Abstract
BackgroundSome organisms can survive extreme desiccation by entering a state of suspended animation known as anhydrobiosis. The free-living mycophagous nematode Aphelenchus avenae can be induced to enter anhydrobiosis by pre-exposure to moderate reductions in relative humidity (RH) prior to extreme desiccation. This preconditioning phase is thought to allow modification of the transcriptome by activation of genes required for desiccation tolerance.ResultsTo identify such genes, a panel of expressed sequence tags (ESTs) enriched for sequences upregulated in A. avenae during preconditioning was created. A subset of 30 genes with significant matches in databases, together with a number of apparently novel sequences, were chosen for further study. Several of the recognisable genes are associated with water stress, encoding, for example, two new hydrophilic proteins related to the late embryogenesis abundant (LEA) protein family. Expression studies confirmed EST panel members to be upregulated by evaporative water loss, and the majority of genes was also induced by osmotic stress and cold, but rather fewer by heat. We attempted to use RNA interference (RNAi) to demonstrate the importance of this gene set for anhydrobiosis, but found A. avenae to be recalcitrant with the techniques used. Instead, therefore, we developed a cross-species RNAi procedure using A. avenae sequences in another anhydrobiotic nematode, Panagrolaimus superbus, which is amenable to gene silencing. Of 20 A. avenae ESTs screened, a significant reduction in survival of desiccation in treated P. superbus populations was observed with two sequences, one of which was novel, while the other encoded a glutathione peroxidase. To confirm a role for glutathione peroxidases in anhydrobiosis, RNAi with cognate sequences from P. superbus was performed and was also shown to reduce desiccation tolerance in this species.ConclusionsThis study has identified and characterised the expression profiles of members of the anhydrobiotic gene set in A. avenae. It also demonstrates the potential of RNAi for the analysis of anhydrobiosis and provides the first genetic data to underline the importance of effective antioxidant systems in metazoan desiccation tolerance.
Highlights
Some organisms can survive extreme desiccation by entering a state of suspended animation known as anhydrobiosis
An expressed sequence tags (ESTs) dataset from A. avenae enriched for dehydrationinduced sequences A 5’ oligo capping method was used to construct an EST library of genes enriched in those upregulated on drying in A. avenae: an initial screen of 984 clones by reverse Northern dot blots, using cDNA from both dried and control nematodes in parallel experiments, yielded 88 ESTs which appeared to be induced by desiccation
The closest BLASTX match is with lea-1 of Caenorhabditis elegans and, the BLAST score is not highly significant (0.001), other features of the predicted protein sequence identify it as a Accession Closest BLAST hita no
Summary
Some organisms can survive extreme desiccation by entering a state of suspended animation known as anhydrobiosis. The free-living mycophagous nematode Aphelenchus avenae can be induced to enter anhydrobiosis by pre-exposure to moderate reductions in relative humidity (RH) prior to extreme desiccation. This preconditioning phase is thought to allow modification of the transcriptome by activation of genes required for desiccation tolerance. Anhydrobiotic organisms are capable of surviving in this state for an indefinite period of time after which, when environmental conditions become favourable, the organism can resume normal metabolic activity [2,3] These organisms are widespread throughout nature and include yeasts, bdelloid rotifers, tardigrades, plants and nematodes. Some steinernemitid species exhibit partial desiccation tolerance [8,9]
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