Peptidomics Approaches for the Identification of Bioactive Molecules from Diaphorina citri
- Laura A. FleitesLaura A. FleitesBoyce Thompson Institute for Plant Research, Ithaca, New York 14853, United StatesUSDA Agricultural Research Service, Robert W. Holley Center for Agriculture and Health, Ithaca, New York, 14853-2901, United StatesDepartment of Plant Pathology and Plant Microbe Biology, Cornell University, Ithaca, New York 14850-5905, United StatesMore by Laura A. Fleites
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- Richard JohnsonRichard JohnsonDepartment of Genome Sciences, University of Washington, Seattle, Washington 98195, United StatesMore by Richard Johnson
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- Angela R. KruseAngela R. KruseBoyce Thompson Institute for Plant Research, Ithaca, New York 14853, United StatesDepartment of Plant Pathology and Plant Microbe Biology, Cornell University, Ithaca, New York 14850-5905, United StatesMore by Angela R. Kruse
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- Ronald J. NachmanRonald J. NachmanUSDA Agricultural Research Service, Insect Control and Cotton Disease Research Unit, College Station, Texas 77845, United StatesMore by Ronald J. Nachman
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- David G. HallDavid G. HallUSDA Agricultural Research Service, US Horticulture Research Laboratory, Fort Pierce, Florida 34945, United StatesMore by David G. Hall
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- Michael MacCossMichael MacCossDepartment of Genome Sciences, University of Washington, Seattle, Washington 98195, United StatesMore by Michael MacCoss
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- Michelle L. Heck*Michelle L. Heck*E-mail: [email protected]Boyce Thompson Institute for Plant Research, Ithaca, New York 14853, United StatesUSDA Agricultural Research Service, Robert W. Holley Center for Agriculture and Health, Ithaca, New York, 14853-2901, United StatesDepartment of Plant Pathology and Plant Microbe Biology, Cornell University, Ithaca, New York 14850-5905, United StatesMore by Michelle L. Heck
Abstract
Huanglongbing (HLB), a deadly citrus disease, is primarily associated with Candidatus Liberibacter asiaticus (CLas) and spread by the hemipteran insect Diaphorina citri. Control strategies to combat HLB are urgently needed. In this work, we developed and compared workflows for the extraction of the D. citri peptidome, a dynamic set of polypeptides produced by proteolysis and other cellular processes. High-resolution mass spectrometry revealed bias among methods reflecting the physiochemical properties of the peptides: while TCA/acetone-based methods resulted in enrichment of C-terminally amidated peptides, a modification characteristic of bioactive peptides, larger peptides were overrepresented in the aqueous phase of chloroform/methanol extracts, possibly indicative of reduced co-analytical degradation during sample preparation. Parallel reaction monitoring (PRM) was used to validate the structure and upregulation of peptides derived from hemocyanin, a D. citri immune system protein, in insects reared on healthy and CLas-infected trees. Mining of the data sets also revealed 122 candidate neuropeptides, including PK/PBAN family neuropeptides and kinins, biostable analogs of which have known insecticidal properties. Taken together, this information yields new, in-depth insights into peptidomics methodology. Additionally, the putative neuropeptides identified may lead to psyllid mortality if applied to or expressed in citrus, consequently blocking the spread of HLB disease in citrus groves.
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