Limits...
Linoleic acid isomerase gene FgLAI12 affects sensitivity to salicylic acid, mycelial growth and virulence of Fusarium graminearum

View Article: PubMed Central - PubMed

ABSTRACT

Fusarium graminearum is the major causal agent of fusarium head blight in wheat, a serious disease worldwide. Linoleic acid isomerase (LAI) catalyses the transformation of linoleic acid (LA) to conjugated linoleic acid (CLA), which is beneficial for human health. We characterised a cis-12 LAI gene of F. graminearum (FGSG_02668; FgLAI12), which was downregulated by salicylic acid (SA), a plant defence hormone. Disruption of FgLAI12 in F. graminearum resulted in decreased accumulation of cis-9,trans-11 CLA, enhanced sensitivity to SA, and increased accumulation of LA and SA in wheat spikes during infection. In addition, mycelial growth, accumulation of deoxynivalenol, and pathogenicity in wheat spikes were reduced. Re-introduction of a functional FgLAI12 gene into ΔFgLAI12 recovered the wild-type phenotype. Fluorescent microscopic analysis showed that FgLAI12 protein was usually expressed in the septa zone of conidia and the vacuole of hyphae, but was expressed in the cell membrane of hyphae in response to exogenous LA, which may be an element of LA metabolism during infection by F. graminearum. The cis-12 LAI enzyme encoded by FgLAI12 is critical for fungal response to SA, mycelial growth and virulence in wheat. The gene FgLAI12 is potentially valuable for biotechnological synthesis of cis-9,trans-11 CLA.

No MeSH data available.


Related in: MedlinePlus

Deletion and complementation of FgLAI12 in F. graminearum.(A) The left border (LB) and right border (RB) were amplified from the wild type (a). ΔFgLAI12 mutant (d) was created by recombination between the knockout vector (b) and FgLAI12 (c). (B) The nucleotide sequence of FgLAI12 was cloned (a), and ligated into the complementation vector (b). The T-DNA region of the complementation vector was recombined into ΔFgLAI12 to create C-FgLAI12 (c). SacI, ApaI, SpeI, HindIII, NcoI and SpeI indicate the restriction enzymes used. (C) PCR verification of ΔFgLAI12 using the primer pairs P5 forward + P5 reverse and P7 forward + P7 reverse. (D) Verification of expression of FgLAI12 using the primer pair RJ-FgLAI12 forward + RJ-FgLAI12 reverse. FgGAPDH were used as a control. (E) Copy numbers of HPH determined by qPCR. Different letters (a and b) above each column indicate a significant difference (P < 0.05; n = 3). (F) Effect of SA and LA on expression of FgLAI12 in hyphae, as determined using the primer pair RJ-FgLAI12 (Table 1). Hyphae were collected on the 4th day after adding SA and LA. FgGAPDH was used as a reference. “F”, forward; “R”, reverse.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5384231&req=5

f2: Deletion and complementation of FgLAI12 in F. graminearum.(A) The left border (LB) and right border (RB) were amplified from the wild type (a). ΔFgLAI12 mutant (d) was created by recombination between the knockout vector (b) and FgLAI12 (c). (B) The nucleotide sequence of FgLAI12 was cloned (a), and ligated into the complementation vector (b). The T-DNA region of the complementation vector was recombined into ΔFgLAI12 to create C-FgLAI12 (c). SacI, ApaI, SpeI, HindIII, NcoI and SpeI indicate the restriction enzymes used. (C) PCR verification of ΔFgLAI12 using the primer pairs P5 forward + P5 reverse and P7 forward + P7 reverse. (D) Verification of expression of FgLAI12 using the primer pair RJ-FgLAI12 forward + RJ-FgLAI12 reverse. FgGAPDH were used as a control. (E) Copy numbers of HPH determined by qPCR. Different letters (a and b) above each column indicate a significant difference (P < 0.05; n = 3). (F) Effect of SA and LA on expression of FgLAI12 in hyphae, as determined using the primer pair RJ-FgLAI12 (Table 1). Hyphae were collected on the 4th day after adding SA and LA. FgGAPDH was used as a reference. “F”, forward; “R”, reverse.

Mentions: The FgLAI12 gene contains three exons and two introns (Fig. 2A). To determine the function of FgLAI12 in F. graminearum, knockout mutants (ΔFgLAI12) and complementation mutants (C-FgLAI12) were created (Fig. 2A,B). The mutants were verified by PCR (Fig. 2C,D) and sequencing (data not shown).


Linoleic acid isomerase gene FgLAI12 affects sensitivity to salicylic acid, mycelial growth and virulence of Fusarium graminearum
Deletion and complementation of FgLAI12 in F. graminearum.(A) The left border (LB) and right border (RB) were amplified from the wild type (a). ΔFgLAI12 mutant (d) was created by recombination between the knockout vector (b) and FgLAI12 (c). (B) The nucleotide sequence of FgLAI12 was cloned (a), and ligated into the complementation vector (b). The T-DNA region of the complementation vector was recombined into ΔFgLAI12 to create C-FgLAI12 (c). SacI, ApaI, SpeI, HindIII, NcoI and SpeI indicate the restriction enzymes used. (C) PCR verification of ΔFgLAI12 using the primer pairs P5 forward + P5 reverse and P7 forward + P7 reverse. (D) Verification of expression of FgLAI12 using the primer pair RJ-FgLAI12 forward + RJ-FgLAI12 reverse. FgGAPDH were used as a control. (E) Copy numbers of HPH determined by qPCR. Different letters (a and b) above each column indicate a significant difference (P < 0.05; n = 3). (F) Effect of SA and LA on expression of FgLAI12 in hyphae, as determined using the primer pair RJ-FgLAI12 (Table 1). Hyphae were collected on the 4th day after adding SA and LA. FgGAPDH was used as a reference. “F”, forward; “R”, reverse.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5384231&req=5

f2: Deletion and complementation of FgLAI12 in F. graminearum.(A) The left border (LB) and right border (RB) were amplified from the wild type (a). ΔFgLAI12 mutant (d) was created by recombination between the knockout vector (b) and FgLAI12 (c). (B) The nucleotide sequence of FgLAI12 was cloned (a), and ligated into the complementation vector (b). The T-DNA region of the complementation vector was recombined into ΔFgLAI12 to create C-FgLAI12 (c). SacI, ApaI, SpeI, HindIII, NcoI and SpeI indicate the restriction enzymes used. (C) PCR verification of ΔFgLAI12 using the primer pairs P5 forward + P5 reverse and P7 forward + P7 reverse. (D) Verification of expression of FgLAI12 using the primer pair RJ-FgLAI12 forward + RJ-FgLAI12 reverse. FgGAPDH were used as a control. (E) Copy numbers of HPH determined by qPCR. Different letters (a and b) above each column indicate a significant difference (P < 0.05; n = 3). (F) Effect of SA and LA on expression of FgLAI12 in hyphae, as determined using the primer pair RJ-FgLAI12 (Table 1). Hyphae were collected on the 4th day after adding SA and LA. FgGAPDH was used as a reference. “F”, forward; “R”, reverse.
Mentions: The FgLAI12 gene contains three exons and two introns (Fig. 2A). To determine the function of FgLAI12 in F. graminearum, knockout mutants (ΔFgLAI12) and complementation mutants (C-FgLAI12) were created (Fig. 2A,B). The mutants were verified by PCR (Fig. 2C,D) and sequencing (data not shown).

View Article: PubMed Central - PubMed

ABSTRACT

Fusarium graminearum is the major causal agent of fusarium head blight in wheat, a serious disease worldwide. Linoleic acid isomerase (LAI) catalyses the transformation of linoleic acid (LA) to conjugated linoleic acid (CLA), which is beneficial for human health. We characterised a cis-12 LAI gene of F. graminearum (FGSG_02668; FgLAI12), which was downregulated by salicylic acid (SA), a plant defence hormone. Disruption of FgLAI12 in F. graminearum resulted in decreased accumulation of cis-9,trans-11 CLA, enhanced sensitivity to SA, and increased accumulation of LA and SA in wheat spikes during infection. In addition, mycelial growth, accumulation of deoxynivalenol, and pathogenicity in wheat spikes were reduced. Re-introduction of a functional FgLAI12 gene into &Delta;FgLAI12 recovered the wild-type phenotype. Fluorescent microscopic analysis showed that FgLAI12 protein was usually expressed in the septa zone of conidia and the vacuole of hyphae, but was expressed in the cell membrane of hyphae in response to exogenous LA, which may be an element of LA metabolism during infection by F. graminearum. The cis-12 LAI enzyme encoded by FgLAI12 is critical for fungal response to SA, mycelial growth and virulence in wheat. The gene FgLAI12 is potentially valuable for biotechnological synthesis of cis-9,trans-11 CLA.

No MeSH data available.


Related in: MedlinePlus