Renal NF-κB activation impairs uric acid homeostasis to promote tumor-associated mortality independent of wasting

Highlights

• Fly yki^3SA-gut tumors cause bacterial overload via suppression of gut PGRP-SC2

• Gut bacterial clearance increases survival of yki^3SA-tumor-bearing flies

• Renal-specific IMD-NF-κB blockade increases survival of yki^3SA-tumor-bearing flies

• Renal IMD-NF-κB blockade diminishes yki^3SA-tumor-associated uric acid accumulation

Summary

Tumor-induced host wasting and mortality are general phenomena across species. Many groups have previously demonstrated endocrinal impacts of malignant tumors on host wasting in rodents and Drosophila. Whether and how environmental factors and host immune response contribute to tumor-associated host wasting and survival, however, are largely unknown. Here, we report that flies bearing malignant yki^3SA-gut tumors exhibited the exponential increase of commensal bacteria, which were mostly acquired from the environment, and systemic IMD-NF-κB activation due to suppression of a gut antibacterial amidase PGRP-SC2. Either gut microbial elimination or specific IMD-NF-κB blockade in the renal-like Malpighian tubules potently improved mortality of yki^3SA-tumor-bearing flies in a manner independent of host wasting. We further indicate that renal IMD-NF-κB activation caused uric acid (UA) overload to reduce survival of tumor-bearing flies. Therefore, our results uncover a fundamental mechanism whereby gut commensal dysbiosis, renal immune activation, and UA imbalance potentiate tumor-associated host death.

Introduction

Cancer cachexia, or tumor-induced host wasting, is a process characterized by body weight loss, atrophy of skeletal and adipose tissue, hyperglycemia, and mortality (Fearon et al., 2013; Tisdale, 2002). Due to the unclear pathogenic mechanisms, different model organisms from rodents to Drosophila have been used to mimic the wasting development and address the unmet clinical needs. Recent studies using tumor-bearing mice have demonstrated the cachectic roles of circulating secreted proteins, such as activins, myostatin, transforming growth factor β (TGF-β), parathyroid hormone-related protein (PTHrP), interleukin-6 (IL-6), IL-1, tumor necrosis factor α (TNF-α), and interferon γ (IFN-γ), in mediating host organ wasting in response to tumor growth (Ballaro et al., 2016; Baracos et al., 2018). However, most of these mice studies were performed in specific-pathogen-free (SPF) conditions and ignored the potential participation of environmental microorganisms such as bacteria, fungi, and viruses that inevitably interact with the host to modulate host immune response and physiological outputs. Whether and how environmental factors and host immune response contribute to tumor-induced organ wasting or mortality are largely unknown.

Drosophila has emerged as an evolutionarily conserved model organism to investigate the mechanism of cancer cachexia in the past years. We have established that induction of an active transcription factor yki^3SA, a homolog of the human oncogene YAP1, in intestinal stem cells (ISCs) leads to overproliferation of gut tumor cells and subsequent wasting phenotypes, including muscle dysfunction, lipid loss, hyperglycemia, and mortality (Kwon et al., 2015). We further characterized the molecular mechanisms through which malignant yki^3SA-gut-tumor cells produce cachectic ligands, including ImpL2, Pvf1, and Upd3, to remotely impair anabolism-catabolism balance in host organs via modulation of insulin, MEK, and Jak/Stak signaling pathways, causing wasting phenotypes (Ding et al., 2021; Kwon et al., 2015; Song et al., 2019). Other groups using different tumor models in Drosophila and mice also observe similar cachectic roles of tumor-secreted proteins in the wasting development (Dong et al., 2021; Figueroa-Clarevega and Bilder, 2015; Kandarian et al., 2018; Newton et al., 2020; Nie et al., 2019; Yeom et al., 2021). These findings collectively demonstrate that tumor-induced wasting is a general phenomenon and that Drosophila can be used to dissect the molecular mechanisms involved in tumor-host interaction. Moreover, different from SPF mice, tumor-bearing flies are cultured at a less-restricted condition with direct interaction with natural air and external microorganisms and provide a convenient tool to investigate microbial-host interaction in the context of malignant tumor growth.

Drosophila has also emerged as an excellent model to conduct genetic analysis of microbial recognition, immune defense, and host physiological perturbation (Buchon et al., 2014; Capo et al., 2019; Charroux and Royet, 2012; Lee and Brey, 2013) due to the simpler gut commensal communities and conserved immune regulation. Two innate immune NF-κB pathways, Toll and immune deficiency (IMD), have been identified as the major signaling cascades to regulate the humoral reactions in response to the diverse bacteria and fungi (Lemaitre and Hoffmann, 2007). The Toll-NF-κB signaling is triggered by Gram-positive bacterial Lysine-type peptidoglycans (PGNs) and fungal β-(1,3)-glucan via binding to peptidoglycan recognition proteins (PGRPs) and glucan-binding proteins (GNBPs), respectively, to cleave cytokine Spaetzle (Spz) and activate Toll receptor as well as the Dorsal-Dif transcriptional complex (Ferrandon et al., 2007). On the other hand, Gram-negative bacterial DAP-type PGNs trigger IMD-NF-κB signaling through interaction with PGRP-LC to activate IMD and the downstream transcriptional factor Relish (Rel) (Ferrandon et al., 2007). A subgroup of PGRPs, such as PGRP-LB with the amidase enzymatic activity, have been found to degrade PGNs and suppress the immune responses upon infection (Charroux et al., 2018; Zaidman-Remy et al., 2006). Activated Toll and IMD pathways in multiple tissues like gut and fat body induce expression of anti-microbial peptide (AMP) genes such as Drosomycin (Drs) and Diptericin (Dpt), respectively, and others to restrain the microbial growth in a feedback loop (Lemaitre and Hoffmann, 2007; Royet and Dziarski, 2007). In addition, activation of NF-κB pathways in Drosophila also involves regulation of energy balance and host death (Buchon et al., 2014; Garschall and Flatt, 2018; Molaei et al., 2019), the major features of tumor-induced host wasting or mortality. It raises a hypothesis whereby systemic immune response plays pathogenic roles in the context of tumor growth.

In this study, we observed a robust systemic activation of IMD-NF-κB signaling in yki^3SA-tumor bearing flies due to suppressed production of gut PGRP-SC2, a key antibacterial amidase characterized by us, and increased abundance of commensal bacteria. We found that bacteria-induced IMD-NF-κB activation in the renal organ impaired UA metabolism to decrease survival of yki^3SA flies without affecting tumor growth or energy wasting. Thus, our results indicate that gut-microbial dysbiosis and renal-IMD-NF-κB activation promote mortality independent of host wasting in yki^3SA-tumor-bearing flies.