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An ARV1 homologue from a filarial nematode is functional in yeast

Published online by Cambridge University Press:  29 November 2017

H.M.L.P.B. Herath ,
Y.I.N.S. Gunawardene ,
M. Pathiranage ,
P.D.S.U. Wickramasinghe ,
P.G.T.S. Wickramatunge  and
R.S. Dassanayake
H.M.L.P.B. Herath
Affiliation:
Department of Chemistry, Faculty of Science, University of Colombo, Colombo 03, Sri Lanka
Y.I.N.S. Gunawardene
Affiliation:
Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, Kelaniya, Sri Lanka
M. Pathiranage
Affiliation:
Department of Chemistry, Faculty of Science, University of Colombo, Colombo 03, Sri Lanka
P.D.S.U. Wickramasinghe
Affiliation:
Department of Chemistry, Faculty of Science, University of Colombo, Colombo 03, Sri Lanka
P.G.T.S. Wickramatunge
Affiliation:
Department of Chemistry, Faculty of Science, University of Colombo, Colombo 03, Sri Lanka
R.S. Dassanayake*
Affiliation:
Department of Chemistry, Faculty of Science, University of Colombo, Colombo 03, Sri Lanka
*
Author for correspondence: R.S. Dassanayake, E-mail: [email protected]
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Abstract

The transmembrane protein, ARV1, plays a key role in intracellular sterol homeostasis by controlling sterol distribution and cellular uptake. To date, only the ARV1s from yeast and humans have been characterized to some extent. In this study, the ARV1 of an animal filarial parasite, Setaria digitata (SdARV1), was characterized; its cDNA was 761 bp and encoded a protein of 217 amino acids, with a predicted molecular weight of 25 kDa, containing a highly conserved ARV1 homology domain and three transmembrane domains in the bioinformatic analyses. Information required to cluster members belonging to a particular taxon has been revealed in phylogenetic analyses of ARV1 sequences derived from different organisms. Reverse transcription-polymerase chain reaction (RT-PCR) analyses indicated that SdARV1 was expressed in different developmental stages – microfilariae and adult male and female worms. Experiments carried out with a single copy of the SdARV1 under the control of the PMA-1 promoter in a temperature-sensitive Saccharomyces cerevisiae mutant strain indicated full complementation of the mutant phenotype, with growth at a non-permissive temperature (37°C). Microscopic observations of cellular morphology with Gram staining revealed alteration of the shape from shrunken to oval, in mutant and complemented strains, respectively. Assessment of free sterol levels extracted from mutant yeast and complemented strains indicated that the level of sterol was significantly higher in the former compared to the latter, which had sterol levels similar to those of the wild type. Thus, the results of the current study suggest that SdARV1 is ubiquitously expressed in different developmental stages of S. digitata, and that it is a true functional homologue of mammalian and yeast ARV1s, which have crucial phylogenetic information that follows classical evolutionary trends. Finally, this is the first study to report the biological function of nematode ARV1.

Type
Research Paper
Information
Journal of Helminthology , Volume 93 , Issue 1 , January 2019 , pp. 12 - 20
Copyright
Copyright © Cambridge University Press 2017 

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