The Nf-actin gene is an important factor for food-cup formation and cytotoxicity of pathogenic Naegleria fowleri.
Naegleria fowleri destroys target cells by trogocytosis, a phagocytosis mechanism, and a process of piecemeal ingestion of target cells by food-cups. Phagocytosis is an actin-dependent process that involves polymerization of monomeric G-actin into filamentous F-actin. However, despite the numerous studies concerning phagocytosis, its role in the N. fowleri food-cup formation related with trogocytosis has been poorly reported. In this study, we cloned and characterized an Nf-actin gene to elucidate the role of Nf-actin gene in N. fowleri pathogenesis. The Nf-actin gene is composed of 1,128-bp and produced a 54.1-kDa recombinant protein (Nf-actin). The sequence identity was 82% with nonpathogenic Naegleria gruberi but has no sequence identity with other mammals or human actin gene. Anti-Nf-actin polyclonal antibody was produced in BALB/c mice immunized with recombinant Nf-actin. The Nf-actin was localized on the cytoplasm, pseudopodia, and especially, food-cup structure (amoebastome) in N. fowleri trophozoites using immunofluorescence assay. When N. fowleri co-cultured with Chinese hamster ovary cells, Nf-actin was observed to localize around on phagocytic food-cups. We also observed that N. fowleri treated with cytochalasin D as actin polymerization inhibitor or transfected with antisense oligomer of Nf-actin gene had shown the reduced ability of food-cup formation and in vitro cytotoxicity. Finally, it suggests that Nf-actin plays an important role in phagocytic activity of pathogenic N. fowleri.
Department of Microbiology, Ajou University School of Medicine, Suwon, South Korea.
Hae-Jin
HJ
Sohn
Jong-Hyun
JH
Kim
Myeong-Heon
MH
Shin
Kyoung-Ju
KJ
Song
Ho-Joon
HJ
Shin
D000199:Actins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D003593:Cytoplasm / Q000737:chemistry; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016848:Naegleria fowleri / Q000472:pathogenicity*; D010587:Phagocytosis; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D011554:Pseudopodia / Q000737:chemistry; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D037521:Virulence Factors / Q000737:chemistry / Q000235:genetics / Q000502:physiology*
D000199:Actins; D016054:DNA, Protozoan; D015800:Protozoan Proteins; D037521:Virulence Factors