Ectoine, the compatible solute of Halomonas elongata, confers hyperosmotictolerance in cultured tobacco cells

1,4,5,6-Tetrahydro-2-methyl-4-pyrimidinecarboxylic acid (ectoine) functions as a compatible osmolyte in the moderate halophile Halomonas elongata OUT3 0018. Ectoine is biosynthesized by three successive enzyme reactions from a spartic beta-semialdehyde. The genes encoding the enzymes involved in the b iosynthesis, ectA, ectB, and ectC, encoding L-2,4-diaminobutyric acid acety ltransferase, L-2,4-diaminobutyric acid transaminase, and L-ectoine synthas e, respectively, have been previously cloned. To investigate the function o f ectoine as a compatible solute in plant cells, the three genes were indiv idually placed under the control of the cauliflower mosaic virus 35S promot er and introduced together into cultured tobacco (Nicotiana tabacum L.) cv Bright Yellow 2 (BY2) cells. The transgenic BY2 cells accumulated a small q uantity of ectoine (14-79 nmol g(-1) fresh weight) and showed increased tol erance to hyperosmotic shock (900 mOsm). Furthermore, the transgenic BY2 ce lls exhibited a normal growth pattern even under hyperosmotic conditions (u p to 530 mOsm), in which the growth of the untransformed BY2 (wild type) ce lls was obviously delayed. These results suggest that genetically engineere d synthesis of ectoine results in the increased hyperosmotic tolerance of c ultured tobacco BY2 cells despite the low level of accumulation of the solu te.