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Open Access Highly Accessed Research article

Rapid and dynamic subcellular reorganization following mechanical stimulation of Arabidopsis epidermal cells mimics responses to fungal and oomycete attack

Adrienne R Hardham1*, Daigo Takemoto12 and Rosemary G White3

Author Affiliations

1 Plant Cell Biology Group, Research School of Biological Sciences, The Australian National University, Canberra, ACT 2601, Australia

2 Plant Pathology Laboratory, Graduate School of Bioagricultural Sciences, Nagoya University, Chikusa, Nagoya, 464-8601, Japan

3 Division of Plant Industry, C.S.I.R.O., Canberra, ACT 2601, Australia

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BMC Plant Biology 2008, 8:63  doi:10.1186/1471-2229-8-63

Published: 2 June 2008

Additional files

Additional file 1:

Rapid development of a patch of actin microfilaments at the contact site. Movie composed of 94 images of the actin array underneath the point of contact with a glass microneedle in A. thaliana expressing hTalin-GFP. Each image in the movie is a projection of six optical sections through the cortical cytoplasm underlying the outer epidermal cell wall. The movie commences at the time of touching the cell surface with the needle and ends 1 hour and 5 minutes later. The time between most images is about 40 seconds. Selected images from the movie are illustrated in Fig. 1. The movie plays at about 200 times real-time.

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Additional file 2:

Formation, dispersal and re-formation of an actin patch at the contact site. Actin microfilaments visualized in the cortical cytoplasm underlying the outer epidermal cell wall in a cotyledon of A. thaliana expressing hTalin-GFP. The surface of the epidermal cell was touched with a glass microneedle at the site indicated by the asterisk in A. Times in minutes and seconds show elapsed time after the image in A. Images A-C show formation of a patch of actin microfilaments about 6 minutes after touching the cell surface. Needle contact was made about 10 minutes after the image in A was taken. Images D-F show dispersal of the actin patch after the needle lifted off the cotyledon. Images G-I show reformation of the actin patch when the needle was again brought into contact with the cell at the same location. The image in G was taken 1 minute after re-positioning the needle to touch the surface again. Images are projections of 7 (B, G-I), 8 (C, E), 9 (D), 12 (F) or 13 (A) optical sections. Bar = 10 μm.

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Additional file 3:

Focusing of actin cables on the contact site. Movie composed of 53 images of the actin array underneath the point of contact with a tungsten microneedle in A. thaliana expressing hTalin-GFP. Each image in the movie is a projection of six optical sections through the cortical cytoplasm underlying the outer epidermal cell wall. The position of the needle tip is indicated by the vertical line of reflected light. The movie commences just before touching the cell surface with the needle and ends 28 min later. The time between most images is 20–25 s. During the second half of the movie sequence, actin cables become focused on the contact site. The movie plays at about 150 times real-time. Selected images from the sequence are illustrated in Fig. 2.

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Additional file 4:

Dynamics of a bright cloud of actin at the point of contact. Movie composed of 73 images of the actin array underneath the point of contact with a tungsten microneedle in A. thaliana expressing hTalin-GFP. Each image in the movie is a projection of five optical sections through the cortical cytoplasm underlying the outer epidermal cell wall. The position of the needle is indicated by the V-shaped shadow. The movie commences just before touching the cell surface with the needle and ends 28 minutes later. The time between most images is about 20 seconds. Accumulation of actin below the point of contact forms a bright patch of fluorescence that appears to swirl around the needle tip. The movie plays at about 150 times real-time.

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Additional file 5:

Changes in the microtubule array beneath the point of contact. Movie composed of 115 images of the microtubule array underneath the point of contact with a tungsten needle in A. thaliana expressing TUA6-GFP [70]. Each image in the movie is a projection of five optical sections through the cortical cytoplasm underlying the outer epidermal cell wall. The position of the needle is indicated by the V-shaped shadow (top centre). The movie commences just before touching the cell surface with the needle and ends 1 hour 17 minutes later. The time between most images is about 30–40 seconds. A bright diffuse cloud of GFP-tubulin begins to accumulate at the point of contact about 4 min after touching the cell with the needle. Linear arrays of diffuse fluorescence appear to sweep below the cortical microtubule array. The movie plays at about 200 times real-time.

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Additional file 6:

Dynamic changes in the microtubule array as the contact point moves across the cell wall. Movie composed of 95 images of the microtubule array underneath the point of contact with a tungsten needle in A. thaliana expressing TUA6-GFP. Each image in the movie is a projection of five optical sections through the cortical cytoplasm underlying the outer epidermal cell wall. The position of the needle is indicated by the V-shaped shadow (top centre). The movie commences just before touching the cell surface with the needle and ends 1 hour 11 minutes later. The time between most images is about 30–40 seconds. A bright diffuse cloud of GFP-tubulin begins to accumulate at the point of contact about 3.5 minutes after touching the cell with the needle. As the needle drifts across the cotyledon surface, the cloud of concentrated GFP-tubulin moves with it to remain beneath the point of contact. When the needle tip moves from one cell to an adjacent cell the fluorescent cloud dissipates in the first cell and forms in the second. The movie plays at about 200 times real-time.

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Additional file 7:

Dynamics of the microtubule array in a control cotyledon. Movie composed of 100 images of the microtubule array underneath the point of contact with a tungsten needle in A. thaliana expressing TUA6-GFP. All images in the movie are single optical sections in the same focal plane taken at intervals of 3–4 seconds for a period of 6 minutes. Against a framework of stable microtubules in the cell cortex just below the cell wall, mobile, linear strands of diffuse fluorescence sweep through the cell beneath the cortical array. The movie plays at about 18 times real-time.

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Additional file 8:

Dynamics of the ER in epidermal cells in control cotyledons. ER in the cortical cytoplasm underlying the outer epidermal cell wall in a cotyledon of Arabidopsis thaliana expressing GFP-KKXX [71]. The cotyledon has been mounted onto a microscope slide but had not been touched with a microprobe. In general, the network of ER is stable and its organization does not change during the observation period. However, some transient flaring of strands of diffuse fluorescence occurs in the top right hand corner of the cell in the first three images (arrowheads) and near the left hand side of the cell in the images taken at 4 minutes 18 seconds and 5 minutes 12 seconds (arrows). Bar = 10 μm.

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Additional file 9:

Dynamic changes in the organization of the cortical ER beneath the contact site. Movie composed of 77 images of the ER in an epidermal cell touched with a tungsten needle in A. thaliana expressing GFP-KKXX. Each image in the movie is a projection of five optical sections; the interval between each z-series is about 35 seconds. The movie begins when the cell is touched with the needle and shows images collected over the next 52 minutes. Bright fluorescence indicative of aggregated ER accumulates beneath the contact site. The movie plays at about 200 times real-time.

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Additional file 10:

Clustering of peroxisomes at the site of contact with the microneedle. Movie composed of 43 images of peroxisomes in an epidermal cell touched with a tungsten needle in A. thaliana expressing PTS-GFP [46]. Each image in the movie is a projection of three optical sections and the interval between each z-series is about 15 seconds. The movie begins when the cell is touched with the needle and shows images collected over the next 14 minutes. About 5 minutes after touching the cotyledon, peroxisomes begin to cluster beneath the tip of the needle. Selected images from the first 8 minutes of observation are shown in Fig. 6. The movie plays at about 100 times real-time. Selected images from this sequence are shown in Fig. 7.

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