Limits...
Pleurotus ostreatus manganese-dependent peroxidase silencing impairs decolourization of Orange II.

Salame TM, Yarden O, Hadar Y - Microb Biotechnol (2009)

Bottom Line: Relative real-time PCR quantification analysis confirmed that all the nine genes are transcribed, and that Mn(2+) amendment results in a drastic increase in the transcript levels of the predominantly expressed MnP genes (mnp 3 and mnp 9), while decreasing versatile peroxidase gene transcription (mnp 4).Knock-down of mnp 3 resulted in the reduction of fungal OII decolourization capacity, which was co-linear with marked silencing of the Mn(2+)-dependent peroxidase genes mnp 3 and mnp 9.This is the first direct genetic proof of an association between MnP gene expression levels and azo dye decolourization capacity in P. ostreatus, which may have significant implication on understanding the mechanisms governing lignin biodegradation.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Pathology and Microbiology, The Robert H. Smith Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem, Rehovot 76100, Israel.

Show MeSH

Related in: MedlinePlus

Relative transcript abundance of MnP gene family members in RNA extracted from P. ostreatus PC9 transformants grown for 7 days in liquid GP medium amended with 27 µM Mn2+ (+Mn treatment), compared with that of the TC3 strain grown in non‐amended medium (−Mn treatment). The abundance of mnp1–9 genes transcripts, relative to β‐tubulin, was measured by real‐time PCR, and quantified by the ΔΔCT method. TC3 – control strain (pTM1 transformant); TS1, TS9, TS24, TS30 – Mn2+‐dependent peroxidase silenced strains (pTMS1 transformants). Data represent the average of three biological replicates. Bars denote the standard deviation; when not visible, the standard deviation is included within the graph line width. Note the log10 scale, and different axis ranges.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3815951&req=5

f6: Relative transcript abundance of MnP gene family members in RNA extracted from P. ostreatus PC9 transformants grown for 7 days in liquid GP medium amended with 27 µM Mn2+ (+Mn treatment), compared with that of the TC3 strain grown in non‐amended medium (−Mn treatment). The abundance of mnp1–9 genes transcripts, relative to β‐tubulin, was measured by real‐time PCR, and quantified by the ΔΔCT method. TC3 – control strain (pTM1 transformant); TS1, TS9, TS24, TS30 – Mn2+‐dependent peroxidase silenced strains (pTMS1 transformants). Data represent the average of three biological replicates. Bars denote the standard deviation; when not visible, the standard deviation is included within the graph line width. Note the log10 scale, and different axis ranges.

Mentions: In order to determine whether alterations in MnP gene family transcript levels accompany the changes observed in OII decolourization we used relative real‐time PCR quantification analysis. The selected strains (TS1, TS9, TS24, TS30 and the TC3 control) were grown for 7 days in liquid culture, in the presence of carboxin, in either a medium amended with 27 µM Mn2+ (+Mn treatment) or a non‐amended medium (−Mn treatment). The results presented in Fig. 6 show the relative expression of the nine different members of the MnP gene family in the five selected strains. The overall abundance of the endogenous control gene (β‐tubulin) transcript was found to be similar in all the strains in all the tested treatments. As TC3 exhibited similar gene expression levels to the PC9 wild‐type strain, we used the former, in the −Mn treatment, as our calibrators for relative quantifications. The results obtained (Fig. 6) indicate that the predominantly expressed MnP gene family members expression in the pTMS1 transformants varied in intensity in response to Mn2+ amendment, similarly to the calibrator (TC3) and PC9 (Fig. 2), but at different levels. Specifically, relatively to the calibrator, mnp3 and mnp9 gene expression, though not substantially affected in the −Mn treatment (less than twofold difference), were strongly inhibited in the +Mn treatment which exhibited 24‐ to 79‐fold higher levels in the calibrator. Therefore, mnp3 and mnp9 were not even remotely upregulated at levels comparable to the control in response to Mn2+ amendment. mnp4 gene expression was not substantially affected in the −Mn treatment (less than 2.2‐fold difference); however, in the +Mn treatment it was 8‐ to 24‐fold higher in the calibrator; mnp8 gene expression was not substantially affected in the −Mn treatment (less than 2 fold difference), but in the +Mn treatment it was 4‐ to 9‐fold higher in the calibrator. Thus, in contrast to the striking results obtained in the case of mnp3, 4 and 9, the transcript abundance of mnp1, 2, 5, 6 and 7 was not substantially affected (less than fourfold difference), generally showing the same trend as the calibrator both in the −Mn and +Mn treatments.


Pleurotus ostreatus manganese-dependent peroxidase silencing impairs decolourization of Orange II.

Salame TM, Yarden O, Hadar Y - Microb Biotechnol (2009)

Relative transcript abundance of MnP gene family members in RNA extracted from P. ostreatus PC9 transformants grown for 7 days in liquid GP medium amended with 27 µM Mn2+ (+Mn treatment), compared with that of the TC3 strain grown in non‐amended medium (−Mn treatment). The abundance of mnp1–9 genes transcripts, relative to β‐tubulin, was measured by real‐time PCR, and quantified by the ΔΔCT method. TC3 – control strain (pTM1 transformant); TS1, TS9, TS24, TS30 – Mn2+‐dependent peroxidase silenced strains (pTMS1 transformants). Data represent the average of three biological replicates. Bars denote the standard deviation; when not visible, the standard deviation is included within the graph line width. Note the log10 scale, and different axis ranges.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3815951&req=5

f6: Relative transcript abundance of MnP gene family members in RNA extracted from P. ostreatus PC9 transformants grown for 7 days in liquid GP medium amended with 27 µM Mn2+ (+Mn treatment), compared with that of the TC3 strain grown in non‐amended medium (−Mn treatment). The abundance of mnp1–9 genes transcripts, relative to β‐tubulin, was measured by real‐time PCR, and quantified by the ΔΔCT method. TC3 – control strain (pTM1 transformant); TS1, TS9, TS24, TS30 – Mn2+‐dependent peroxidase silenced strains (pTMS1 transformants). Data represent the average of three biological replicates. Bars denote the standard deviation; when not visible, the standard deviation is included within the graph line width. Note the log10 scale, and different axis ranges.
Mentions: In order to determine whether alterations in MnP gene family transcript levels accompany the changes observed in OII decolourization we used relative real‐time PCR quantification analysis. The selected strains (TS1, TS9, TS24, TS30 and the TC3 control) were grown for 7 days in liquid culture, in the presence of carboxin, in either a medium amended with 27 µM Mn2+ (+Mn treatment) or a non‐amended medium (−Mn treatment). The results presented in Fig. 6 show the relative expression of the nine different members of the MnP gene family in the five selected strains. The overall abundance of the endogenous control gene (β‐tubulin) transcript was found to be similar in all the strains in all the tested treatments. As TC3 exhibited similar gene expression levels to the PC9 wild‐type strain, we used the former, in the −Mn treatment, as our calibrators for relative quantifications. The results obtained (Fig. 6) indicate that the predominantly expressed MnP gene family members expression in the pTMS1 transformants varied in intensity in response to Mn2+ amendment, similarly to the calibrator (TC3) and PC9 (Fig. 2), but at different levels. Specifically, relatively to the calibrator, mnp3 and mnp9 gene expression, though not substantially affected in the −Mn treatment (less than twofold difference), were strongly inhibited in the +Mn treatment which exhibited 24‐ to 79‐fold higher levels in the calibrator. Therefore, mnp3 and mnp9 were not even remotely upregulated at levels comparable to the control in response to Mn2+ amendment. mnp4 gene expression was not substantially affected in the −Mn treatment (less than 2.2‐fold difference); however, in the +Mn treatment it was 8‐ to 24‐fold higher in the calibrator; mnp8 gene expression was not substantially affected in the −Mn treatment (less than 2 fold difference), but in the +Mn treatment it was 4‐ to 9‐fold higher in the calibrator. Thus, in contrast to the striking results obtained in the case of mnp3, 4 and 9, the transcript abundance of mnp1, 2, 5, 6 and 7 was not substantially affected (less than fourfold difference), generally showing the same trend as the calibrator both in the −Mn and +Mn treatments.

Bottom Line: Relative real-time PCR quantification analysis confirmed that all the nine genes are transcribed, and that Mn(2+) amendment results in a drastic increase in the transcript levels of the predominantly expressed MnP genes (mnp 3 and mnp 9), while decreasing versatile peroxidase gene transcription (mnp 4).Knock-down of mnp 3 resulted in the reduction of fungal OII decolourization capacity, which was co-linear with marked silencing of the Mn(2+)-dependent peroxidase genes mnp 3 and mnp 9.This is the first direct genetic proof of an association between MnP gene expression levels and azo dye decolourization capacity in P. ostreatus, which may have significant implication on understanding the mechanisms governing lignin biodegradation.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Pathology and Microbiology, The Robert H. Smith Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem, Rehovot 76100, Israel.

Show MeSH
Related in: MedlinePlus