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Acta Agronomica Sinica ›› 2024, Vol. 50 ›› Issue (12): 2962-2970.doi: 10.3724/SP.J.1006.2024.42021

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

Overexpression of OsPIN2 increases tiller angle by reducing shoot gravitropic response in rice

WU Jia-Jun(), TU Ran-Ran, ZHANG Qiu-Li, ZOU Qin-Wen, SUN Zhi-Hao, WANG Hong, HE Guang-Hua()   

  1. Rice Research Institute, Southwest University / Academy of Agricultural Sciences, Southwest University / Key Laboratory of Crop Molecular Improvement, Chongqing 400715, China
  • Received:2024-04-22 Accepted:2024-08-15 Online:2024-12-12 Published:2024-08-29
  • Contact: *E-mail: heghswu@163.com
  • Supported by:
    National Natural Science Foundation of Chongqing, China(cstc2021jcyj-cxttX0004);Chongqing Modern Agricultural Industry Technology System Rice Innovation Team of Chongqing, China(CQMAITS202301)

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Abstract:

Shoot gravitropism is closely related to the formation of the tiller angle, and understanding its regulatory mechanism is crucial for rationally designing tiller angles to improve crop plant architecture. In the rice auxin efflux carrier gene OsPIN2 overexpression lines, two lines with significantly increased tiller angles, OE-OsPIN2-1 and OE-OsPIN2-2, were identified. Scanning electron microscopy revealed that the near-ground and far-ground parts of the tiller bases in OE-OsPIN2-1/2 grew nearly symmetrically. Shoot gravitropism assays indicated that the shoot gravitropic responses of OE-OsPIN2-1/2 seedlings were reduced, and the asymmetric expression of the auxin marker gene OsIAA20 and WUSCHEL RELATED HOMEOBOX6/11 (WOX6/11) was weakened upon gravistimulation. This suggests that overexpression of OsPIN2 attenuates the asymmetric distribution of auxin following gravistimulation. Furthermore, the expressions of positive regulators involved in the gravitropic response at the tiller base in OE-OsPIN2-1/2 lines were downregulated, while the expressions of negative regulators were upregulated. This further indicates that overexpression of OsPIN2 leads to a reduced shoot gravitropic response. This study elucidates the mechanism by which OsPIN2 controls rice tiller angle by regulating shoot gravitropism, providing a theoretical basis for in-depth studies of shoot gravitropic responses.

Key words: rice, OsPIN2, tiller angle, shoot gravitropism

Table 1

Primers used in this study"

名称 Name 上游引物 Upstream primer (5′-3′) 下游引物 Downstream primer (5′-3′)
OsPIN2-OE GCAGGATCCCCGGGTACCATGATCACCGGACGCGAC AATGTTTGAACGGAGCTCCTATATCCCAAGAAGCACATAGTAG
OsPIN2-qPCR CAACACCTACTCCAGCCTC TGGACCAGTCAAGAACCTC
UBQ-qPCR GCTCCGTGGCGGTATCAT CGGCAGTTGACAGCCCTAG
OsIAA20-qPCR TGGCGGATATGTGAAGGTGAA TATGAGCCGAGGATGGACAAG
WOX6-qPCR TCCAATAGACTTGCGAGCCAT GCATTAGGATTCCATAGTCGTT
WOX11-qPCR CGGTGTTCATCAACGGAGTG TCTGGAGAGAATGGAGGAGGAT
CRCT-qPCR TTCTGGGTGCCTCAACTCA AACGCTGTCTCAAAGTCCAATC
FucT-qPCR GGAGTCTGCTGTGCTTGCTA ACTGGTATAATGCCTGTCGTTGTG
AGPL1-qPCR TTGATTCCACATGGCAGAGAAC GTTGCTGCTGCTACTTCACT
OsHOX1-qPCR AGCACAACACCCTCAATC GTTCTGGAACCACACCTC
OsHOX28-qPCR CATTGACCACCCTCACAA CATTGACCACCCTCACAA
LPA1-qPCR GCGTATGTATGTAAAGCAAG GAAACGACCTACGAAACTAC
LA1-qPCR GAGATGAACGGCAACAAG TTCCAGCACCAAGTAGTC
LA2-qPCR GAACCAGCAGCCTGTAAGA AGCCATCCTCTCCTTCATTG
LA3-qPCR GGCGTCGCGCCAGCCTCA CTTCGGCGGAGTATCACG
TAC1-qPCR AGATGGCTCTAAAGGTGTTCAA TCTTCCATGGCCTTGTTCTC
TAC4-qPCR AAGGTCGCAAACAAGCAG AACTGCCAGGAGCAGAGAG
HSFA2D-qPCR CAGCAGGCACTTGGCACC TTCTTGTCACGCTTTAGCCTGT
OsARF12-qPCR GTTGGGAGGTCGTTGGACATAA AAGCACATCATTCTCCCTGTCG
OsARF17-qPCR TTTACAAATCGGGAACCTATGG TTTATGCAGGAGACGCTATTCA
OsARF25-qPCR TGACATCTCCAGATTCAGCAGC CGTCTCCACCACGAACCAA

Fig. 1

Phenotypic analysis of WT and OE-OsPIN2-1/2 lines A: comparison of WT and OE-OsPIN2-1/2 plants at tillering stage, with scale bars of 20 cm; B: expression analysis of OsPIN2 in WT and OE-OsPIN2-1/2 tiller bases (n = 3); C: comparison of tiller angle of WT and OE-OsPIN2-1/2 lines, with scale bars of 4 cm; D: measurements of tiller angle of WT and OE-OsPIN2-1/2 lines (n = 10). Data are shown as means ± SD, two-tailed Student’s t-test, **: P < 0.01."

Fig. 2

Analysis of tiller base of WT and OE-OsPIN2-1/2 plants A-C: comparison of tiller base between WT and OE-OsPIN2-1/2 lines at heading stage, with scale bars of 1 cm; D-F: scanning electron microscopy of NG cells of tiller base in WT and OE-OsPIN2-1/2 lines, with scale bars of 50 μm; G-I: scanning electron microscopy of FG cells of tiller base in WT and OE-OsPIN2-1/2 lines, with scale bars of 50 μm; J-K: measurements of cell length (J) and width (K) in NG and FG cells of tiller base in WT and OE-OsPIN2-1/2 lines (n = 20); L: cell number per unit area in NG and FG cells of tiller base in wild type and OE-OsPIN2-1/2 (n = 3). NG represents near-ground part, FG represents far-ground part. Means ± SD, two-tailed Student’s t-test, **: P < 0.01."

Fig. 3

Shoot gravitropism assay of WT and OE-OsPIN2-1/2 seedlings A: comparison of WT and OE-OsPIN2-1/2 seedlings at 0 h, 12 h, 24 h, 36 h, and 48 h after gravitropic stimulation; B: measurements of shoot curvature of wild type and OE-OsPIN2-1/2 seedlings at 0 h, 12 h, 24 h, 36 h, and 48 h after gravitropic stimulation (n = 5). Data are shown as means ± SD. Multiple comparisons were performed using Duncan’s test, and different letters (lowercase letters a and b in B) indicate significant differences at the 0.05 level."

Fig. 4

Expression analysis of OsIAA20 and WOX6/11 before and after gravity treatment, and of tiller angle-related genes in the tiller base A: a schematic diagram depicting the lower site (LS) and upper site (US) of the shoot base in seedlings after gravity treatment; ‘g’ represents gravity, and arrows indicate the direction of gravity; B: analysis of the transcription levels of the auxin marker gene OsIAA20 and WUSCHEL-Related HOMEOBOX 6/11 (WOX6/11) in the lower and upper sites of the shoot base of WT and OE-OsPIN2-1/2 seedlings at 0 h and 6 h after gravity treatment; C: analysis of the expression levels of tiller angle-related genes in the tiller base of WT and OE-OsPIN2-1/2 at the tillering stage; D: network planning of shoot gravitropism regulating rice tiller angle; the red upward arrows indicate upregulation of the gene expression, while the red downward arrows indicate downregulation of the gene expression. Means ± SD (n = 3), two-tailed Student’s t-test, **: P < 0.01."

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