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Cellular characterization of genes concerned with the contact-dependent mechanisms on Naegleria fowleri

Authors
Sohn, Hae-Jin; Kim, Jong-Hyun; Lee, Jinyoung; Sim, Moon-Hee; Yang, Hee-Jong; Kang, Hee-Kyoung; Shin, Ho-Joon
Department
Department of Microbiology
Abstract
Naegleria fowleri, pathogenic free-living amoeba has been isolated from widespread areas in soil, water, swimming pools, exists as a virulent pathogen which causes fatal primary amoebic meningoencephalitis (PAME) in experimental animal and humans. N. fowleri destroy target cells require adhesion of amoeba to host cell, secretion of proteases, and phagocytosis. In this study, cellular characterization of gene concerned with contact-dependent mechanisms on N. fowleri. The nfa1 gene and nf-actin gene was amplified from a gene-cloned vector, pEXQP5-T7/NT TOPO. The nfa1 gene and nf-actin gene was inserted into pEGFP-C2 vector, and then the pEGFP-C2/nfa1 and pEGFP-C2/nf-actin was transfected to N. fowleri and G418 selection. The strong EGFP fluorescence was observed in N. fowleri transfected with pEGFP-C2/nfa1 vector and pEGFP-C2/nf-actin. And the expression of EGFP-Nfa1 protein was well checked by Western blot. When transgenic N. fowleri trophozoites were co-cultured with U87MG target cells, the nfa1 protein was also located on cytoplasm and pseudopodia. Finally, these results suggest that nfa1 and nf-actin plays an important role in motility and phagocytic activity of N. fowleri.
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