Expression of a Bacterial Ice Nucleation Gene in Plants

Abstract

We have introduced an ice nucleation gene (inaZ) from Pseudomonas syringae pv. syringae into Nicotiana tabacum, a freezingsensitive species, and Solanum commersonii, a freezing-tolerant species. Transformants of both species showed increased ice nucleation activity over untransformed controls. The concentration of ice nuclei detected at -10.5°C in 15 different primary transformants of S. commersonii varied by over 1000-fold, and the most active transformant contained over 100 ice nuclei/mg of tissue. The temperature of the warmest freezing event in plant samples of small mass was increased from approximately -12°C in the untransformed controls to -4°C in inaZ-expressing transformants. The threshold nucleation temperature of samples from transformed plants did not increase appreciably with the mass of the sample. The most abundant protein detected in transgenic plants using immunological probes specific to the inaZ protein exhibited a higher mobility on sodium dodecyl sulfate polyacrylamide gels than the inaZ protein from bacterial sources. However, some protein with a similar mobility to the inaZ protein could be detected. Although the warmest ice nucleation temperature detected in transgenic plants is lower than that conferred by this gene in P. syringae (-2°C), our results demonstrate that the ice nucleation gene of P. syringae can be expressed in plant cells to produce functional ice nuclei.

Bacteria in the ECHAM5-HAM global climate model

“…Its ice nucleation activity is conferred by a single gene (inaZ) that encodes the outer membrane InaZ protein. Individual InaZ proteins cannot serve as IN, but form large, homogeneous aggregates that collectively orient water molecules into a conformation mimicking the crystalline structure of ice, thereby catalysing ice formation (Baertlein et al, 1992). It has also been shown that there are ice nucleation active fungi (Pouleur et al, 1992;Iannone et al, 2011) and pollen (Diehl et al, 2001(Diehl et al, , 2002von Blohn et al, 2005), though they initiate freezing at colder temperatures than bacteria do.…”

Functional display of ice nucleation protein InaZ on the surface of bacterial ghosts

“…23,24 Recombinant bacterial INPs were already functionally expressed in Gram-negative non-ice nucleation active Escherichia coli, as well as in plants. 25,26 INPs are also used for molecular biologic applications, using ice-nucleation activity as a reporter gene system. 27 In addition, the N-terminal domain of INPs, which link the protein to the outer cell membrane is used as cell surface display system.…”

Environmental release of transgenic trees in Canada — potential benefits and assessment of biosafety

“…Tolerance has been achieved in the case of hybrid aspens and poplars to glyphosate (Donahue et al 1994;Strauss et al 1997), to chlorsulfuron (Brasileiro et al 1992), and to phosphinothricin (DeBlock 1990), and in European larch to glyphosate (Shin et al 1994). Genetic engineering of cold tolerance has been a focal point for a g r i c u l d crops where various strategies have been developed such as insertion of ice nucleation genes from bacteria (Baertlein et al 1992), antifreeze proteins from cold water fish species (Georges et al 1990), and altered lipid composition of cell and plastid membranes (Murata et al 1992;Wolter et al 1992). Increased cold tolerance would enhance the safety of northerly seed transfers that take advantage of longer day-length, and would expand the safe planting zones for valuable exotic species.…”