Auxin metabolome profiling in the arabidopsis endoplasmic reticulum using an optimised organelle isolation protocol

0551225 - ÚEB 2022 RIV CH eng J - Článek v odborném periodiku
Včelařová, Ludmila - Skalický, Vladimír - Chamrád, Ivo - Lenobel, René - Kubeš, Martin - Pěnčík, Aleš - Novák, Ondřej
Auxin metabolome profiling in the arabidopsis endoplasmic reticulum using an optimised organelle isolation protocol.
International Journal of Molecular Sciences. Roč. 22, č. 17 (2021), č. článku 9370. E-ISSN 1422-0067
Grant CEP: GA ČR(CZ) GJ17-21581Y; GA MŠMT(CZ) EF16_019/0000827
Institucionální podpora: RVO:61389030
Klíčová slova: Auxin * Density gradient centrifugation * Endoplasmic reticulum * Mass spectrometry * Subcellular fractionation
Obor OECD: Biochemistry and molecular biology
Impakt faktor: 6.208, rok: 2021
Způsob publikování: Open access
http://doi.org/10.3390/ijms22179370

The endoplasmic reticulum (ER) is an extensive network of intracellular membranes. Its major functions include proteosynthesis, protein folding, post-transcriptional modification and sorting of proteins within the cell, and lipid anabolism. Moreover, several studies have suggested that it may be involved in regulating intracellular auxin homeostasis in plants by modulating its metabolism. Therefore, to study auxin metabolome in the ER, it is necessary to obtain a highly enriched (ideally, pure) ER fraction. Isolation of the ER is challenging because its biochemical properties are very similar to those of other cellular endomembranes. Most published protocols for ER isolation use density gradient ultracentrifugation, despite its suboptimal resolving power. Here we present an optimised protocol for ER isolation from Arabidopsis thaliana seedlings for the subsequent mass spectrometric determination of ER-specific auxin metabolite profiles. Auxin metabolite analysis revealed highly elevated levels of active auxin form (IAA) within the ER compared to whole plants. Moreover, samples prepared using our optimised isolation ER protocol are amenable to analysis using various “omics” technologies including analyses of both macromolecular and low molecular weight compounds from the same sample.
Trvalý link: http://hdl.handle.net/11104/0326672