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Identification of novel potential interaction partners of UDP-galactose (SLC35A2), UDP-N-acetylglucosamine (SLC35A3) and an orphan (SLC35A4) nucleotide sugar transporters

ABSTRACT: Nucleotide sugar transporters (NSTs) are ER and Golgi-resident members of the solute carrier 35 (SLC35) family which supply substrates for glycosylation by exchanging lumenal nucleotide monophosphates for cytosolic nucleotide sugars. Defective NSTs have been associated with congenital disorders of glycosylation (CDG), however, molecular basis of many types of CDG remains poorly characterized. To better understand CDG biology, we identified potential interaction partners of UDP-galactose transporter (SLC35A2), UDP-N-acetylglucosamine transporter (SLC35A3) and an orphan nucleotide sugar transporter SLC35A4. For this purpose, each of the SLC35A2-A4 proteins was used as a bait in four independent pull-down experiments and the identity of the immunoprecipitated material was discovered using MS techniques. The consensus findings for each of the NSTs tested were listed as potential interaction partners and should prove useful as a starting point for deciphering the NST interaction network.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Maciej Wiktor

PROVIDER: MSV000086622 | MassIVE | Mon Dec 21 07:58:00 GMT 2020

SECONDARY ACCESSION(S): PXD023254

REPOSITORIES: MassIVE

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Project description:Secondary metabolites play a key role in coordinating ecology and defense strategies of plants. Diversity of these metabolites arise by conjugation of core structures with diverse chemical moieties, such as sugars in glycosylation. Active pools of phytohormones, including those involved in plant stress response are also regulated by glycosylation. While, much is known about the enzymes involved in glycosylation, we know little about their regulation or coordination with other processes. We characterized the flavonoid pathway transcription factor, TRANSPARENT TESTA 8 (TT8) in Arabidopsis thaliana, using an integrative omics strategy. This approach provides a systems level understanding of the cellular machinery that is used to generate metabolite diversity by glycosylation. Metabolomics analysis of TT8 loss-of-function and inducible overexpression lines showed that TT8 coordinates glycosylation of not only flavonoids, but also nucleotides, thus, implicating TT8 in regulating pools of activated nucleotide sugars. Transcriptome and promoter network analyses revealed that TT8 regulome included sugar transporters, proteins involved in sugar binding and sequestration, and a number of carbohydrate active enzymes. Importantly, TT8 affects stress response, along with brassinosteroid and jasmonic acid biosynthesis, by directly binding to the promoters of key genes of these processes. This combined effect on metabolites glycosylation and stress hormones by TT8 inducible overexpression led to significant increase in tolerance towards multiple abiotic and biotic stresses. Conversely, loss of TT8 leads to increased sensitivity to these stresses. Thus, the transcription factor TT8 is an integrator of secondary metabolism and stress response. These findings provide novel approaches to improve broad-spectrum stress tolerance. Gene expression analysis for 6-day old Arabidopsis thaliana seedling were performed for TT8 loss-of-function line, tt8-3 (in Ws background), and its wild type background control, Ws. Two independent biological replicates for each line were hybridized onto two slides of Agilent SurePrint G2 Agilent Arabidopsis V4 (4x44K), with each slide having two technical replicates for each line. Contributors: Amit Rai, Shivshankar Umashankar, Megha, Lim Boon Kiat, Johanan Aow Shao Bing, Sanjay Swarup.

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Identification of novel potential interaction partners of UDP-galactose (SLC35A2), UDP-N-acetylglucosamine (SLC35A3) and an orphan (SLC35A4) nucleotide sugar transporters

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