Multidimensional mapping of root responses to soil environmental cues using a luminescence-based imaging system

Root systems develop different root types that individually sense cues from their local environment and integrate them with systemic signals. This complex multi-dimensional amalgam of inputs leads to continuous adjustment of root growth rates, direction and metabolic activity to define a dynamic physical network. Current methods for analyzing root biology balance physiological relevance with imaging capability. To bridge this divide, we developed an integrated imaging system called Growth and Luminescence Observatory for Roots (GLO-Roots) that uses luminescence-based reporters to enable studies of root architecture and gene expression patterns in soil-grown, light-shielded roots. We have developed image analysis algorithms that allow the spatial integration of soil properties such as soil moisture with root traits. We propose GLO-Roots as a system that has great utility in both presenting environmental stimuli to roots in ways that evoke natural adaptive responses, and in providing tools for developing a multi-dimensional understanding of such processes.

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1. Multidimensional mapping of root responses to soil environmental cues ubén Rellán Álvarez bit.ly/glo-roots-preprint bit.ly/glo-roots-slides www.rrlab.org/glo_roots @rubenrellan rrellan@langebio.cinvestav.mx www.rrlab.org
2. Aaron Escobar: http://www.flickr.com/photos/aaronescobar/2569091622/
3. Aaron Escobar: http://www.flickr.com/photos/aaronescobar/2569091622/ water
4. Aaron Escobar: http://www.flickr.com/photos/aaronescobar/2569091622/ water Phosphate
5. Roots integrate a variety of signals and adjust function and shape accordingly Drew et al. 1978 JXB 109:435 Lima et al. 2011 PMID: 21119058 Tabata et al. 2014PMID: 25324386 AMT1:GFP
7. Yun et al. 2014 PMID: 24927545 From metabolite in vivo imaging to 3D in soil root architecture Grossman et al. 2011 PMID: 22186371 Soil Artificial Substrate
8. Yun et al. 2014 PMID: 24927545 From metabolite in vivo imaging to 3D in soil root architecture Grossman et al. 2011 PMID: 22186371 Soil Artificial Substrate
9. Soil A multidimensional map of root responses to soil environmental cues
10. Soil Time A multidimensional map of root responses to soil environmental cues
11. Soil Soil Properties Time A multidimensional map of root responses to soil environmental cues
12. Soil Soil Properties Time Gene Expression A multidimensional map of root responses to soil environmental cues
13. Soil Soil Properties Time Root Structure Gene Expression A multidimensional map of root responses to soil environmental cues
14. Soil Microbe Colonization Soil Properties Time Root Structure Gene Expression A multidimensional map of root responses to soil environmental cues
15. Soil Microbe Colonization Soil Properties Time Root Structure Gene Expression A multidimensional map of root responses to soil environmental cues
16. Transpiring environment Roots protected from light
17. 30 cm 15 cm 2 mm 100 cm3 of soil Rhizotrons: a 2D sample of soil
18. 30 cm 15 cm 2 mm 100 cm3 of soil Rhizotrons: a 2D sample of soil
19. The expression profile of roots grown in pots and rhizotrons is very similar Fresh Weight FW (mg) 0 60 120 180 240 MS Pot Rhizotron
20. Aaron Escobar: http://www.flickr.com/photos/aaronescobar/2569091622/ water Phosphate
21. Different environmental cues can be recreated and quantified in rhizotrons water
22. Different environmental cues can be recreated and quantified in rhizotrons water
23. Different environmental cues can be recreated and quantified in rhizotrons water
24. Different environmental cues can be recreated and quantified in rhizotrons water 80 60 40 20 0 moisture content (%)
25. Different environmental cues can be recreated and quantified in rhizotrons water 80 60 40 20 0 moisture content (%)
26. Different environmental cues can be recreated and quantified in rhizotrons water Phosphate 80 60 40 20 0 moisture content (%)
27. Different environmental cues can be recreated and quantified in rhizotrons water Phosphate 80 60 40 20 0 moisture content (%)
28. Different environmental cues can be recreated and quantified in rhizotrons water Phosphate 80 60 40 20 0 moisture content (%)
31. We use luciferase to visualize roots in soil www.firefly.org/firefly-pictures.html Luciferase is great! • Does not require any light excitation • No background luminescence • LUC can be used in temporal studies • Dynamic response to gene expression • Multiple color reporters are possible luciferin + ATP + O2 → oxyluciferin +
32. Roots are visualized with the GLO1 imaging system
33. Roots are visualized with the GLO1 imaging system
34. Both sides of the rhizotron are combined visualize the full root system
35. Both sides of the rhizotron are combined visualize the full root system
36. Root system architecture can be studied until late stages of development
37. Root system architecture can be studied until late stages of development
38. Images are data
39. Images are data
40. GLO-RIA is an ImageJ plugin developed to analyze root traits www.guillaumelobet.be
41. GLO-RIA is an ImageJ plugin developed to analyze root traits GLO-RIA Demo www.guillaumelobet.be
42. GLO-RIA enables semi-automatic quantification of RSA traits 11 12 13 14 15 16 17 18 19 20 21 Angle Count 0 45 90 0 45 90 0 45 90 0 45 90 0 45 90 0 45 90 0 45 90 0 45 90 0 45 90 0 45 90 0 45 90 Root system area
43. Different accessions show distinct root traits including lateral root angle
44. Different accessions show distinct root traits including lateral root angle
45. X Y DAS 0 21 Soil Soil Properties Time Root Structure water Phosphate
46. Root system architecture is light regulated
47. Root system architecture is light regulated
48. Root system architecture is light regulated
49. Root system architecture is light regulated
50. P-deficient plants have wider RSA and invest more resources in root growth 22 DAS 27 P P
51. Several degrees of water deficit can be induced
52. Several degrees of water deficit can be induced
53. Several degrees of water deficit can be induced
54. Early withdrawal of water may affect lateral root emergence
55. Early withdrawal of water may affect lateral root emergence
56. Early withdrawal of water may affect lateral root emergence
57. Early withdrawal of water may affect lateral root emergence
58. Early withdrawal of water may affect lateral root emergence
59. + 24 h Early withdrawal of water may affect lateral root emergence
60. If water deprivation starts later there is an induction of lateral root growth and a higher proliferation of tertiary roots
61. Lateral root angle is affected differently in different ecotypes in response to water deprivation
62. This response is different from hydrotropism and auxin dependent
63. Root system architecture can be revealed days after luciferin addition 16 17 18 19 20 21 22 23 24
64. Root system architecture can be revealed days after luciferin addition 16 17 18 19 20 21 22 23 24
65. Root system architecture can be revealed days after luciferin addition 16 17 18 19 20 21 22 23 24
66. Root direction can be locally correlated with soil moisture content WW WD
67. 500 540 580 620 660 700 UBQ10:vLUC2 ACT2:PpyREo We have generated a golden gate ready collection of different luciferases Intensity 1005 1064 1123 1182 1241 1300 UBQ10:CBRo Intensity 1005 1064 1123 1182 1241 1300 500 540 580 620 660 700 UBQ10:LUC2o 500 540 580 620 660 700 UBQ10:CBGo LUC2o Maximum Intensity vLUC CBG99o PpyRE8o CBRo λ (nm)
68. Microbe interactions imaged using luminescence emitting bacteria
69. Simultaneous imaging of root structure and gene is expression is possible ProACT2:PPyRE8o ProDR5rev:LUC+
70. Rapid changes in gene expression can be imaged ProACT2:PPyRE8o x ProZAT12:LUC ProZAT12:LUC
71. Rapid changes in gene expression can be imaged ProACT2:PPyRE8o x ProZAT12:LUC ProZAT12:LUC
72. Rapid changes in gene expression can be imaged ProACT2:PPyRE8o x ProZAT12:LUC ProZAT12:LUC GLO-RIA Demo
73. Soil Microbe Colonization Soil Properties Time Root Structure Gene Expression A multidimensional map of root responses to soil environmental cues
74. Soil Microbe Colonization Soil Properties Time Root Structure Gene Expression A multidimensional map of root responses to soil environmental cues
75. Soil Microbe Colonization Soil Properties Time Root Structure Gene Expression A multidimensional map of root responses to soil environmental cues
76. Dinneny Lab Muh-Ching Yee Geng Yu Emilie Winfield Pierre-Luc Pradier Charlotte Trontin Thérèse LaRue Heike Lindner Guillaume Lobet Image Analysis Jose Sebastian John Vogel Julin Maloof Other Species Cara Haney
79. Other species can also be imaged : Dual Color Tomato
80. Other species can also be imaged: Brachypodium dystachion