Plastid targeting strategies for cyanophycin synthetase to achieve high-level polymer accumulation in Nicotiana tabacum

Autor: Hühns, M.; Neumann, K. ; Hausmann, T.; Ziegler, K. ; Klemke, F. ; Kahmann, U. ; Staiger, D. ; Lockau, W.; Pistorius E.K.; Broer, I.
Mitarbeiter: Inge Broer, Maja Hühns, Tina Hausmann
weitere Bearbeiter: Maja Hühns
In:

Plant Biotechnology Journal

Herausgeber: Blackwell Publishing Ltd
Bandangabe: 6
Auflage: 4
ISBN: ISSN 1467-7644
Seite: 321 - 336
Jahr: 2008

Einordung:
Institut: Professur Agrobiotechnologie

Abstract:
The production of biodegradable polymers in transgenic plants is an important challenge in plant biotechnology; nevertheless it is often accompanied by reduced plant fitness. In order to diminish the phenotypic abnormalities caused by cytosolic production of the biodegradable polymer cyanophycin, and to increase polymer accumulation four translocation pathway signal sequences for import into chloroplasts were individually fused to the coding region of the cyanophycin synthetase gene (cphATe) of Thermosynechococcus elongatus BP-1 resulting in the constructs named pRieske-cphATe, pCP24-cphATe, pFNR-cphATe and pPsbY-cphATe. These constructs were expressed in Nicotiana tabacum var. Petit Havanna SRI under the control of the constitutive CaMV35S promoter. Three of the four constructs led to polymer production. However, only the construct pPsbY-cphATe led to cyanophycin accumulation exclusively in chloroplasts. In plants transformed with the pCP24-cphATe and pFNR-cphATe construct water-soluble and insoluble forms of cyanophycin were only located in the cytoplasm, which resulted in phenotypic changes similar to those observed in plants, transformed with constructs lacking a targeting sequence. The plants transformed with pPsbY-cphATe produced predominantly the water insoluble form of cyanophycin. The polymer accumulated to up to 1.7% of dry matter in primary (T0) transformants. Specific T2 plants produced 6.8% of dry weight as cyanophycin, which is more than five-fold higher than the previously published value. Although all lines tested were fertile, the progeny of the highest cyanophycin producing line had reduced seed production compared to control plants.

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