J-proteins in plant development and stress response

Dittmer, Sophie (2020): J-proteins in plant development and stress response. Dissertation, LMU München: Fakultät für Biologie

Abstract

Molecular chaperones are involved in a broad range of different processes, like protein folding, trafficking or degradation. Furthermore, many chaperones also play an important role during the response to different stresses. Since plants are sessile organisms, they need to be able to quickly adapt to different conditions. To do so, plants possess a complex chaperone machinery, composed of HSP70, HSP90, J-proteins and other factors. How the different chaperones cooperate, and in which processes they are involved in is so far not well understood. DJC31 and DJC62 are two J-proteins in Arabidopsis thaliana and have previously been described to be located inside the chloroplast. Using GFP localization studies and splitGFP, the actual localization could be determined to be the cytosolic side of the endoplasmic reticulum membrane, which could be confirmed by sucrose density centrifugation using isolated microsomes. Moreover, they were found to be attached to the membrane, which is presumably mediated by the N-terminal halves. Both proteins are composed of a long, disordered N-terminal part followed by several TPR repeats, forming two TPR domains, and a J-domain at the C terminus. This domain composition hints towards a function as co-chaperones of both HSP70 and HSP90. An interaction with cytosolic HSP70 and HSP90 could be shown by bimolecular fluorescence complementation. Additionally, performing a yeast two-hybrid library screening, potential client proteins of DJC31 could be identified. Knockout of either DJC31 or DJC62 caused only a mild phenotype, which was overall comparable to wild type. However, the double mutant exhibited severe defects in growth and development, which affected almost all organs. Furthermore, it could be shown that the double mutant is more sensitive to osmotic stress and treatment with abscisic acid, but surprisingly exhibited enhanced tolerance to drought. On the molecular level, up- or downregulation of DJC31 or DJC62 could not be observed under different stress conditions by qPCR. However, salt treatment of protoplasts, expressing the respective N-terminus of DJC31 or DJC62 fused to GFP, revealed that both proteins are released from the ER membrane into the cytosol under salt stress conditions, which indicates a regulation of DJC31 and DJC62 on the protein level. Transcriptome analysis could show, that under non-stress conditions expression levels of factors involved in biotic or abiotic stress response, as well as hormonal signaling are altered in the mutants. Taken together, these findings indicate that DJC31 and DJC62 might be regulators of different signaling pathways involved in development and stress response.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften, Biologie
• Fakultäten: Fakultät für Biologie
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 17. Dezember 2020
• 1. Bericht­erstatter:in: Schwenkert, Serena