Participation of 14-3-3ε and 14-3-3ζ proteins in the phagocytosis, component of cellular immune response, in Aedes mosquito cell linesReportar como inadecuado




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Parasites and Vectors

, 10:362

First Online: 01 August 2017Received: 27 December 2016Accepted: 03 July 2017

Abstract

BackgroundBetter knowledge of the innate immune system of insects will improve our understanding of mosquitoes as potential vectors of diverse pathogens. The ubiquitously expressed 14-3-3 protein family is evolutionarily conserved from yeast to mammals, and at least two isoforms of 14-3-3, the ε and ζ, have been identified in insects. These proteins have been shown to participate in both humoral and cellular immune responses in Drosophila. As mosquitoes of the genus Aedes are the primary vectors for arboviruses, causing several diseases such as dengue fever, yellow fever, Zika and chikungunya fevers, cell lines derived from these mosquitoes, Aag-2 from Aedes aegypti and C6-36 HT from Aedes albopictus, are currently used to study the insect immune system. Here, we investigated the role of 14-3-3 proteins ε and ζ isoform in phagocytosis, the main cellular immune responses executed by the insects, using Aedes spp. cell lines.

ResultsWe evaluated the mRNA and protein expression of 14-3-3ε and 14-3-3ζ in C6-36 HT and Aag-2 cells, and demonstrated that both proteins were localised in the cytoplasm. Further, in C6-36 HT cells treated with a 14-3-3 specific inhibitor we observed a notable modification of cell morphology with filopodia-like structure caused through cytoskeleton reorganisation co-localization of 14-3-3 proteins with F-actin, more importantly the decrease in Salmonella typhimurium, Staphylococcus aureus and E. coli phagocytosis and reduction in phagolysosome formation. Additionally, silencing of 14-3-3ε and 14-3-3ζ expression by mean of specific DsiRNA confirmed the decreased phagocytosis and phagolysosome formation of pHrodo labelled E. coli and S. aureus bacteria by Aag-2 cells.

ConclusionThe 14-3-3ε and 14-3-3ζ proteins modulate cytoskeletal remodelling, and are essential for phagocytosis of Gram-positive and Gram-negative bacteria in Aedes spp. cell lines.

KeywordsAag-2 Ae. aegypti C6-36 HT Ae. albopictus 14-3-3ε 14-3-3ζ DsiRNA 14-3-3 inhibitor Phagocytosis AbbreviationsDMSODimethyl sulfoxide

DsiRNAsDicer-substrate siRNAs

dsRNADouble-strand RNA

FITCFluorescein

GFPGreen fluorescent protein

HBSSHank’s balanced salt solution

INH14-3-3 inhibitor

MEMMinimum essential medium

RNAiRibonucleic acid interference

RT-PCRReverse transcription polymerase chain reaction

siRNAsSmall interference RNA

Electronic supplementary materialThe online version of this article doi:10.1186-s13071-017-2267-5 contains supplementary material, which is available to authorized users.

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Autor: Abel Trujillo-Ocampo - Febe Elena Cázares-Raga - Rosa María del Angel - Fernando Medina-Ramírez - Leopoldo Santos-Argume

Fuente: https://link.springer.com/



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