PMID- 29951075
OWN - NLM
STAT- PubMed-not-MEDLINE
LR  - 20230926
IS  - 1664-462X (Print)
IS  - 1664-462X (Electronic)
IS  - 1664-462X (Linking)
VI  - 9
DP  - 2018
TI  - Using RT-qPCR, Proteomics, and Microscopy to Unravel the Spatio-Temporal 
      Expression and Subcellular Localization of Hordoindolines Across Development in 
      Barley Endosperm.
PG  - 775
LID - 10.3389/fpls.2018.00775 [doi]
LID - 775
AB  - Hordeum vulgare (barley) hordoindolines (HINs), HINa, HINb1, and HINb2, are 
      orthologous proteins of wheat puroindolines (PINs) that are small, basic, 
      cysteine-rich seed-specific proteins and responsible for grain hardness. Grain 
      hardness is, next to its protein content, a major quality trait. In barley, HINb 
      is most highly expressed in the mid-stage developed endosperm and is associated 
      with both major endosperm texture and grain hardness. However, data required to 
      understand the spatio-temporal dynamics of HIN transcripts and HIN protein 
      regulation during grain filling processes are missing. Using reverse 
      transcription quantitative PCR (RT-qPCR) and proteomics, we analyzed HIN 
      transcript and HIN protein abundance from whole seeds (WSs) at four [6 days after 
      pollination (dap), 10, 12, and >/=20 dap] as well as from aleurone, subaleurone, 
      and starchy endosperm at two (12 and >/=20 dap) developmental stages. At the WS 
      level, results from RT-qPCR, proteomics, and western blot showed a continuous 
      increase of HIN transcript and HIN protein abundance across these four 
      developmental stages. Miroscopic studies revealed HIN localization mainly at the 
      vacuolar membrane in the aleurone, at protein bodies (PBs) in subaleurone and at 
      the periphery of starch granules in the starchy endosperm. Laser microdissetion 
      (LMD) proteomic analyses identified HINb2 as the most prominent HIN protein in 
      starchy endosperm at >/=20 dap. Additionally, our quantification data revealed a 
      poor correlation between transcript and protein levels of HINs in subaleurone 
      during development. Here, we correlated data achieved by RT-qPCR, proteomics, and 
      microscopy that reveal different expression and localization pattern of HINs in 
      each layer during barley endosperm development. This indicates a contribution of 
      each tissue to the regulation of HINs during grain filling. The effect of the 
      high protein abundance of HINs in the starchy endosperm and their localization at 
      the periphery of starch granules at late development stages at the cereal-based 
      end-product quality is discussed. Understanding the spatio-temporal regulated 
      HINs is essential to improve barley quality traits for high end-product quality, 
      as hard texture of the barley grain is regulated by the ratio between HINb/HINa.
FAU - Shabrangy, Azita
AU  - Shabrangy A
AD  - Department of Ecogenomics and Systems Biology, University of Vienna, Vienna, 
      Austria.
FAU - Roustan, Valentin
AU  - Roustan V
AD  - Department of Ecogenomics and Systems Biology, University of Vienna, Vienna, 
      Austria.
FAU - Reipert, Siegfried
AU  - Reipert S
AD  - Core Facility Cell Imaging and Ultrastructure Research, University of Vienna, 
      Vienna, Austria.
FAU - Weidinger, Marieluise
AU  - Weidinger M
AD  - Core Facility Cell Imaging and Ultrastructure Research, University of Vienna, 
      Vienna, Austria.
FAU - Roustan, Pierre-Jean
AU  - Roustan PJ
AD  - Department of Ecogenomics and Systems Biology, University of Vienna, Vienna, 
      Austria.
FAU - Stoger, Eva
AU  - Stoger E
AD  - Department of Applied Genetics and Cell Biology, University of Natural Resources 
      and Life Sciences, Vienna, Austria.
FAU - Weckwerth, Wolfram
AU  - Weckwerth W
AD  - Department of Ecogenomics and Systems Biology, University of Vienna, Vienna, 
      Austria.
AD  - Vienna Metabolomics Center, University of Vienna, Vienna, Austria.
FAU - Ibl, Verena
AU  - Ibl V
AD  - Department of Ecogenomics and Systems Biology, University of Vienna, Vienna, 
      Austria.
LA  - eng
GR  - P 29303/FWF_/Austrian Science Fund FWF/Austria
PT  - Journal Article
DEP - 20180613
PL  - Switzerland
TA  - Front Plant Sci
JT  - Frontiers in plant science
JID - 101568200
PMC - PMC6008550
OTO - NOTNLM
OT  - RT-qPCR
OT  - barley endosperm
OT  - grain hardness
OT  - hordoindoline
OT  - label-free shotgun proteomics
OT  - laser microdissection
OT  - reference genes
OT  - spatio-temporal
EDAT- 2018/06/29 06:00
MHDA- 2018/06/29 06:01
PMCR- 2018/01/01
CRDT- 2018/06/29 06:00
PHST- 2018/01/21 00:00 [received]
PHST- 2018/05/22 00:00 [accepted]
PHST- 2018/06/29 06:00 [entrez]
PHST- 2018/06/29 06:00 [pubmed]
PHST- 2018/06/29 06:01 [medline]
PHST- 2018/01/01 00:00 [pmc-release]
AID - 10.3389/fpls.2018.00775 [doi]
PST - epublish
SO  - Front Plant Sci. 2018 Jun 13;9:775. doi: 10.3389/fpls.2018.00775. eCollection 
      2018.