Modularity of plant metabolic gene clusters: a trio of linked genes that are collectively required for acylation of triterpenes in oat

Operon-like gene clusters are an emerging phenomenon in the field of plant natural products. The genes encoding some of the best-characterized plant secondary metabolite biosynthetic pathways are scattered across plant genomes. However, an increasing number of gene clusters encoding the synthesis of diverse natural products have recently been reported in plant genomes. These clusters have arisen through the neo-functionalization and relocation of existing genes within the genome, and not by horizontal gene transfer from microbes. The reasons for clustering are not yet clear, although this form of gene organization is likely to facilitate co-inheritance and co-regulation. Oats (Avena spp) synthesize antimicrobial triterpenoids (avenacins) that provide protection against disease. The synthesis of these compounds is encoded by a gene cluster. Here we show that a module of three adjacent genes within the wider biosynthetic gene cluster is required for avenacin acylation. Through the characterization of these genes and their encoded proteins we present a model of the subcellular organization of triterpenoid biosynthesis

The complete nucleotide sequence of tobacco necrosis virus strain D

The complete sequence of the RNA genome of tobacco necrosis virus strain D (TNV-D) consisting of 3759 nucleotides has been determined. The positive strand contains five open reading frames (ORFs). The 5'proximal ORF encodes a 22K protein terminating with an amber codon which may be read through to produce a 82K protein (p82). Two small centrally located ORFs each encode two out-of-frame 7K proteins (p7a and p7b). The 3'-proximal ORF encodes the 29K coat protein (CP), the N terminus of which has been sequenced directly. The genomic organization of TNVD is very similar to that of TNV-A but differs in the placement of the p7a ORF, which does not overlap the p82 ORF in TNV-D, and in the absence of an ORF downstream of the CP gene in TNV-D. The p82 ORF shows extensive sequence similarity with the putative polymerases of the carmovirus group. This ORF is also as closely related to the corresponding ORF of TNV-A as it is to the corresponding ORF of the tombusvirus cucumber necrosis virus. The amino acid sequence of the TNV-D CP gene is similar to both the TNV-A and southern bean mosaic virus CP genes. Of the two p7 ORFs, p7a exhibits amino acid sequence similarity with corresponding proteins from TNV-A, melon necrotic spot virus, carnation mottle virus, turnip crinkle virus and maize chlorotic mottle virus, whereas the p7b ORF appears to be unique to TNV-A and TNV-D. Only the 3'-terminal three nucleotides of TNV-D genomic RNA are identical to the 3'-terminal nucleotides of the TNV satellite virus.Peer reviewe

Self-assembly of filamentous virus-like particles (VLPs) through transient expression in N. benthamiana of the coat protein (CP) of different sweet potato viruses

Trabajo presentado al International Advances In Plant Virology, celebrado on-line del 20 al 22 de abril de 2021.Peer reviewe

Engineering Cowpea Mosaic Virus RNA-2 into a vector to express heterologous proteins in plants

series of new cowpea mosaic virus (CPMV) RNA-2-based expression vectors were designed. The jellyfish green fluorescent protein (GFP) was introduced between the movement protein (MP) and the large (L) coat protein or downstream of the small (S) coat protein. Release of the GFP inserted between the MP and L proteins was achieved by creating artificial processing sites each side of the insert, either by duplicating the MP-L cleavage site or by introducing a sequence encoding the foot-and-mouth disease virus (FMDV) 2A catalytic peptide. Eight amino acids derived from the C-terminus of the MP and 14–19 amino acids from the N-terminus of the L coat protein were necessary for efficient processing of the artificial Gln/Met sites. Insertion of the FMDV 2A sequence at the C-terminus of the GFP resulted in a genetically stable construct, which produced particles containing about 10 GFP-2A-L fusion proteins. Immunocapture experiments indicated that some of the GFP is present on the virion surface. Direct fusion of GFP to the C-terminus of the S coat protein resulted in a virus which was barely viable. However, when the sequence of GFP was linked to the C-terminus by an active FMDV 2A sequence, a highly infectious construct was obtained