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Involvement of calmodulin in regulation of primary root elongation by N-3-oxo-hexanoyl homoserine lactone in Arabidopsis thaliana.

Zhao Q, Zhang C, Jia Z, Huang Y, Li H, Song S - Front Plant Sci (2015)

Bottom Line: In this study, we found that the stimulatory effect of N-3-oxo-hexanoyl homoserine lactone (3OC6-HSL) on primary root elongation of Arabidopsis was abolished by the calmodulin (CaM) antagonists N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7) and trifluoperazine (TFP).Nevertheless, our results suggest that CaM is involved in the bacterial 3OC6-HSL signaling in plant cells.These data offer new insight into the mechanism of plant response to bacterial QS signals.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, Biology Institute, Hebei Academy of Sciences Shijiazhuang, China ; Hebei Engineering and Technology Center of Microbiological Control on Main Crop Disease Shijiazhuang, China.

ABSTRACT
Many bacteria use signal molecules of low molecular weight to monitor their local population density and to coordinate their collective behavior in a process called "quorum sensing" (QS). N-acyl-homoserine lactones (AHLs) are the primary QS signals among Gram-negative bacteria. AHL-mediated QS plays an essential role in diverse bacterial physiological processes. Recent evidence shows that plants are able to sense bacterial AHLs and respond to them appropriately. However, little is known about the mechanism by which plants perceive and transduce the bacterial AHLs within cells. In this study, we found that the stimulatory effect of N-3-oxo-hexanoyl homoserine lactone (3OC6-HSL) on primary root elongation of Arabidopsis was abolished by the calmodulin (CaM) antagonists N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7) and trifluoperazine (TFP). Western-blot and ELISA analysis revealed that the concentration of CaM protein in Arabidopsis roots increased after treatment with 1 μM 3OC6-HSL. Results from quantitative RT-PCR demonstrated that the transcription of all nine CaM genes in Arabidopsis genome was up-regulated in the plants treated with 3OC6-HSL. The loss-of-function mutants of each AtCaM gene (AtCaM1-9) were insensitive to 3OC6-HSL-stimulation of primary root elongation. On the other hand, the genetic evidence showed that CaM may not participates the inhibition of primary root length caused by application of long-chained AHLs such as C10-HSL and C12-HSL. Nevertheless, our results suggest that CaM is involved in the bacterial 3OC6-HSL signaling in plant cells. These data offer new insight into the mechanism of plant response to bacterial QS signals.

No MeSH data available.


Related in: MedlinePlus

Effects of 3OC6-HSL on the accumulation of CaM protein in Arabidopsis roots. The plants were grown in hydroponic Hoagland medium with or without 1 μM 3OC6-HSL. Total protein was extracted from roots at different time interval after 3OC6-HSL treatment. (A) The change of CaM protein level detected by western blot. Actin was used as an internal quantification control. (B) The change of CaM protein level detected by ELISA using polyclonal antibody against wheat CaM. The results were normalized to the CaM protein level of untreated samples. Each value was the average of three independent experiments. Entries with p-values < 0.05 shown with asterisk.
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Figure 4: Effects of 3OC6-HSL on the accumulation of CaM protein in Arabidopsis roots. The plants were grown in hydroponic Hoagland medium with or without 1 μM 3OC6-HSL. Total protein was extracted from roots at different time interval after 3OC6-HSL treatment. (A) The change of CaM protein level detected by western blot. Actin was used as an internal quantification control. (B) The change of CaM protein level detected by ELISA using polyclonal antibody against wheat CaM. The results were normalized to the CaM protein level of untreated samples. Each value was the average of three independent experiments. Entries with p-values < 0.05 shown with asterisk.

Mentions: Pharmacological analysis implicates that CaM is required for 3OC6-HSL-mediated primary root elongation (Figure 3). Whether the level of CaM in plant cells is influenced by treatment with 3OC6-HSL has not been resolved. To address this point, we examined the level of CaM protein in Arabidopsis roots reacting to 3OC6-HSL. Total protein was extracted from Arabidopsis roots exposed to 1 μM 3OC6-HSL at different time points and the overall concentration of CaM protein was determined by western blot and ELISA using specific polyclonal antibody against CaM. Western analysis showed that the concentration of CaM protein in roots increased after treatment with 1 μM 3OC6-HSL and reached the maximum at 6 h post treatment (Figure 4A). Similarly, ELISA result indicated that treatment with 1 μM 3OC6-HSL promoted the accumulation of CaM protein and the CaM concentration reached a maximum 2-fold increase after 6 h of treatment with 3OC6-HSL (Figure 4B). These results provide the evidence for a positive regulation of 3OC6-HSL on CaM protein level.


Involvement of calmodulin in regulation of primary root elongation by N-3-oxo-hexanoyl homoserine lactone in Arabidopsis thaliana.

Zhao Q, Zhang C, Jia Z, Huang Y, Li H, Song S - Front Plant Sci (2015)

Effects of 3OC6-HSL on the accumulation of CaM protein in Arabidopsis roots. The plants were grown in hydroponic Hoagland medium with or without 1 μM 3OC6-HSL. Total protein was extracted from roots at different time interval after 3OC6-HSL treatment. (A) The change of CaM protein level detected by western blot. Actin was used as an internal quantification control. (B) The change of CaM protein level detected by ELISA using polyclonal antibody against wheat CaM. The results were normalized to the CaM protein level of untreated samples. Each value was the average of three independent experiments. Entries with p-values < 0.05 shown with asterisk.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4292405&req=5

Figure 4: Effects of 3OC6-HSL on the accumulation of CaM protein in Arabidopsis roots. The plants were grown in hydroponic Hoagland medium with or without 1 μM 3OC6-HSL. Total protein was extracted from roots at different time interval after 3OC6-HSL treatment. (A) The change of CaM protein level detected by western blot. Actin was used as an internal quantification control. (B) The change of CaM protein level detected by ELISA using polyclonal antibody against wheat CaM. The results were normalized to the CaM protein level of untreated samples. Each value was the average of three independent experiments. Entries with p-values < 0.05 shown with asterisk.
Mentions: Pharmacological analysis implicates that CaM is required for 3OC6-HSL-mediated primary root elongation (Figure 3). Whether the level of CaM in plant cells is influenced by treatment with 3OC6-HSL has not been resolved. To address this point, we examined the level of CaM protein in Arabidopsis roots reacting to 3OC6-HSL. Total protein was extracted from Arabidopsis roots exposed to 1 μM 3OC6-HSL at different time points and the overall concentration of CaM protein was determined by western blot and ELISA using specific polyclonal antibody against CaM. Western analysis showed that the concentration of CaM protein in roots increased after treatment with 1 μM 3OC6-HSL and reached the maximum at 6 h post treatment (Figure 4A). Similarly, ELISA result indicated that treatment with 1 μM 3OC6-HSL promoted the accumulation of CaM protein and the CaM concentration reached a maximum 2-fold increase after 6 h of treatment with 3OC6-HSL (Figure 4B). These results provide the evidence for a positive regulation of 3OC6-HSL on CaM protein level.

Bottom Line: In this study, we found that the stimulatory effect of N-3-oxo-hexanoyl homoserine lactone (3OC6-HSL) on primary root elongation of Arabidopsis was abolished by the calmodulin (CaM) antagonists N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7) and trifluoperazine (TFP).Nevertheless, our results suggest that CaM is involved in the bacterial 3OC6-HSL signaling in plant cells.These data offer new insight into the mechanism of plant response to bacterial QS signals.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, Biology Institute, Hebei Academy of Sciences Shijiazhuang, China ; Hebei Engineering and Technology Center of Microbiological Control on Main Crop Disease Shijiazhuang, China.

ABSTRACT
Many bacteria use signal molecules of low molecular weight to monitor their local population density and to coordinate their collective behavior in a process called "quorum sensing" (QS). N-acyl-homoserine lactones (AHLs) are the primary QS signals among Gram-negative bacteria. AHL-mediated QS plays an essential role in diverse bacterial physiological processes. Recent evidence shows that plants are able to sense bacterial AHLs and respond to them appropriately. However, little is known about the mechanism by which plants perceive and transduce the bacterial AHLs within cells. In this study, we found that the stimulatory effect of N-3-oxo-hexanoyl homoserine lactone (3OC6-HSL) on primary root elongation of Arabidopsis was abolished by the calmodulin (CaM) antagonists N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7) and trifluoperazine (TFP). Western-blot and ELISA analysis revealed that the concentration of CaM protein in Arabidopsis roots increased after treatment with 1 μM 3OC6-HSL. Results from quantitative RT-PCR demonstrated that the transcription of all nine CaM genes in Arabidopsis genome was up-regulated in the plants treated with 3OC6-HSL. The loss-of-function mutants of each AtCaM gene (AtCaM1-9) were insensitive to 3OC6-HSL-stimulation of primary root elongation. On the other hand, the genetic evidence showed that CaM may not participates the inhibition of primary root length caused by application of long-chained AHLs such as C10-HSL and C12-HSL. Nevertheless, our results suggest that CaM is involved in the bacterial 3OC6-HSL signaling in plant cells. These data offer new insight into the mechanism of plant response to bacterial QS signals.

No MeSH data available.


Related in: MedlinePlus