Enzymatic control of anhydrobiosis-related accumulation of trehalose in the sleeping chironomid, Polypedilum vanderplanki.
Bottom Line: Although computational prediction indicated that the alternative splicing variants (PvTpsα/β) obtained encoded probable functional motifs consisting of a typical consensus domain of TPS and a conserved sequence of TPP, PvTpsα did not exert activity as TPP, but only as TPS.Trehalose was synthesized abundantly, consistent with increased activities of TPS and TPP and suppressed TREH activity.These results show that trehalose accumulation observed during anhydrobiosis induction in desiccating larvae can be attributed to the activation of the trehalose synthetic pathway and to the depression of trehalose hydrolysis.
Affiliation: Anhydrobiosis Research Unit, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, Japan.Show MeSH
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Mentions: As a result of 3′-RACE on PvTps, we isolated two distinct mRNAs, named PvTpsα and PvTpsβ, that were different at each 3′-end of the nucleotide sequence. PvTpsα cDNA consisted of 3026 bp (Fig. 3A). Because nucleotides (nt) 69–71 represent a stop codon (TAA), the downstream nt 90–92 were regarded as the initiation codon (ATG). nt 2628–2630 also represented a stop codon (TGA), thus suggesting a 2538-bp ORF (846 amino acids with a molecular mass of 95 300). PvTpsβ cDNA consisted of 3094 bp; 68 nucleotides were inserted between nt 2291 and 2292 of PvTpsα. Because a frame shift occurred by insertion, the ORF in PvTpsβ was shortened to 2373 bp, encoding 791 amino acids with a calculated molecular mass of 89 500 (Fig. 3A). The genomic DNA sequence of the PvTps gene confirmed that PvTpsα and PvTpsβ were generated by alternative splicing (Fig. 3A). In the same manner, cDNAs of PvTpp and PvTreh were defined to consist of 1044 bp, including an 882-bp ORF (294 amino acids with a molecular mass of 33 400), and 2177 bp, including a 1734-bp ORF (578 amino acids with a molecular mass of 66 400), respectively (Fig. 3B, C).
Affiliation: Anhydrobiosis Research Unit, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, Japan.