-expressed SECRET AGENT -GlcNAc modifies threonine 829 of GIGANTEA.

The glycosyl transferases SPINDLY (SPY) and SECRET AGENT (SEC) modify nuclear and cytosolic proteins with -linked fucose or -linked -acetylglucosamine (-GlcNAc), respectively. -fucose and -GlcNAc modifications can occur at the same sites. SPY interacts physically and genetically with GIGANTEA (GI), suggesting that it could be modified by both enzymes.

A deoxyviolacein-based transposon insertion vector for pigmented tracer studies.

Pigments provide a simple means to rapidly visually ascertain the quantities or presence of specific microbes in a complex community. The selection of pigment-producing colonies that are simple to differentiate from common colony phenotypes provides a high degree of certainty for the identity of pigment-tagged strains. Successful employment of pigment production is dependent on various intrinsic factors related to proper levels of gene expression and pigment production that are not always easy to predict and vary within each microbe.

Enhanced extracellular ammonium release in the plant endophyte through genome editing.

Our results demonstrate increased extracellular ammonium release in the endophyte plant growth-promoting bacterium . Strains were constructed in a manner that leaves no antibiotic markers behind, such that these strains contain no transgenes. Levels of ammonium achieved by cultures of modified strains reached concentrations of approximately 18 mM ammonium, while wild-type remained much lower (below 50 µM).

Gluconacetobacter diazotrophicus Gene Fitness during Diazotrophic Growth.

Plant growth-promoting (PGP) bacteria are important to the development of sustainable agricultural systems. PGP microbes that fix atmospheric nitrogen (diazotrophs) could minimize the application of industrially derived fertilizers and function as a biofertilizer. The bacterium Gluconacetobacter diazotrophicus is a nitrogen-fixing PGP microbe originally discovered in association with sugarcane plants, where it functions as an endophyte.

Genomic characterization of cycad leaf necrosis virus, the first badnavirus identified in a gymnosperm.

A previously undescribed badnavirus was isolated from Zamia fischeri showing symptoms of chlorosis, necrosis, and ringspot. The virus has bacilliform virions 30 nm in diameter and averaging 120 nm in length. The viral genome is 9227 bp in length and contains three open reading frames characteristic of members of the genus Badnavirus.

Complete genome sequence of aglaonema bacilliform virus (ABV).

Aglaonema bacilliform virus (ABV), a member of the genus Badnavirus in the family Caulimoviridae, is associated with leaf deformation and chlorosis in Aglaonema modestum. The complete genome sequence of a Minnesota isolate of ABV was determined. The ABV genome is 7,178 bp in length and similar in size and organization to those of the members of the genus Badnavirus, containing three open reading frames (ORFs) with the potential to encode three proteins of 14.

Using CRISPR/Cas9 genome editing in tomato to create a gibberellin-responsive dominant dwarf DELLA allele.

The tomato PROCERA gene encodes a DELLA protein, and loss-of-function mutations derepress growth. We used CRISPR/Cas9 and a single guide RNAs (sgRNA) to target mutations to the PROCERA DELLA domain, and recovered several loss-of-function mutations and a dominant dwarf mutation that carries a deletion of one amino acid in the DELLA domain. This is the first report of a dominant dwarf PROCERA allele.

The Arabidopsis O-fucosyltransferase SPINDLY activates nuclear growth repressor DELLA.

Plant development requires coordination among complex signaling networks to enhance the plant's adaptation to changing environments. DELLAs, transcription regulators originally identified as repressors of phytohormone gibberellin signaling, play a central role in integrating multiple signaling activities via direct protein interactions with key transcription factors. Here, we found that DELLA is mono-O-fucosylated by the novel O-fucosyltransferase SPINDLY (SPY) in Arabidopsis thaliana.

Constitutive Expression of Arabidopsis SMALL AUXIN UP RNA19 (SAUR19) in Tomato Confers Auxin-Independent Hypocotyl Elongation.

The plant hormone indole-3-acetic acid (IAA or auxin) mediates the elongation growth of shoot tissues by promoting cell expansion. According to the acid growth theory proposed in the 1970s, auxin activates plasma membrane H-ATPases (PM H-ATPases) to facilitate cell expansion by both loosening the cell wall through acidification and promoting solute uptake. Mechanistically, however, this process is poorly understood.

O-GlcNAcylation of master growth repressor DELLA by SECRET AGENT modulates multiple signaling pathways in Arabidopsis.

The DELLA family of transcription regulators functions as master growth repressors in plants by inhibiting phytohormone gibberellin (GA) signaling in response to developmental and environmental cues. DELLAs also play a central role in mediating cross-talk between GA and other signaling pathways via antagonistic direct interactions with key transcription factors. However, how these crucial protein-protein interactions can be dynamically regulated during plant development remains unclear.

GA signalling and cross-talk with other signalling pathways.

Gibberellins (GAs) are phytohormones that regulate growth and development. DELLA proteins repress GA responses. GA binding to its receptor triggers a series of events that culminate in the destruction of DELLA proteins by the 26S proteasome, which removes the repression of GA signalling.

Uncovering DELLA-Independent Gibberellin Responses by Characterizing New Tomato procera Mutants.

Gibberellin (GA) regulates plant development primarily by triggering the degradation/deactivation of the DELLA proteins. However, it remains unclear whether all GA responses are regulated by DELLAs. Tomato (Solanum lycopersicum) has a single DELLA gene named PROCERA (PRO), and its recessive pro allele exhibits constitutive GA activity but retains responsiveness to external GA.

The putative eukaryote-like O-GlcNAc transferase of the cyanobacterium Synechococcus elongatus PCC 7942 hydrolyzes UDP-GlcNAc and is involved in multiple cellular processes.

The posttranslational addition of a single O-linked β-N-acetylglucosamine (O-GlcNAc) to serine or threonine residues regulates numerous metazoan cellular processes. The enzyme responsible for this modification, O-GlcNAc transferase (OGT), is conserved among a wide variety of organisms and is critical for the viability of many eukaryotes. Although OGTs with domain structures similar to those of eukaryotic OGTs are predicted for many bacterial species, the cellular roles of these OGTs are unknown.

Targeted mutagenesis of the tomato PROCERA gene using transcription activator-like effector nucleases.

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O-GlcNAc modification of the coat protein of the potyvirus Plum pox virus enhances viral infection.

O-GlcNAcylation is a dynamic protein modification which has been studied mainly in metazoans. We reported previously that an Arabidopsis thaliana O-GlcNAc transferase modifies at least two threonine residues of the Plum pox virus (PPV) capsid protein (CP). Now, six additional residues were shown to be involved in O-GlcNAc modification of PPV CP.

Identification and origin of N-linked β-D-N-acetylglucosamine monosaccharide modifications on Arabidopsis proteins.

Many plant proteins are modified with N-linked oligosaccharides at asparagine-X-serine/threonine sites during transit through the endoplasmic reticulum and the Golgi. We have identified a number of Arabidopsis (Arabidopsis thaliana) proteins with modifications consisting of an N-linked N-acetyl-D-glucosamine monosaccharide (N-GlcNAc). Electron transfer dissociation mass spectrometry analysis of peptides bearing this modification mapped the modification to asparagine-X-serine/threonine sites on proteins that are predicted to transit through the endoplasmic reticulum and Golgi.

O-GlcNAcylation of the Plum pox virus capsid protein catalyzed by SECRET AGENT: characterization of O-GlcNAc sites by electron transfer dissociation mass spectrometry.

The capsid protein of Plum pox virus (PPV-CP) is modified with O-linked β-N-acetylglucosamine (O-GlcNAc). In Arabidopsis thaliana this modification is made by an O-GlcNAc transferase named SECRET AGENT (SEC). Modification of PPV-CP by SEC is hypothesized to have a direct role in the infection process, because virus titer and rate of spread are reduced in SEC mutants.

O-GlcNAc protein modification in plants: Evolution and function.

The role in plants of posttranslational modification of proteins with O-linked N-acetylglucosamine and the evolution and function of O-GlcNAc transferases responsible for this modification are reviewed. Phylogenetic analysis of eukaryotic O-GlcNAc transferases (OGTs) leads us to propose that plants have two distinct OGTs, SEC- and SPY-like, that originated in prokaryotes. Animals and some fungi have a SEC-like enzyme while plants have both.

Characterization of a Strain of Turnip vein-clearing virus Causing Red Ringspot of Penstemon.

A disease of penstemon (Penstemon digitalis) occurring in commercial nurseries in Minnesota in 2004 to 2006 and characterized by red foliar ringspots, leaf deformation, and plant stunting was found to be caused by a strain of Turnip vein-clearing virus (TVCV) that was named Penstemon ringspot virus (PenRSV). This is the first report of a viral disease of penstemon. The genome organization of PenRSV was similar to that of the crucifer-infecting tobamoviruses.

Biodegradation of atrazine in transgenic plants expressing a modified bacterial atrazine chlorohydrolase (atzA) gene.

Atrazine is one of the most widely used herbicides in the USA. Atrazine chlorohydrolase (AtzA), the first enzyme in a six-step pathway leading to the mineralization of atrazine in Gram-negative soil bacteria, catalyses the hydrolytic dechlorination and detoxification of atrazine to hydroxyatrazine. In this study, we investigated the potential use of transgenic plants expressing atzA to take up, dechlorinate and detoxify atrazine.

Functional analysis of SPINDLY in gibberellin signaling in Arabidopsis.

The Arabidopsis (Arabidopsis thaliana) SPINDLY (SPY) protein negatively regulates the gibberellin (GA) signaling pathway. SPY is an O-linked N-acetylglucosamine (GlcNAc) transferase (OGT) with a protein-protein interaction domain consisting of 10 tetratricopeptide repeats (TPR). OGTs add a GlcNAc monosaccharide to serine/threonine residues of nuclear and cytosolic proteins.

SECRET AGENT, an Arabidopsis thaliana O-GlcNAc transferase, modifies the Plum pox virus capsid protein.

The capsid protein of Plum pox virus (PPV-CP) is modified with O-linked GlcNAc (O-GlcNAc). While Arabidopsis has two O-GlcNAc transferases, SECRET AGENT (SEC) and SPINDLY (SPY), previous work suggests that SEC modifies PPV-CP and that the modification plays a role in the infection process. Here, we show that when co-expressed in Escherichia coli SEC modifies PPV-CP.

Mapping of two O-GlcNAc modification sites in the capsid protein of the potyvirus Plum pox virus.

A large number of O-linked N-acetylglucosamine (O-GlcNAc) residues have been mapped in vertebrate proteins, however targets of O-GlcNAcylation in plants still have not been characterized. We show here that O-GlcNAcylation of the N-terminal region of the capsid protein of Plum pox virus resembles that of animal proteins in introducing O-GlcNAc monomers. Thr-19 and Thr-24 were specifically O-GlcNAcylated.