Cold stress regulation of a bi-functional 3-dehydroquinate dehydratase/shikimate dehydrogenase (DHQ/SDH)-like gene in the freshwater green alga Spirogyra varians

A low temperature up-regulated gene was isolated from Spirogyra varians using the differentially expressed gene method. The full cDNA sequence consisted of 1798 bp with an open reading frame of 1560 bp encoding 520 amino acid residues, which had neither a cleavage site nor a signal peptide. The deduced amino acid sequence was highly similar to the bi-functional 3-dehydroquinate dehydratase/shikimate dehydrogenase (DHQ/SDH) gene of higher plants. As the gene was cold regulated, we named it SVCR3 (Spirogyra varians cold regulated gene). SVCR3 had two conserved domains of DHQase I and SDH (AroE), as well as a shikimate-binding site as found in other DHQ/SDH genes. Northern blot analysis showed that SVCR3 was up-regulated at 4 degrees C. When combined with light (> 50 mu mol photon m(-2) s(-1)) the expression of SVCR3 increased more at 4 degrees C, but the expression was not affected by light intensity alone up to 200 mmol photon m(-2) s(-1) at temperatures higher than 10 degrees C. Using the zymogram method (which is a visualization method for gel areas containing specific enzymes based on chromogenic reactions), we showed that there were three isozymes of DHQ/SDH in Spirogyra varians. This is the first report of DHQ/SDH-like gene regulation by cold stress.