Overexpression of a Na+/H+ antiporter confers salt tolerance on a freshwater cyanobacterium, making it capable of growth in sea water

Abstract

The salt tolerance of a freshwater cyanobacterium, Synechococcus sp. PCC 7942, transformed with genes involved in the synthesis of a Na(+)/H(+) antiporter, betaine, catalase, and a chaperone was examined. Compared with the expression of betaine, catalase, and the chaperone, the expression of the Na(+)/H(+) antiporter gene from a halotolerant cyanobacterium (ApNhaP) drastically improved the salt tolerance of the freshwater cyanobacterium. The Synechococcus cells expressing ApNhaP could grow in BG11 medium containing 0.5 M NaCl as well as in sea water, whereas those expressing betaine, catalase, and the chaperone could not grow under those conditions. The coexpression of ApNhaP with catalase or ApNhaP with catalase and betaine did not further enhance the salt tolerance of Synechococcus cells expressing ApNhaP alone when grown in BG11 medium containing 0.5 M NaCl. Interestingly, the coexpression of ApNhaP with catalase resulted in enhanced salt tolerance of cells grown in sea water. These results demonstrate a key role of sodium ion exclusion by the Na(+)/H(+) antiporter for the salt tolerance of photosynthetic organisms.

Figure 1

Schematic structure of expression vectors.

Figure 2

Effects of salt stress on the growth rates of the control and transformed Synechococcus cells. The control and transformed cells at logarithmic phase in BG11 medium were subjected to salt stress by inoculation into fresh medium containing the indicated concentrations of NaCl.

Figure 3

Exclusion of Na⁺ from Na⁺ loaded cells. The control and ApNhaP-expressing cells were grown in BG11 medium until the optical density at 730 nm reached 0.5 and then they were transferred to medium containing 0.5 M NaCl. After incubation for 1 day, diethanolamine-induced exclusion of Na⁺ was measured.

Figure 4

Effects of salt shock on the growth rate (A) and photosynthetic electron transport activity (B) in control and ApNhaP-expressing Synechococcus cells. The control and ApNhaP-expressing cells were grown in BG11 medium until the optical density at 730 nm reached 0.5 and then transferred to the new medium containing 0.65 M NaCl.

Figure 5

Growth curve (A) and absorption spectra (B) of control and transformed Synechococcus cells in sea water. The control and transformed cells at logarithmic phase in BG11 medium were transferred to sea water. In growth curve (A), the initial growth rates ([doubling times (day)]⁻¹) of the cells expressing ApNhaP, ApNhaP + KatE, and ApNhaP + KatE + Bet were 0.88 ± 0.06, 0.76 ± 0.03, and 0.64 ± 0.03, respectively. The absorption spectra (B) of the control and ApNhaP-expressing cells were measured 3 days after transfer to sea water. The difference spectrum between the ApNhaP and control cells was normalized to chlorophyll absorption.