cDNA representational difference analysis for identifying transcripts regulated under anhydrobiosis in the tardigrade Milnesium tardigradum

Although water is a prerequisite for life, some organisms are capable of surviving complete cellular dehydration by entering a state called anhydrobiosis. In this state, no metabolic activity is measurable and these organisms can survive several years in dry form until water becomes available again. Anhydrobiosis has been described in bacteria, fungi, plants and many invertebrates. Among those invertebrates, tardigrades are also known to be capable to undergo anhydrobiosis by forming a so-called 'tun' stage. Therefore, they represent an interesting model system for studying the cellular response to desiccation. We have applied cDNA representational difference analysis to investigate the transcriptional response of the tardigrade Milnesium tardigradum to desiccation. This resulted in the identification of two transcripts that showed significant regulation between active and anhydrobiotic animals as validated by quantitative real-time PCR. One of these putative transcripts was downregulated fivefold. It did not show any significant similarity to known transcripts, but had previously been cloned in M. tardigradum. The second transcript could be identified as the M. tardigradum homologue of a subunit of replication factor C and was downregulated more than sixfold in the anhydrobiotic state. Replication factor C plays an important role in DNA replication during cell division. Both candidates showed an even lower expression in the transition stage between active and the tun stage (27- and 25-fold reduction, respectively), implying active inhibition of transcription of these two transcripts. Our results suggest that the potential modulation of DNA replication and its role in protecting genome stability should attract more attention when studying anhydrobiotic animals.