title: Putative protease inhibitor gene discovery and transcript profiling during fruit development and leaf damage in grapefruit (Citrus paradisi Macf.).
abstract: Seven putative protease inhibitor (PPI) cDNAs, representing four protein families, were isolated from a grapefruit (Citrus paradisi Macf. Cv. Marsh) immature fruit flavedo cDNA library. Cloned open reading frames encoded proteins with similarity to, and protein signatures for: legume Kuntiz inhibitors (lkiL-1, lkiL-2, lkiL-3), potato trypsin inhibitor I (ptiIL-1), serpins (serpL-1), cystatins (cystL-1), and gamma thionins (gthL-1). Response of transcript abundance to fruit development and leaf wounding was determined for all but lkiL-1 using real-time RT-PCR. Immature leaves had the highest transcript levels for all PPIs. The gthL-1 transcript in immature leaves was the most abundant transcript but was absent from healthy mature leaves. In fruit flavedo, transcripts for all PPIs were most abundant in youngest fruit (<15 mm dia. fruit), and declined during development, but displayed different patterns of developmental change. Mechanical or Diaprepes root weevil (DRW) feeding damage to leaves caused a <10-fold reduction or had no effect on transcript level with the exception of gthL-1 which, as a result of damage, increased >50-fold in mature leaves and decreased >1400-fold in immature leaves. This developmental control of transcript response to wounding in a woody perennial is opposite of what has been observed for defensive proteinase inhibitors (PIs) in other plants (typically herbaceous and/or annual plants), where younger leaves typically invoke a higher defensive proteinase inhibitor transcript accumulation than older tissues. Except for gthL-1, the PPI transcripts were minimally responsive or unresponsive to wounding. Changes in PPI transcript levels suggest diverse roles for the products of these genes in citrus, with only gthL-1 responding in a defense-like manner.
authors.affiliation: USDA, ARS, U.S. Horticultural Research Laboratory, 2001 South Rock Road, Fort Pierce, FL 34945, USA. rshatters@ushrl.ars.usda.gov
authors.forename: Robert G
authors.initials: RG
authors.lastname: Shatters
authors.forename: Michael G
authors.initials: MG
authors.lastname: Bausher
authors.forename: Wayne B
authors.initials: WB
authors.lastname: Hunter
authors.forename: José X
authors.initials: JX
authors.lastname: Chaparro
authors.forename: Phat M
authors.initials: PM
authors.lastname: Dang
authors.forename: Randall P
authors.initials: RP
authors.lastname: Niedz
authors.forename: Richard T
authors.initials: RT
authors.lastname: Mayer
authors.forename: T Greg
authors.initials: TG
authors.lastname: McCollum
authors.forename: Xiomara
authors.initials: X
authors.lastname: Sinisterra
mesh_terms: D002957:Citrus / Q000235:genetics* / Q000254:growth & development; D015853:Cysteine Proteinase Inhibitors / Q000235:genetics*; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D020224:Expressed Sequence Tags; D005638:Fruit / Q000235:genetics* / Q000254:growth & development; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D015723:Gene Library; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000235:genetics*; D010940:Plant Proteins / Q000235:genetics*; D017422:Sequence Analysis, DNA; D013314:Stress, Mechanical; D014158:Transcription, Genetic
chemical_list: D015853:Cysteine Proteinase Inhibitors; D018076:DNA, Complementary; D010940:Plant Proteins