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Acta Agronomica Sinica ›› 2024, Vol. 50 ›› Issue (7): 1647-1657.doi: 10.3724/SP.J.1006.2024.31077

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• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Effects of high light stress on photosynthesis and physiological characteristics of wheat with maize C4-type ZmPEPC+ZmPPDK gene

FANG Yu-Hui(), QI Xue-Li, LI Yan, ZHANG Yu, PENG Chao-Jun, HUA Xia, CHEN Yan-Yan, GUO Rui, HU Lin, XU Wei-Gang*()   

  1. Institute of Crop Molecular Breeding, Henan Academy of Agricultural Sciences / Key Laboratory for Wheat Biology of Henan Province / Shennong Laboratory, Zhengzhou 450002, Henan, China
  • Received:2023-12-01 Accepted:2024-01-30 Online:2024-07-12 Published:2024-02-20
  • Contact: *E-mail: xuwg1958@163.com
  • Supported by:
    Independent Innovation Foundation of Henan Academy of Agricultural Sciences(2023ZC090);Henan Provincial Science and Technology Research Project(232102110203);‘First-class Project’ of Shennong Laboratory(SN01-2022-01);Major Project of Agricultural Biological Breeding(2023ZD0402302);China Agriculture Research System of MOF and MARA(CARS-03-7)

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Abstract:

To study the photosynthetic and physiological response of maize C4-type PEPC (phosphoenolpyruvate carboxylase gene) and PPDK (pyruvate phosphate double kinase gene) dual-gene wheat to high light, the ZmPEPC + ZmPPDK wheat lines PCK30 and PCK60 and their wild type control material (WT) were used as experimental materials. The relative expression levels of exogenous genes in transgenic wheat were identified. The photosynthetic enzyme activity, chlorophyll content, gas exchange parameters, chlorophyll fluorescence parameters, active oxygen content, and antioxidant enzyme activity of transgenic wheat were measured under normal light intensity (NL) and high light stress (HL) at heading and grain-filling stages. The results showed that the two transgenic lines expressed the PEPC and PPDK genes efficiently at the transcriptional level. The enzyme activity of PEPC, PPDK, NADP-ME, and Rubisco of transgenic wheat was significantly higher than WT under NL or HL stress at different stage, and the increase of WT was more obvious under HL stress. Compared with NL stress, the chlorophyll content of transgenic wheat and WT decreased significantly under HL stress, but the decrease of transgenic lines was smaller, and the chlorophyll content of transgenic lines was significantly higher than WT under HL stress. Under two treatment stresses, the net photosynthetic rate (Pn) of PCK30 and PCK60 was significantly higher than WT, and the higher range was more obvious under HL stress. The increases were 15.26% and 17.57% at heading stage and 13.41% and 15.82% at grain-filling stage, respectively. The variation trend of stomatal conductance, Fv/Fm and qp was consistent with Pn, while the variation trend of intercellular carbon dioxide concentration was opposite to Pn. The content of reactive oxygen species and malondialdehyde produced by transgenic lines under HL stress was significantly lower than WT, while the trend of antioxidant enzymes was opposite. In two consecutive years of field plot yield experiments, PCK30 and PCK60 were 8.37% and 10.16% higher than WT on average. The overexpression of PEPC and PPDK in wheat enhanced the endogenous photosynthetic enzyme, photochemical efficiency, and antioxidant enzyme activities of wheat, enhanced the stability of leaf cell membrane under high light stress, protected the photosynthetic apparatus, and maintained a high photosynthetic efficiency, thus improving the tolerance of transgenic wheat to high light stress.

Key words: wheat, maize C4-type photosynthetic gene, high light stress, photosynthesis, physiological characteristics

Fig. 1

PCR analysis of ZmPEPC in ZmPEPC and ZmPPDK transgenic wheats M: marker DL2000 plus; 1: positive control; 2: wile type control (Zhengmai 1860); 3-6: PCK30 transgenic plants; 7-9: PCK60 transgenic plants."

Fig. 2

PCR analysis of ZmPPDK in ZmPEPC and ZmPPDK transgenic wheats M: marker DL2000 plus; 1: positive control; 2: wile type control (Zhengmai 1860); 3-6: PCK30 transgenic plants; 7-9: PCK60 transgenic plants."

Fig. 3

Relative expression level of ZmPEPC and ZmPPDK in flag leaves of transgenic wheat lines and wild type Lowercases letters indicate significant difference at the 0.05 probability level. Error bar: ±SE."

Fig. 4

PEPC, PPDK, NADP-ME, and Rubisco activities in flag leaves of transgenic wheat lines and wild type Lowercases letters indicate significant difference at the 0.05 probability level. Error bar: ±SE."

Table 1

Chlorophyll content in flag leaves of transgenic wheat and control (WT) under high light"

叶绿素含量
Chlorophyll content		 材料
Material		 抽穗期 Heading 灌浆期 Grain-filling
NL HL NL HL
总叶绿素含量
Total chlorophyll content		 WT 5.046±0.132 a 4.197±0.157 c 5.071±0.112 b 4.078±0.127 d
PCK30 5.271±0.243 a 4.704±0.223 b 5.418±0.151 a 4.676±0.178 c
PCK60 5.253±0.108 a 4.595±0.176 b 5.360±0.127 a 4.760±0.223 c
叶绿素a/b
Chlorophyll a/b		 WT 2.073±0.068 a 1.453±0.047 c 1.614±0.065 b 1.334±0.047 c
PCK30 2.141±0.109 a 1.650±0.075 b 1.742±0.076 a 1.554±0.037 b
PCK60 2.149±0.078 a 1.635±0.056 b 1.776±0.097 a 1.617±0.066 b

Table 2

Gas exchange parameter in flag leaves of transgenic wheat and control (WT) under high light"

气体交换参数
Gas exchange parameter		 材料
Material		 抽穗期 Heading 灌浆期 Grain-filling
NL HL NL HL
Pn (μmol CO2 m-2 s-1) WT 26.32±0.61 b 20.35±0.99 d 24.79±0.92 b 20.18±1.14 d
PCK30 28.14±0.88 a 23.46±1.58 c 26.94±0.75 a 22.89±0.76 c
PCK60 28.48±0.92 a 23.93±0.65 c 27.23±0.76 a 23.37±1.04 bc
Gs (mmol H2O m-2 s-1) WT 287.25±10.60 a 225.96±7.21 c 297.67±4.39 b 243.00±7.27 d
PCK30 296.67±7.30 a 261.25±9.46 b 323.67±6.90 a 273.25±13.67 c
PCK60 298.00±12.05 a 261.67±12.27 b 319.96±10.17 a 276.33±6.44 c
Tr (mmol H2O m-2 s-1) WT 4.76±0.16 a 3.60±0.15 c 5.44±0.15 b 4.32±0.20 d
PCK30 4.88±0.16 a 4.32±0.18 b 5.80±0.13 a 4.92±0.16 c
PCK60 4.89±0.22 a 4.27±0.14 b 5.78±0.21 a 4.95±0.25 c
Ci (μmol mol-1) WT 248.00±9.36 c 290.67±5.06 a 241.67±6.30 bc 273.67±4.88 a
PCK30 236.25±10.61 c 265.33±10.54 b 226.96±7.69 d 255.00±8.80 b
PCK60 234.67±7.98 c 269.96±8.47 b 228.33±10.22 d 252.67±7.61 b

Table 3

Chlorophyll fluorescence parameter in flag leaves of transgenic wheat and control (WT) under high light"

叶绿素荧光参数
Chlorophyll fluorescence parameters		 材料
Material		 抽穗期 Heading 灌浆期 Grain-filling
NL HL NL HL
Fv/Fm WT 0.833±0.016 b 0.714±0.019 d 0.821±0.028 b 0.710±0.012 d
PCK30 0.886±0.025 a 0.785±0.008 c 0.873±0.012 a 0.779±0.019 c
PCK60 0.884±0.013 a 0.794±0.016 c 0.870±0.021 a 0.802±0.013 bc
qp WT 0.705±0.019 b 0.548±0.012 c 0.692±0.028 b 0.497±0.014 d
PCK30 0.813±0.020 a 0.691±0.026 b 0.750±0.011 a 0.599±0.032 c
PCK60 0.822±0.015 a 0.668±0.034 b 0.748±0.026 a 0.614±0.015 c

Table 4

Reactive oxygen species and MDA content in flag leaves of transgenic wheat and control (WT) under high light"

指标
Index		 材料
Material		 抽穗期 Heading 灌浆期 Grain-filling
NL HL NL HL
MDA含量
MDA content (μmol g-1 FW)		 WT 24.51±0.42 b 31.50±1.35 a 27.95±0.64 c 34.15±0.98 a
PCK30 22.88±0.64 c 26.20±0.92 b 25.60±0.58 d 29.27±1.26 bc
PCK60 22.50±0.55 c 26.11±1.18 b 26.01±0.71 d 29.87±1.57 b
超氧阴离子生成速率
Production rate of superoxide anion (nmol min-1 g-1 FW)		 WT 20.38±0.41 c 25.61±0.89 a 26.11±0.35 c 34.27±1.15 a
PCK30 19.29±0.43 d 22.21±0.75 b 24.48±0.56 d 30.67±1.33 b
PCK60 18.88±0.55 d 20.90±1.24 bc 24.46±0.42 d 30.93±1.38 b
H2O2含量
H2O2 content (μmol g-1 FW)		 WT 11.62±0.38 c 15.85±0.58 a 16.48±0.66 c 21.06±0.67 a
PCK30 10.71±0.52 d 14.04±0.64 b 15.06±0.35 d 18.06±0.52 b
PCK60 10.66±0.44 d 13.53±0.43 b 14.92±0.46 d 17.46±0.86 bc

Table 5

Antioxidant enzyme activity in flag leaves of transgenic wheat and control (WT) under high light (U g-1)"

抗氧化酶
Antioxidant enzyme		 材料
Material		 抽穗期 Heading 灌浆期 Grain-filling
NL HL NL HL
SOD
 WT 319.48±8.20 c 401.39±12.51 b 509.36±6.22 d 624.12±15.76 b
PCK30 332.90±10.48 c 463.13±10.36 a 541.09±12.37 c 710.31±22.15 a
PCK60 334.75±13.35 c 472.16±17.77 a 549.34±16.23 c 720.61±17.50 a
POD
 WT 179.75±8.21 c 236.60±7.94 b 221.83±6.84 d 271.81±10.58 b
PCK30 187.53±11.36 c 266.46±13.21 a 240.71±7.61 c 303.47±8.63 a
PCK60 186.60±9.48 c 268.24±10.25 a 240.33±10.48 c 300.95±12.66 a
CAT
 WT 50.83±2.28 c 61.54±2.45 b 62.46±2.24 c 72.40±2.57 b
PCK30 52.24±3.15 c 71.23±1.86 a 69.49±1.66 b 84.52±3.22 a
PCK60 53.20±1.24 c 70.36±1.59 a 70.84±3.18 b 85.08±2.64 a

Table 6

Plot yield and its composition of transgenic wheat and control (WT)"

年度
Year		 材料
Material		 穗数
Spike number
(×104 hm-2)		 穗粒数
Grains per spike		 千粒重
1000-kernel weight
(g)		 产量
Grain yield
(kg hm-2)
2021-2022 WT 423.59±18.32 a 35.33±2.42 a 51.27±0.67 b 7551.12±167.45 b
PCK30 429.83±20.37 a 35.17±2.18 a 54.03±0.95 a 8168.27±204.42 a
PCK60 428.60±13.03 a 35.71±1.64 a 54.18±0.62 a 8291.38±162.33 a
2022-2023 WT 470.11±21.72 a 34.84±2.32 a 50.09±0.65 b 8062.50±233.39 b
PCK30 477.76±17.30 a 34.75±1.04 a 52.72±0.54 a 8752.81±139.63 a
PCK60 471.65±24.51 a 35.62±2.19 a 53.04±0.71 a 8910.90±199.02 a
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