finally, RNAseq data [20][21][22] previously obtained to study encystment was used to analyze protease expression throughout the amoeba life cycle to look for differentially expressed proteases. Automated TruSeq-stranded mRNA-seq from total RNA were used to prepare the libraries.…”

“…The data presented in this study are available throughout the article and more in depth can be found in de Obeso Fernández del Valle [20].…”