Universität zu Köln

Zhang, Pingze ORCID: 0000-0002-6156-9722 (2023). S6K regulates inflammageing, immunosenescence and lifespan through the endolysosomal system. PhD thesis, Universität zu Köln.

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Abstract

Suppression of Target of Rapamycin complex 1 (TORC1) induces longevity and healthspan in diverse species. Suppression of S6 kinase (S6K) is an essential downstream mediator of the effect of TORC1 on ageing in Drosophila, but the mechanisms at work remain largely obscure. In this thesis, the role of S6K activity in ageing was investigated in Drosophila. I showed that reducing S6K activity ubiquitously in adult flies, using the GeneSwitch system, increased lifespan. Tissue-specific analysis revealed that the lifespan-extending effect was only observed when S6K activity was suppressed specifically in the fat body, the tissue that is functionally equivalent to the liver and white adipose tissue in mammals. I also found that expression of a constitutively active S6K protein in the fat body was sufficient to block the lifespan-extending effect of the TOR inhibitor rapamycin, suggesting that the activity of S6K specifically in the Drosophila fat body is essential for rapamycin-mediated longevity. Using proteomics profiling and network propagation analysis, I found that in young adults, the proteins directly affected by TORC1-S6K signalling were endosome/lysosome-, oogenesis-, and translation-related, while at later age immune-, translation-, and lipid-related processes were altered. Therefore, I tested the effect of TORC1-S6K signalling on fecundity, global translation and triacylglycerol homeostasis, but none of these were altered upon TORC1-S6K manipulation. Although the Drosophila intestine has recently been shown to play a crucial role in TORC1-dependent longevity, I found that S6K activity in the fat body does not affect gut health. By using electron microscopy, lysosome-specific live cell staining and genetic reporter flies, I found that TORC1 inhibition by rapamycin treatment repressed enlarged multilamellar lysosomes in the Drosophila fat body, and that this effect was blocked by elevating S6K activity. Inhibition of autophagy by Atg5 RNAi did not affect lysosomal size, but impairment of late endosomes by expressing a dominant negative form of Rab7 induced lysosomal enlargement. By proteomic analysis, I found that the protein level of Syntaxin 13 (Syx13), a SNARE family protein, was elevated by the inhibition of TORC1-S6K signalling. Furthermore, repressing Syx13 induced enlarged lysosomes in the fat body and overexpressing Syx13 attenuated the S6K-induced enlarged multilamellar lysosomes. Thus, Syx13 is a downstream effector of the TORC1-S6K pathway that mediates the lysosomal structural changes in Drosophila. During ageing in Drosophila chronic inflammation (termed “inflammageing” and mainly represented by the activation of the immune deficiency (IMD) pathway) increases, and pathogen clearance decreases (termed “immunosenescence”). Activation of the IMD pathway, represented by elevated nuclear translocalisation of the NF-κB like transcription factor Relish and expression of Diptericin A, and decline of pathogen clearance in old flies, were both suppressed by reduced S6K activity or rapamycin treatment, whereas activation of S6K in the fat tissue blocked this effect. Syx13 inhibition also blocked the rapamycin-related effect on IMD activation. S6K activation-related IMD activation and immune dysfunction were blocked by Syx13 overexpression. Furthermore, I found that ameliorating inflammageing by knocking down Relish level in the fat body from middle-adulthood on significantly improved bacterial clearance and extended fly lifespan. These results suggest that the TORC1-S6K-Syx13 signalling plays a crucial role in regulating the endolysosomal system and the aged immune system in the fat body of Drosophila. To assess if the regulation of Syx13 by rapamycin is also conserved in mammals, I used mice as a model system. I found that the chronic rapamycin treatment significantly increased Syntaxin 12/13 (Stx12) protein levels in the liver. The impact of reduced TORC1-S6K signalling on immunoageing was assessed in mice by proteomics profiling and network propagation analysis of liver samples from old mice treated with rapamycin and integrating these data with previous published transcriptome datasets of livers of aged mice with rapamycin treatment or S6K1 deficiency. I revealed that immune-related processes, such as inflammation and leukocyte proliferation, were commonly down-regulated by S6K1 deficiency or rapamycin treatment. These findings suggest that the regulation of Syntaxin 12/13 expression and its effect on immune ageing by TORC1-S6K signalling are conserved between flies and mammals. In summary, my work established that suppression of the TORC1-S6K-Syx13 axis ameliorates both inflammageing and immunosenescence in hepatic tissues through the endolysosomal system, and thereby extends longevity in Drosophila, providing a mechanistic explanation for the effects of rapamycin and suppression of S6K on immune function and lifespan in model organisms and, potentially, humans.

Item Type:	Thesis (PhD thesis)
Creators:	Creators Email ORCID ORCID Put Code Zhang, Pingze UNSPECIFIED orcid.org/0000-0002-6156-9722 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-704901
Date:	10 July 2023
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Außeruniversitäre Forschungseinrichtungen > MPI for Biology of Ageing
Subjects:	Natural sciences and mathematics Life sciences
Uncontrolled Keywords:	Keywords Language ageing UNSPECIFIED rapamycin UNSPECIFIED S6K UNSPECIFIED lysosome UNSPECIFIED inflammageing UNSPECIFIED immunosenescence UNSPECIFIED
Date of oral exam:	10 July 2023
Referee:	Name Academic Title Partridge, Linda Prof. Dr. Uhlirova, Mirka Prof. Dr. Schumacher, Björn Prof. Dr.
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/70490