Autophagy protein 16-mediated autophagy is required for the encystation of Acanthamoeba castellanii

Song, Su Min; Han, Byung In; Moon, Eun Kyung; Lee, Yu Ran; Yu, Hak Sun; Jha, Bijay Kumar; Danne, Dinzouna Boutamba Sylvatrie; Kong, Hyun-Hee; Chung, Dong Il; Hong, Yeonchul

doi:10.1016/j.molbiopara.2012.02.013

Cited by 44 publications

(45 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In our previous studies, we determined that autophagy is essential for the encystation of Acanthamoeba (58,59), and it is likely that the higher expression levels of cysteine proteases during encystation are associated with degradation of intracellular contents in autophagosomes (60). Murine cystatin C (CysC) is known to play a protective role under conditions of neuronal challenge by inducing autophagy via the mTOR pathway, which leads to enhanced proteolytic clearance of autophagy substrates by lysosomes, whereas the inhibitory effects of CysC on cathepsin B are not required for these neuroprotective functions (61).…”

Section: Discussionmentioning

confidence: 99%

Cysteine Protease Inhibitor (AcStefin) Is Required for Complete Cyst Formation of Acanthamoeba

et al. 2013

Self Cite

View full text Add to dashboard Cite

dThe encystation of Acanthamoeba leads to the formation of resilient cysts from vegetative trophozoites. This process is essential for parasite survival under unfavorable conditions, such as those associated with starvation, low temperatures, and biocides. Furthermore, cysteine proteases have been implicated in the massive turnover of intracellular components required for encystation. Thus, strict modulation of the activities of cysteine proteases is required to protect Acanthamoeba from intracellular damage. However, mechanisms underlying the control of protease activity during encystation have not been established in Acanthamoeba. In the present study, we identified and characterized Acanthamoeba cysteine protease inhibitor (AcStefin), which was found to be highly expressed during encystation and to be associated with lysosomes by fluorescence microscopy. Recombinant AcStefin inhibited various cysteine proteases, including human cathepsin B, human cathepsin L, and papain. Transfection with small interfering RNA against AcStefin increased cysteine protease activity during encystation and resulted in incomplete cyst formation, reduced excystation efficiency, and a significant reduction in cytoplasmic area. Taken together, these results indicate that AcStefin is involved in the modulation of cysteine proteases and that it plays an essential role during the encystation of Acanthamoeba.A canthamoeba is the causative agent of granulomatous amoebic encephalitis (GAE) and amoebic keratitis. The life cycle of Acanthamoeba is divided into two stages, that is, the vegetative trophozoite stage and the dormant cystic stage. Under challenging conditions, such as those associated with starvation, low temperatures, and biocide treatment, the trophozoite converts to the resilient cyst form (1-5). This differentiation, which is termed encystation, protects Acanthamoeba against host immune responses and allows it to evade the effect of chemical treatments, and thus, encystation decreases treatment efficacies (3, 6). Therefore, the inhibition of encystation during medical treatment could lead to outcomes that are more favorable in cases of amoebic infection. However, this goal is hindered by a lack of information about the encystation mechanism involved.The cysteine proteases are regarded as major drug targets, and the papain family of cysteine proteases, such as cathepsin B and cathepsin L, are localized in lysosomes for intracellular protein degradation (7,8). Previous studies on Giardia and Entamoeba have shown that cysteine proteases are implicated in parasite survival under nutrient-limited conditions and during developmental differentiation (9-14). In Acanthamoeba, the cysteine protease inhibitor E64d was found to slow encystment (15), and in a previous study, we detected the upregulations of various proteases, including cysteine proteases, by cDNA microarray analysis and comparative expression analysis of expressed sequence tags (ESTs) during encystation (16,17). Cysteine proteases in lysosomes cause massive proteolytic de...

show abstract

Section: Discussionmentioning

confidence: 99%

Cysteine Protease Inhibitor (AcStefin) Is Required for Complete Cyst Formation of Acanthamoeba

et al. 2013

Self Cite

View full text Add to dashboard Cite

show abstract

“…The biological significance of autophagy associated with starvation, differentiation, development, and immunity has been previously demonstrated (10)(11)(12)(13). In Acanthamoeba cells, several genes associated with autophagy, including those encoding Atg8, Atg8 isoform, Atg3, and Atg16L, were identified and their roles in the encystation of Acanthamoeba cells were studied (14)(15)(16)(17). Autophagy is a major cellular pathway for the encystation of Acanthamoeba cells.…”

mentioning

confidence: 99%

Autophagy Inhibitors as a Potential Antiamoebic Treatment for Acanthamoeba Keratitis

Moon

Kim

Hong

et al. 2015

Antimicrob Agents Chemother

Self Cite

View full text Add to dashboard Cite

bAcanthamoeba cysts are resistant to extreme physical and chemical conditions. Autophagy is an essential pathway for encystation of Acanthamoeba cells. To evaluate the possibility of an autophagic Acanthamoeba encystation mechanism, we evaluated autophagy inhibitors, such as 3-methyladenine (3MA), LY294002, wortmannin, bafilomycin A, and chloroquine. Among these autophagy inhibitors, the use of 3MA and chloroquine showed a significant reduction in the encystation ratio in Acanthamoeba cells. Wortmannin also inhibited the formation of mature cysts, while LY294002 and bafilomycin A did not affect the encystation of Acanthamoeba cells. Transmission electron microscopy revealed that 3MA and wortmannin inhibited autophagy formation and that chloroquine interfered with the formation of autolysosomes. Inhibition of autophagy or autolysosome formation resulted in a significant block in the encystation in Acanthamoeba cells. Clinical treatment with 0.02% polyhexamethylene biguanide (PHMB) showed high cytopathic effects on Acanthamoeba trophozoites and cysts; however, it also revealed high cytopathic effects on human corneal epithelial cells. In this study, we investigated effects of the combination of a low (0.00125%) concentration of PHMB with each of the autophagy inhibitors 3MA, wortmannin, and chloroquine on Acanthamoeba and human corneal epithelial cells. These new combination treatments showed low cytopathic effects on human corneal cells and high cytopathic effects on Acanthamoeba cells. Taken together, these results provide fundamental information for optimizing the treatment of Acanthamoeba keratitis.A canthamoeba trophozoites differentiate into cysts under conditions of physical stresses, starvation, and chemical exposure (1-3); the cysts are resistant to several chemical biocides (4). Resistance of the cysts to chemotherapeutic drugs makes the treatment of Acanthamoeba infections difficult (4, 5). Although polyhexamethylene biguanide (PHMB) (0.02%), alone or in combination with a diamidine (0.1% propamidine or 0.1% hexamidine), showed therapeutic efficacy against Acanthamoeba keratitis (6), PHMB was reported to be cytotoxic to human corneal epithelial (HCE) cells (7). Although therapy for Acanthamoeba keratitis needs better-acting drugs, due to the encystation of Acanthamoeba cells, cytotoxicity toward mammalian cells must also be considered.Recent studies have demonstrated anticancer therapies that include autophagy targeting in different tumor cell lines (8, 9). One change in the cellular composition during the encystation of protozoa is the formation of an autophagy (10). The biological significance of autophagy associated with starvation, differentiation, development, and immunity has been previously demonstrated (10-13). In Acanthamoeba cells, several genes associated with autophagy, including those encoding Atg8, Atg8 isoform, Atg3, and Atg16L, were identified and their roles in the encystation of Acanthamoeba cells were studied (14-17). Autophagy is a major cellular pathway for the encystation of Acanth...

show abstract

“…Previous reports of cyst-specific gene expression profiles suggest that autophagy genes are highly expressed and have crucial roles during encystation of A. castellanii (13,14,32). Accordingly, we speculated that inhibiting the autophagic process could block encystation of Acanthamoeba, ultimately leading to decreased cell viability.…”

Section: Resultsmentioning

confidence: 86%

“…Because it blocks autophagy, chloroquine has also been used as a treatment for cancer, lupus erythematosus, and rheumatoid arthritis (50). Recent studies suggest that autophagy plays an important role as a survival mechanism during the Acanthamoeba encystation process (13,14). In this sense, we hypothesized that chloroquine treatment might have a specific effect during encystation and might induce cell death by inhibiting autophagy.…”

Section: Discussionmentioning

confidence: 98%

See 1 more Smart Citation

Chloroquine Has a Cytotoxic Effect on Acanthamoeba Encystation through Modulation of Autophagy

Jha

Jung

Seo

et al. 2014

Antimicrob Agents Chemother

Self Cite

View full text Add to dashboard Cite

bEncystation of Acanthamoeba castellanii is associated with resistance to chemotherapeutic agents. Blocking the encystation process could potentiate the efficacy of chemotherapeutic agents and biocides. During encystation, autophagy is highly stimulated and required for proper encystation of Acanthamoeba. In this study, the cytotoxic effect of chloroquine, a well-known autophagy-inhibitory drug, was tested in A. castellanii. Chloroquine was able to selectively reduce cell survival during the encystation of A. castellanii. However, A. castellanii trophozoites and mature cysts were resistant to chloroquine. Chloroquine treatment led to an increase in the number and size of lysosomes in encysting cells. Moreover, chloroquine inhibited the degradation of long-lived proteins in the encysting cells. Decreased autophagic flux, indicated by an increased number of lysosomes and decreased degradation of long-lived proteins, may be the mechanism by which cell death is induced by chloroquine in encysting Acanthamoeba. These results suggest a potential novel therapeutic application of chloroquine as an anti-Acanthamoeba drug. Our findings also suggest that targeting autophagy could be a therapeutic strategy against Acanthamoeba infection.

show abstract

Autophagy protein 16-mediated autophagy is required for the encystation of Acanthamoeba castellanii

Cited by 44 publications

References 37 publications

Cysteine Protease Inhibitor (AcStefin) Is Required for Complete Cyst Formation of Acanthamoeba

Cysteine Protease Inhibitor (AcStefin) Is Required for Complete Cyst Formation of Acanthamoeba

Autophagy Inhibitors as a Potential Antiamoebic Treatment for Acanthamoeba Keratitis

Chloroquine Has a Cytotoxic Effect on Acanthamoeba Encystation through Modulation of Autophagy

Contact Info

Product

Resources

About