Autophagy protein 8 mediating autophagosome in encysting Acanthamoeba.
Autophagy is an evolutionally conserved protein degradation pathway in eukaryotes. It plays essential roles during starvation, cellular differentiation, cell death, and aging by eliminating unwanted or unnecessary organelles and recycling the components for reuse. ATG8, a member of a novel ubiquitin-like protein family, is an essential component of the autophagic machinery. The present study identified and characterized autophagy protein 8 in Acanthamoeba castellanii an amphizoic amoeba causing granulomatous amoebic encephalitis and amoebic keratitis in humans. Real-time polymerase chain reaction demonstrated that the A. castellanii Atg8 (AcAtg8) gene encoding a 118 amino acid protein was highly expressed during encystation. Fluorescence microscopic analysis following transient transfection of enhanced green fluorescent protein-AcAtg8 revealed small or large vacuolar fluorescent structures in an encysting amoeba. The Atg8 fluorescent structures on the membrane were identified as autophagosomes by co-localization analysis with LysoTracker. Chemically synthesized small interfering RNA against AcAtg8 reduced the encystation efficiency and inhibited autophagosome formation in Acanthamoeba.
Moon
Eun-Kyung
EK
Department of Parasitology, Kyungpook National University School of Medicine, 101 Dongin-dong, Joong-gu, Taegu, Republic of Korea.
Chung
Dong-Il
DI
Hong
Yeon-Chul
YC
Kong
Hyun-Hee
HH
D048248:Acanthamoeba castellanii / Q000737:chemistry / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D019677:Artificial Gene Fusion; D001343:Autophagy; D002462:Cell Membrane / Q000737:chemistry; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D020869:Gene Expression Profiling; D020868:Gene Silencing; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D010588:Phagosomes / Q000737:chemistry / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D016054:DNA, Protozoan; D015800:Protozoan Proteins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins