Dataset Information

Citrus vascular proteomics highlights the role of peroxidases and serine proteases during Huanglongbing disease progression

ABSTRACT: The dataset for this project was to identify significantly changing proteins in the vasculature of Washington Navel (Citrus sinensis (L) Osbeck) during Candidatus Liberibacter asiaticus (CLas) infection. The navel trees were graft-inoculated with the Hacienda Heights, CA CLas strain (HHCA) and then sampled ten months after graft inoculation.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Citrus Sinensis (sweet Orange) (citrus Aurantium Var. Sinensis)

TISSUE(S): Phloem, Plant Vascular Cell

SUBMITTER: Jessica Franco

LAB HEAD: Gitta Coaker

PROVIDER: PXD020703 | Pride | 2020-09-07

REPOSITORIES: Pride

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Publications

Citrus Vascular Proteomics Highlights the Role of Peroxidases and Serine Proteases during Huanglongbing Disease Progression.

Franco Jessica Y JY Thapa Shree P SP Pang Zhiqian Z Gurung Fatta B FB Liebrand Thomas W H TWH Stevens Danielle M DM Ancona Veronica V Wang Nian N Coaker Gitta G

Molecular & cellular proteomics : MCP 20200903 12

Huanglongbing (HLB) is the most devastating and widespread citrus disease. All commercial citrus varieties are susceptible to the HLB-associated bacterium, Candidatus Liberibacter asiaticus (CLas), which resides in the phloem. The phloem is part of the plant vascular system and is involved in sugar transport. To investigate the plant response to CLas, we enriched for proteins surrounding the phloem in an HLB susceptible sweet orange variety, Washington navel (Citrus sinen ...[more]

PMID: 32883801

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Project description:The experiment consists of M.hyp 232 cultures digests analyzed on two mass specs, LTQ Velos Pro and LTQ FT Ultra LTQ Velos Pro: Six cell culture replicates of M.hyo 232 were hydrophobically separated using tree detergents: Digitonin, Tween, and SDS. Each francion, including the insoluble pellet, was trypsin digested and then cleaned using an SCX trap follwed by a RP trap to remove detergents. Each fraction was sepated using a Dionex U3000 splitless nanoflow system operating at 333 nl per minute using a gradient of 2% ACN to 50% ACN in 4 hours. Eluate was analyzed using an LTQ Velos Pro mass spectrometer with 20 MS/MS scans of the 20 most intense peaks from each MS scan. Dynamic exclusion was enable for 3 minutes for each m/z with a repeat count of 1. LTQ FT Ultra: Two cell pellets for high resolution analysis were lysed and trypsin digested. Digested peptides were dried, resuspended in 20 mM KH2PO4, 20% ACN, pH 3 (Buffer A) in 2.5 µL and transferred to low retention vials in preparation for separation using an Ultimate 3000 configured for 2D-LC. Each sample was loaded at 15 µl/min onto an SCX microtrap for the first dimension of separation, involving SCX steps of Buffer A + 0, 5, 10, 15, 20, 25, 30, 40, 50, 100, 250, 500, and 1000 mM KCl. For the second dimension of separation, each eluted salt step was desalted with an inline peptide microtrap with 2% ACN, 0.1% FA at 5 µl/min. Once desalted, the microtrap was switched into line with a fritless nano column (75µm x ~10cm) containing C18 media (5µ, 200 Å Magic, Michrom). Peptides were eluted using a gradient of 2% to 36% ACN, 0.1% FA at 350 nl/min over 60 min and electrospray ionized for analysis using an LTQ FT Ultra mass spectrometer. A survey scan m/z 350-1750 was acquired in the FT ICR cell (Resolution = 100,000 at m/z 400, with an accumulation target value of 1,000,000 ions). Up to the 6 most abundant ions (>3,000 counts) with charge states > +2 were sequentially isolated and fragmented within the linear ion trap using collisionally induced dissociation with an activation q = 0.25 and activation time of 30 ms at a target value of 30,000 ions. M/z ratios selected for MS/ MS were dynamically excluded for 30 seconds. Analysis: X!tandem searches were performed using the Mycoplasma hyopneumoniae strain 232 reference protein set from NCBI. The only difference between the searches for the LTQ Velos and LTQ FT was the precursor mass tolenance being set to +-1500ppm and +-24ppm respectively. Decoy searches were performed and the data filtered at e-value <= 0.01 with single peptide proteins discarded. These results are included in the submission as two tab-delimited text files.

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Dataset Information

Citrus vascular proteomics highlights the role of peroxidases and serine proteases during Huanglongbing disease progression

Publications

Citrus Vascular Proteomics Highlights the Role of Peroxidases and Serine Proteases during Huanglongbing Disease Progression.

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