A chalcone synthase with an unusual substrate preference is expressed in barley leaves in response to UV light and pathogen attack.
A cDNA clone was isolated by differential hybridization from a library prepared from barley leaves inoculated with the fungus Blumeria graminis f.sp. hordei (Bgh). The open reading frame of the insert (designated HvCHS2) encoded a polypeptide with 72-79% identity to chalcone synthases (CHS) and 65-68% identity to stilbene synthases. Alignments of the amino acid sequence of HvCHS2 with the consensus sequence of naringenin-CHS (EC 2.3.1.74) reveals significant differences between HvCHS2 and naringenin-CHS. HvCHS2 transcripts accumulate strongly in barley leaves in response to inoculation with Bgh, whereas only insignificant accumulation of barley naringenin-CHS (CHS1) transcripts is seen upon the inoculation. The accumulation of HvCHS2 transcripts is also elicited by UV light. To compare the activity of HvCHS2 with the activity of CHS1, the two enzymes were expressed in Escherichia coli. Both HvCHS2 and CHS1 catalyse the formation of chalcones. However, HvCHS2 and CHS1 differ in their substrate requirements. CHS1 uses cinnamoyl-CoA and 4-coumaroyl-CoA at comparable rates whereas feruloyl-CoA is a poor substrate for this enzyme. In contrast, HvCHS2 converts feruloyl-CoA and caffeoyl-CoA at the highest rate whereas cinnamoyl-CoA is a poor substrate. Thus, HvCHS2 is a novel pathogen and UV light induces homoeriodictyol/eriodictyol CHS involved in the direct production of flavonoids possessing multi-substituted B-rings.
Christensen
A B
AB
Department of Plant Biology, Royal Veterinary and Agricultural University, Frederiksberg, Denmark.
Gregersen
P L
PL
Schröder
J
J
Collinge
D B
DB
D000214:Acyl Coenzyme A / Q000737:chemistry; D000217:Acyltransferases / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D001203:Ascomycota / Q000472:pathogenicity / Q000502:physiology*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D044950:Flavanones; D005419:Flavonoids / Q000737:chemistry / Q000235:genetics; D018506:Gene Expression Regulation, Plant / Q000502:physiology / Q000528:radiation effects; D001467:Hordeum / Q000201:enzymology* / Q000235:genetics / Q000382:microbiology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018515:Plant Leaves / Q000201:enzymology; D012333:RNA, Messenger / Q000032:analysis; D018749:RNA, Plant / Q000032:analysis; D011993:Recombinant Fusion Proteins; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D014466:Ultraviolet Rays
D000214:Acyl Coenzyme A; D044950:Flavanones; D005419:Flavonoids; D012333:RNA, Messenger; D018749:RNA, Plant; D011993:Recombinant Fusion Proteins; D000217:Acyltransferases; C020141:flavanone synthetase; C005273:naringenin