Pradeep Kumar Prabhakar

Pectins are a major class of plant cell wall polysaccharide crucial for plant survival and adaptation. Despite their importance, the biosynthesis of pectin remains poorly understood due to its exceptional structural and functional diversity. Functional interrogation of pectin biosynthetic enzymes in planta is often limited by severe developmental or difficult to interpret carbohydrate phenotypes resulting from genetic disruption. Using a direct biochemical approach, we report the identification and functional characterization of a Galactan Initiation (GalTi) enzyme from the aquatic monocot Spirodela polyrhiza, a member of CAZy glycosyltransferase family 47, that initiates galactan side‑chain biosynthesis on rhamnogalacturonan‑I. We further show that orthologs from Arabidopsis thaliana exhibit identical catalytic activity, revealing conservation of this biosynthetic initiation mechanism across monocots and dicots. Comprehensive carbohydrate structural analyses demonstrate that GalTi enzymes specifically catalyze the transfer of single β‑D‑galactopyranosyl residues to the O‑4 position of backbone rhamnose residues, establishing their direct role in galactan side‑chain biosynthesis. Co-synthesis experiments show 3-O-acetylation of backbone rhamnose residues inhibits SpGalTi‑mediated galactosylation, suggesting a potential regulatory mechanism by which plants use to modulate side‑chain patterning in vivo. Together, these findings provide a new framework for RG‑I galactan assembly, advancing our mechanistic understanding of plant cell wall biosynthesis.

SUMMARY: Caterpillars and beetles are among the most economically damaging defoliating insects, and their economic damage is predicted to increase in the coming decades. Hence the use of genetically derived resistance to supplement other pest control strategies is warranted. In soybean ( Glycine max (L.) Merr.), a major determinant for resistance is the quantitative trait locus, QTL‐M. Glyma07g14530 , the gene underlying QTL‐M, encodes a feeding‐inducible flavonol 3‐ O ‐glycosyltransferase (F3GlcT or UGT78D2) that glucosylates kaempferol, as well as quercetin, myricetin, and isorhamnetin. The resistant allele has a premature stop codon in it, thus preventing the glucosylation and sequestration of flavonols in the vacuole, leading to a concomitant accumulation of proanthocyanidins and manifestation of resistance. Expressing the dominant (susceptible) allele in resistant plants restores susceptibility, and silencing the susceptible allele results in resistance. The discovery and characterization of GmF3GlcT helps clarify the role of flavonoids in resistance to leaf‐chewing insects and facilitates the development of insect‐resistant cultivars that ultimately can lower production costs and reduce insecticide applications.

SUMMARY: Glycosyltransferases (GTs) are the primary enzymes responsible for the biosynthesis of the complex polysaccharides in plant cell walls. Given the important role of GTs in plants, it is necessary to undertake their functional characterization to better understand plant cell wall synthesis pathways to develop improved feedstocks for efficient conversion into fuels and products to support the emerging bioeconomy. The GT47 family in plants represents a unique target for characterization due to the substantial diversity of donor and acceptor substrates observed within a single family. Here, we have carried out the biochemical characterization of MUR3 and XLT2 orthologs from the aquatic monocot Spirodela polyrhiza . Our findings support existing genetic and phylogenetic data classifying these enzymes as regio‐specific galactosyltransferases involved in xyloglucan (XyG) sidechain biosynthesis. In addition, we have identified novel characteristics for both enzymes, such as in vitro arabinopyranosyltransferase activity and distinctiveness in xyloglucan reducing end specificity.

Glycosyltransferases (GTs) are the primary enzymes responsible for the biosynthesis of the complex polysaccharides in plant cell walls. Given the important role of GTs in plants, it is necessary to undertake their functional characterization to better understand plant cell wall synthesis pathways to develop improved feedstocks for efficient conversion into fuels and products to support the emerging bioeconomy. The GT47 family in plants represents a unique target for characterization due to the substantial diversity of donor and acceptor substrates observed within a single family. Here, we have carried out the biochemical characterization of MUR3 and XLT2 orthologs from the aquatic monocot Spirodela polyrhiza. Our findings support existing genetic and phylogenetic data classifying these enzymes as regio-specific galactosyltransferases involved in xyloglucan (XyG) sidechain biosynthesis. In addition, we have identified novel characteristics for both enzymes, such as in vitro arabinopyranosyltransferase activity and distinctiveness in xyloglucan reducing end specificity.

Summary: Caterpillars and beetles are among the most economically damaging defoliating insects, and their economic damage is predicted to increase in the coming decades. Hence the use of genetically derived resistance to supplement other pest control strategies is warranted. In soybean ( Glycine max (L.) Merr.), a major determinant for resistance is the quantitative trait locus, QTL-M. Glyma07g14530 , the gene underlying QTL-M, encodes a feeding-inducible flavonol 3- O -glycosyltransferase (F3GlcT or UGT78D2) that glucosylates kaempferol, as well as quercetin, myricetin, and isorhamnetin. The resistant allele has a premature stop codon in it, thus preventing the glucosylation and sequestration of flavonols in the vacuole, leading to a concomitant accumulation of proanthocyanidins and manifestation of resistance. Expressing the dominant (susceptible) allele in resistant plants restores susceptibility and silencing the susceptible allele results in resistance. The discovery and characterization of GmF3GlcT helps clarify the role of flavonoids in resistance to leaf-chewing insects and facilitates the development of insect-resistant cultivars that ultimately can lower production costs and reduce insecticide applications. Graphical The loss of flavonol 3-O-glucosyltransferase in soybean reduces feeding damage from defoliating insects and is accompanied by a concomitant increase in proanthocyanidins (red arrowhead). Significance: Unraveling the biochemical and genetic basis of soybean resistance to leaf-chewing insects facilitates the development of new, naturally insect resistant varieties. Such varieties contribute to on-farm profitability and reduced concerns over pesticide residues in the field.

Switchgrass, a forage and bioenergy crop, occurs as two main ecotypes with different but overlapping ranges of adaptation. The two ecotypes differ in a range of characteristics, including flowering time. Flowering time determines the duration of vegetative development and therefore biomass accumulation, a key trait in bioenergy crops. No causal variants for flowering time differences between switchgrass ecotypes have, as yet, been identified. In this study, we mapped a robust flowering time quantitative trait locus (QTL) on chromosome 4K in a biparental F₂ population and characterized the flowering-associated transcription factor gene PvHd1, an ortholog of CONSTANS in Arabidopsis and Heading date 1 in rice, as the underlying causal gene. Protein modeling predicted that a serine to glycine substitution at position 35 (p.S35G) in B-Box domain 1 greatly altered the global structure of the PvHd1 protein. The predicted variation in protein compactness was supported in vitro by a 4 °C shift in denaturation temperature. Overexpressing the PvHd1-p.35S allele in a late-flowering CONSTANS-null Arabidopsis mutant rescued earlier flowering, whereas PvHd1-p.35G had a reduced ability to promote flowering, demonstrating that the structural variation led to functional divergence. Our findings provide us with a tool to manipulate the timing of floral transition in switchgrass cultivars and, potentially, expand their cultivation range.

Suppression of immune response is a phenomenon that enables biological processes such as gamete fertilization, cell growth, cell proliferation, endophyte recruitment, parasitism, and pathogenesis. Here, we show for the first time that the Plasminogen-Apple-Nematode (PAN) domain present in G-type lectin receptor-like kinases is essential for immunosuppression in plants. Defense pathways involving jasmonic acid and ethylene are critical for plant immunity against microbes, necrotrophic pathogens, parasites, and insects. Using two Salix purpurea G-type lectin receptor kinases, we demonstrated that intact PAN domains suppress jasmonic acid and ethylene signaling in Arabidopsis and tobacco. Variants of the same receptors with mutated residues in this domain could trigger induction of both defense pathways. Assessment of signaling processes revealed significant differences between receptors with intact and mutated PAN domain in MAPK phosphorylation, global transcriptional reprogramming, induction of downstream signaling components, hormone biosynthesis and resistance to Botrytis cinerea . Further, we demonstrated that the domain is required for oligomerization, ubiquitination, and proteolytic degradation of these receptors. These processes were completely disrupted when conserved residues in the domain were mutated. Additionally, we have tested the hypothesis in recently characterized Arabidopsis mutant which has predicted PAN domain and negatively regulates plant immunity against root nematodes. ern1.1 mutant complemented with mutated PAN shows triggered immune response with elevated WRKY33 expression, hyperphosphorylation of MAPK and resistant to necrotrophic fungus Botrytis cinerea . Collectively, our results suggest that ubiquitination and proteolytic degradation mediated by the PAN domain plays a role in receptor turn-over to suppress jasmonic acid and ethylene defense signaling in plants.