PMID- 25531527
OWN - NLM
STAT- MEDLINE
DCOM- 20151215
LR  - 20201217
IS  - 1932-6203 (Electronic)
IS  - 1932-6203 (Linking)
VI  - 9
IP  - 12
DP  - 2014
TI  - Enzymes from fungal and plant origin required for chemical diversification of 
      insecticidal loline alkaloids in grass-Epichloe symbiota.
PG  - e115590
LID - 10.1371/journal.pone.0115590 [doi]
LID - e115590
AB  - The lolines are a class of bioprotective alkaloids that are produced by Epichloe 
      species, fungal endophytes of grasses. These alkaloids are saturated 
      1-aminopyrrolizidines with a C2 to C7 ether bridge, and are structurally 
      differentiated by the various modifications of the 1-amino group: -NH2 
      (norloline), -NHCH3 (loline), -N(CH3)2 (N-methylloline), -N(CH3)Ac 
      (N-acetylloline), -NHAc (N-acetylnorloline), and -N(CH3)CHO (N-formylloline). 
      Other than the LolP cytochrome P450, which is required for conversion of 
      N-methylloline to N-formylloline, the enzymatic steps for loline diversification 
      have not yet been established. Through isotopic labeling, we determined that 
      N-acetylnorloline is the first fully cyclized loline alkaloid, implying that 
      deacetylation, methylation, and acetylation steps are all involved in loline 
      alkaloid diversification. Two genes of the loline alkaloid biosynthesis (LOL) 
      gene cluster, lolN and lolM, were predicted to encode an N-acetamidase 
      (deacetylase) and a methyltransferase, respectively. A knockout strain lacking 
      both lolN and lolM stopped the biosynthesis at N-acetylnorloline, and 
      complementation with the two wild-type genes restored production of 
      N-formylloline and N-acetylloline. These results indicated that lolN and lolM are 
      required in the steps from N-acetylnorloline to other lolines. The function of 
      LolM as an N-methyltransferase was confirmed by its heterologous expression in 
      yeast resulting in conversion of norloline to loline, and of loline to 
      N-methylloline. One of the more abundant lolines, N-acetylloline, was observed in 
      some but not all plants with symbiotic Epichloe siegelii, and when provided with 
      exogenous loline, asymbiotic meadow fescue (Lolium pratense) plants produced 
      N-acetylloline, suggesting that a plant acetyltransferase catalyzes 
      N-acetylloline formation. We conclude that although most loline alkaloid 
      biosynthesis reactions are catalyzed by fungal enzymes, both fungal and plant 
      enzymes are responsible for the chemical diversification steps in symbio.
FAU - Pan, Juan
AU  - Pan J
AD  - Department of Plant Pathology, University of Kentucky, Lexington, Kentucky, 
      United States of America.
FAU - Bhardwaj, Minakshi
AU  - Bhardwaj M
AD  - Department of Chemistry, University of Kentucky, Lexington, Kentucky, United 
      States of America.
FAU - Nagabhyru, Padmaja
AU  - Nagabhyru P
AD  - Department of Plant Pathology, University of Kentucky, Lexington, Kentucky, 
      United States of America.
FAU - Grossman, Robert B
AU  - Grossman RB
AD  - Department of Chemistry, University of Kentucky, Lexington, Kentucky, United 
      States of America.
FAU - Schardl, Christopher L
AU  - Schardl CL
AD  - Department of Chemistry, University of Kentucky, Lexington, Kentucky, United 
      States of America.
LA  - eng
GR  - R01 GM086888/GM/NIGMS NIH HHS/United States
GR  - R01GM086888/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
PT  - Research Support, U.S. Gov't, P.H.S.
DEP - 20141222
PL  - United States
TA  - PLoS One
JT  - PloS one
JID - 101285081
RN  - 0 (Alkaloids)
RN  - 0 (Fungal Proteins)
RN  - 9035-51-2 (Cytochrome P-450 Enzyme System)
RN  - IC997T2WZ9 (loline)
SB  - IM
MH  - Alkaloids/*chemistry/classification/*metabolism
MH  - Amino Acid Sequence
MH  - Blotting, Southern
MH  - Blotting, Western
MH  - Cytochrome P-450 Enzyme System/genetics/*metabolism
MH  - Epichloe/genetics/*isolation & purification/physiology
MH  - Fungal Proteins/genetics/*metabolism
MH  - Gas Chromatography-Mass Spectrometry
MH  - Genome, Fungal
MH  - Molecular Sequence Data
MH  - Nuclear Magnetic Resonance, Biomolecular
MH  - Phylogeny
MH  - Poaceae/*microbiology
MH  - Sequence Homology, Amino Acid
MH  - *Symbiosis
MH  - Yeasts
PMC - PMC4274035
COIS- Competing Interests: The authors have declared that no competing interests exist.
EDAT- 2014/12/23 06:00
MHDA- 2015/12/17 06:00
PMCR- 2014/12/22
CRDT- 2014/12/23 06:00
PHST- 2014/09/14 00:00 [received]
PHST- 2014/11/29 00:00 [accepted]
PHST- 2014/12/23 06:00 [entrez]
PHST- 2014/12/23 06:00 [pubmed]
PHST- 2015/12/17 06:00 [medline]
PHST- 2014/12/22 00:00 [pmc-release]
AID - PONE-D-14-41406 [pii]
AID - 10.1371/journal.pone.0115590 [doi]
PST - epublish
SO  - PLoS One. 2014 Dec 22;9(12):e115590. doi: 10.1371/journal.pone.0115590. 
      eCollection 2014.