Abstract
The proteasome is a key component for regulation of protein turnover across kingdoms. The proteasome has been shown to be involved in or affected by various stress conditions in multiple model organisms in plants. As such, studying proteasome homeostasis is crucial to understand its participation in different cellular conditions. However, the involvement of the proteasome in many cellular processes and its interplay with other degradation pathways hamper the interpretation of experiments based on a single approach. Thus, it is crucial to formulate a framework to investigate proteasome dynamics in different model organisms including plants. Here, we describe a pipeline to monitor proteasome homeostasis using four different methods including (i) luminescent-based proteasome activity measurement, (ii) immunoblot analysis of ubiquitinated proteins, (iii) evaluation of proteasome subunit protein levels, and (iv) monitoring of the proteasome stress regulon on mRNA levels using quantitative real-time PCR (polymerase chain reaction).
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Vierstra RD (2009) The ubiquitin-26S proteasome system at the nexus of plant biology. Nat Rev Mol Cell Biol 10:385–397
Langin G, Gouguet P, Üstün S (2020) Microbial effector proteins – a journey through the proteolytic landscape. Trends Microbiol 28:523–535
Xu FQ, Xue HW (2019) The ubiquitin-proteasome system in plant responses to environments. Plant Cell Environ 42:2931–2944
Wang H, Schippers JHM (2019) The role and regulation of autophagy and the proteasome during aging and senescence in plants. Genes (Basel) 10:267
Cai YM, Yu J, Ge Y et al (2018) Two proteases with caspase-3-like activity, cathepsin B and proteasome, antagonistically control ER-stress-induced programmed cell death in Arabidopsis. New Phytol 218:1143–1155
Han JJ, Yang X, Wang Q et al (2019) The β5 subunit is essential for intact 26S proteasome assembly to specifically promote plant autotrophic growth under salt stress. New Phytol 221:1359–1368
Üstün S, Sheikh A, Gimenez-Ibanez S et al (2016) The proteasome acts as a hub for plant immunity and is targeted by pseudomonas type III effectors. Plant Physiol 172:1941–1958
Sakamoto T, Sotta N, Suzuki T et al (2019) The 26s proteasome is required for the maintenance of root apical meristem by modulating auxin and cytokinin responses under high-boron stress. Front Plant Sci 10:1–13
Marshall RS, Vierstra RD (2019) Dynamic regulation of the 26S proteasome: from synthesis to degradation. Front Mol Biosci 6:40
Rabl J, Smith DM, Yu Y et al (2008) Mechanism of gate opening in the 20S proteasome by the proteasomal ATPases. Mol Cell 30:360–368
Marshall RS, Li F, Gemperline DC et al (2015) Autophagic degradation of the 26S proteasome is mediated by the dual ATG8/ubiquitin receptor RPN10 in Arabidopsis. Mol Cell 81:2053
Marshall RS, Vierstra RD (2018) Proteasome storage granules protect proteasomes from autophagic degradation upon carbon starvation. elife 7:1–38
Üstün S, Hafrén A, Liu Q et al (2018) Bacteria exploit autophagy for proteasome degradation and enhanced virulence in plants. Plant Cell 30:668–685
Üstün S, Bartetzko V, Börnke F (2013) The Xanthomonas campestris type III effector XopJ targets the host cell proteasome to suppress salicylic-acid mediated plant Defence. PLoS Pathog 9:e1003427
Üstün S, Börnke F (2015) The Xanthomonas campestris type III effector XopJ proteolytically degrades proteasome subunit RPT6. Plant Physiol 168:107–119
Book AJ, Gladman NP, Lee SS et al (2010) Affinity purification of the Arabidopsis 26 S proteasome reveals a diverse array of plant proteolytic complexes. J Biol Chem 285:25554–25569
Gladman NP, Marshall RS, Lee KH et al (2016) The proteasome stress regulon is controlled by a pair of NAC transcription factors in arabidopsis. Plant Cell 28:1279–1296
Sun HH, Fukao Y, Ishida S et al (2013) Proteomics analysis reveals a highly heterogeneous proteasome composition and the post-translational regulation of peptidase activity under pathogen signaling in plants. J Proteome Res 12:5084–5095
Zientara-Rytter K, Sirko A (2016) To deliver or to degrade – an interplay of the ubiquitin–proteasome system, autophagy and vesicular transport in plants. FEBS J 283:3534–3555
Saint-Marcoux D, Proust H, Dolan L et al (2015) Identification of reference genes for real-time quantitative PCR experiments in the liverwort marchantia polymorpha. PLoS One 10:1–14
Acknowledgments
This work was supported by an Emmy Noether Fellowship GZ: UE188/2-1 from the Deutsche Forschungsgemeinschaft (DFG, to S.Ü.) through the collaborative research council 1101 (CRC1101, to G.L.).
Author information
Authors and Affiliations
Corresponding authors
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Langin, G., Üstün, S. (2023). A Pipeline to Monitor Proteasome Homeostasis in Plants. In: Lois, L.M., Trujillo, M. (eds) Plant Proteostasis. Methods in Molecular Biology, vol 2581. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2784-6_25
Download citation
DOI: https://doi.org/10.1007/978-1-0716-2784-6_25
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-2783-9
Online ISBN: 978-1-0716-2784-6
eBook Packages: Springer Protocols