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Botrytis cinerea protein O-mannosyltransferases play critical roles in morphogenesis, growth, and virulence.

González M, Brito N, Frías M, González C - PLoS ONE (2013)

Bottom Line: The most significant case is that of grapevine leaves, whose penetration requires the three functional PMTs.Furthermore, PMT2 also contributes significantly to fungal adherence on grapevine and tobacco leaves.Analysis of extracellular and membrane proteins showed significant changes in the pattern of protein secretion and glycosylation by the pmt mutants, and allowed the identification of new protein substrates putatively glycosylated by specific PMTs.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Bioquímica y Biología Molecular, Universidad de La Laguna, La Laguna (Tenerife), Spain.

ABSTRACT
Protein O-glycosylation is crucial in determining the structure and function of numerous secreted and membrane-bound proteins. In fungi, this process begins with the addition of a mannose residue by protein O-mannosyltransferases (PMTs) in the lumen side of the ER membrane. We have generated mutants of the three Botrytis cinerea pmt genes to study their role in the virulence of this wide-range plant pathogen. B. cinerea PMTs, especially PMT2, are critical for the stability of the cell wall and are necessary for sporulation and for the generation of the extracellular matrix. PMTs are also individually required for full virulence in a variety of hosts, with a special role in the penetration of intact plant leaves. The most significant case is that of grapevine leaves, whose penetration requires the three functional PMTs. Furthermore, PMT2 also contributes significantly to fungal adherence on grapevine and tobacco leaves. Analysis of extracellular and membrane proteins showed significant changes in the pattern of protein secretion and glycosylation by the pmt mutants, and allowed the identification of new protein substrates putatively glycosylated by specific PMTs. Since plants do no possess these enzymes, PMTs constitute a promising target in the development of novel control strategies against B. cinerea.

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Related in: MedlinePlus

Conidia and sclerotia production by Δbcpmt mutants.The indicated strains were grown under continuous light (plain bars) or in darkness (hatched bars) in tomato-agar plates. Osmotic agents were added when indicated. A) Appearance of the colonies 12 days after inoculation. B) Conidia production at day 15. C) Sclerotia production at day 15. Bars represent mean values (n = 3) and standard deviation, and asterisks indicate a statistically significant difference with the wild type in the same medium (p<0.05).
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pone-0065924-g004: Conidia and sclerotia production by Δbcpmt mutants.The indicated strains were grown under continuous light (plain bars) or in darkness (hatched bars) in tomato-agar plates. Osmotic agents were added when indicated. A) Appearance of the colonies 12 days after inoculation. B) Conidia production at day 15. C) Sclerotia production at day 15. Bars represent mean values (n = 3) and standard deviation, and asterisks indicate a statistically significant difference with the wild type in the same medium (p<0.05).

Mentions: The amount of conidia and sclerotia produced was determined quantitatively for the Δbcpmt mutants grown for 15 days on tomato fruit extract, either under continuous visible light, to maximize the production of conidia, or in the dark, to maximize the production of sclerotia [35]. Δbcpmt2 was completely unable to sporulate, even when osmotic stabilizers were added to the growth medium, and no clear sclerotia formation was observed (Figure 4). However, Δbcpmt2 colonies were strongly melanized in the dark, which is one of the features of sclerotia (Figure 4A). Δbcpmt4 and especially Δbcpmt1 were able to sporulate, but the number of conidia produced was reduced (Figure 4B). This difference was larger in the dark, so that light was a more potent inducer of sporulation for the pmt mutants than for the wild type. Addition of sorbitol improved sporulation only slightly in the case of Δbcpmt4 (Figure 4B). Conidia obtained from these two mutants did not show any difference in their germination rate as compared with the wild type (not shown), about 83% of conidia obtained were able to germinate in all cases. Sclerotia production, on the contrary, was higher for Δbcpmt1 and Δbcpmt4, under continuous illumination, than for the wild type (Figure 4C). Light is a strong repressor of sclerotia formation for B. cinerea[35] but it seems that it does not produce the same effect in the pmt mutants. Addition of the osmotic stabilizers KCl and sorbitol completely abolished sclerotia formation for the three mutants and for the wild type (not shown). The sclerotia produced were viable in 100% of the cases.


Botrytis cinerea protein O-mannosyltransferases play critical roles in morphogenesis, growth, and virulence.

González M, Brito N, Frías M, González C - PLoS ONE (2013)

Conidia and sclerotia production by Δbcpmt mutants.The indicated strains were grown under continuous light (plain bars) or in darkness (hatched bars) in tomato-agar plates. Osmotic agents were added when indicated. A) Appearance of the colonies 12 days after inoculation. B) Conidia production at day 15. C) Sclerotia production at day 15. Bars represent mean values (n = 3) and standard deviation, and asterisks indicate a statistically significant difference with the wild type in the same medium (p<0.05).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3675079&req=5

pone-0065924-g004: Conidia and sclerotia production by Δbcpmt mutants.The indicated strains were grown under continuous light (plain bars) or in darkness (hatched bars) in tomato-agar plates. Osmotic agents were added when indicated. A) Appearance of the colonies 12 days after inoculation. B) Conidia production at day 15. C) Sclerotia production at day 15. Bars represent mean values (n = 3) and standard deviation, and asterisks indicate a statistically significant difference with the wild type in the same medium (p<0.05).
Mentions: The amount of conidia and sclerotia produced was determined quantitatively for the Δbcpmt mutants grown for 15 days on tomato fruit extract, either under continuous visible light, to maximize the production of conidia, or in the dark, to maximize the production of sclerotia [35]. Δbcpmt2 was completely unable to sporulate, even when osmotic stabilizers were added to the growth medium, and no clear sclerotia formation was observed (Figure 4). However, Δbcpmt2 colonies were strongly melanized in the dark, which is one of the features of sclerotia (Figure 4A). Δbcpmt4 and especially Δbcpmt1 were able to sporulate, but the number of conidia produced was reduced (Figure 4B). This difference was larger in the dark, so that light was a more potent inducer of sporulation for the pmt mutants than for the wild type. Addition of sorbitol improved sporulation only slightly in the case of Δbcpmt4 (Figure 4B). Conidia obtained from these two mutants did not show any difference in their germination rate as compared with the wild type (not shown), about 83% of conidia obtained were able to germinate in all cases. Sclerotia production, on the contrary, was higher for Δbcpmt1 and Δbcpmt4, under continuous illumination, than for the wild type (Figure 4C). Light is a strong repressor of sclerotia formation for B. cinerea[35] but it seems that it does not produce the same effect in the pmt mutants. Addition of the osmotic stabilizers KCl and sorbitol completely abolished sclerotia formation for the three mutants and for the wild type (not shown). The sclerotia produced were viable in 100% of the cases.

Bottom Line: The most significant case is that of grapevine leaves, whose penetration requires the three functional PMTs.Furthermore, PMT2 also contributes significantly to fungal adherence on grapevine and tobacco leaves.Analysis of extracellular and membrane proteins showed significant changes in the pattern of protein secretion and glycosylation by the pmt mutants, and allowed the identification of new protein substrates putatively glycosylated by specific PMTs.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Bioquímica y Biología Molecular, Universidad de La Laguna, La Laguna (Tenerife), Spain.

ABSTRACT
Protein O-glycosylation is crucial in determining the structure and function of numerous secreted and membrane-bound proteins. In fungi, this process begins with the addition of a mannose residue by protein O-mannosyltransferases (PMTs) in the lumen side of the ER membrane. We have generated mutants of the three Botrytis cinerea pmt genes to study their role in the virulence of this wide-range plant pathogen. B. cinerea PMTs, especially PMT2, are critical for the stability of the cell wall and are necessary for sporulation and for the generation of the extracellular matrix. PMTs are also individually required for full virulence in a variety of hosts, with a special role in the penetration of intact plant leaves. The most significant case is that of grapevine leaves, whose penetration requires the three functional PMTs. Furthermore, PMT2 also contributes significantly to fungal adherence on grapevine and tobacco leaves. Analysis of extracellular and membrane proteins showed significant changes in the pattern of protein secretion and glycosylation by the pmt mutants, and allowed the identification of new protein substrates putatively glycosylated by specific PMTs. Since plants do no possess these enzymes, PMTs constitute a promising target in the development of novel control strategies against B. cinerea.

Show MeSH
Related in: MedlinePlus