Prevotella copri promotes white adipose browning and ameliorates adiposity

Liu, Shuang; Li, Haifang; Li, Rui; Chang, Yaxin; Wang, Senlin; Wang, Zhengang; Sun, Yawen; Li, Mengying; Liu, Min; Zhou, Yunlei; Jiao, Hongchao; Wang, Xiaojuan; Zhao, Jingpeng; Li, Bin; Zhang, Tongtong; Feng, Zhonghui; Xu, Junbo; Lin, Hai

doi:10.1038/s41467-025-67327-x

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Published: 13 December 2025

Prevotella copri promotes white adipose browning and ameliorates adiposity

Nature Communications volume 17, Article number: 680 (2026) Cite this article

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Abstract

Prevotella is a poorly understood symbiont within the gut microbiota, and its role in host health remains unclear. Here, we report a case-control analysis of a Chinese cohort, in which patients with obesity showed significantly lower fecal Prevotella copri abundance. The administration of P. copri can alleviate fat deposition and promote browning in inguinal white adipose tissue (iWAT) of mice with high-fat diet (HFD)-induced adiposity. We identify succinate as a key metabolite that activates succinate receptor 1 (SUCNR1) to mimic the effects of P. copri. Mice treated with either P. copri or succinate have increased iWAT-resident macrophage populations and IL-6-STAT3 pathway activation. Forkhead box M1 (FOXM1) directly upregulates IL-6 to activate IL-6-STAT3 signaling in P. copri- or succinate-induced iWAT browning. In human WAT, we identify that P. copri and succinate levels are positively correlated with TBX1 and UCP1 transcription. This study reveals the function of P. copri in promoting iWAT browning and suggests potential strategies for alleviating adiposity and associated disorders.

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Introduction

Obesity is an escalating epidemic linked to metabolic disorders like hepatic steatosis and insulin resistance¹. Accordingly, understanding the mechanisms of interventions for modulating lipid metabolism represents a long-standing goal of clinical research. There are three types of adipocytes in mammals: white, brown, and beige, the latter of which is a distinct subset of brown-like thermogenic adipocytes transdifferentiated from white adipose tissue (WAT)^2,3. Both mesenchymal stem cells located at the WAT perivascular region and some differentiated adipocytes are the progenitors for de novo beige adipogenesis^3,4,5. Brown and beige are thermogenic adipocytes that could be manipulated to alleviate metabolic disorders due to their specific expression of mitochondrial uncoupling protein 1 (UCP1)^2,3,4,5. In addition to UCP1, several beige-specific markers, such as transmembrane protein 26 (TMEM26) and T-box1 (TBX1), are known to be activated during the browning of WAT³. Emerging evidence demonstrated that the browning capability of subcutaneous WAT is substantially higher than the visceral WAT^2,3. Since brown adipose tissue (BAT) is present primarily in small mammals and human neonates but declines with age and obesity^2,6, stimulation of WAT browning represents a promising alternative intervention strategy to combat obesity and associated disorders^7,8.

The gut microbiome participates in regulating diverse host metabolic processes⁹, including thermogenesis in adipose tissues^10,11. Certain commensal gut bacteria (e.g. Ruminococcus torques, Limosilactobacillus reuteri, and Lactobacillus reuteri), could promote WAT browning and mitigate obesity or metabolic comorbidities. These effects are primarily mediated by specific bacterial metabolites, such as deoxycholic acid and short-chain fatty acids^12,13,14. Genus Prevotella is a pivotal keystone symbiont within the gut microbiota¹⁵, with Prevotella copri being the most widely known and abundant species¹⁶. However, host - P. copri interactions have received relatively little attention, and interpretations of this relationship remain controversial. For instance, intestinal P. copri abundance is reportedly higher in children with obesity¹⁷ but lower in adults with obesity versus healthy controls¹⁸. In pigs, P. copri promotes fat accumulation via activating inflammation¹⁹. However, in mice, P. copri improves glucose homeostasis under high-fiber intake by promoting hepatic glycogen storage²⁰, whereas it leads to insulin resistance and glucose intolerance when fed a high-fat diet (HFD)²¹. Despite mounting evidence of positive or negative links of P. copri to obesity and other metabolic disorders^{17,18,19,20,21}, whether and how P. copri may play a specific role in white adipose browning remains largely unknown.

In the present study, we found reduced fecal P. copri abundance in patients with obesity. In HFD-fed mice, oral P. copri administration increased inguinal WAT (iWAT) browning and attenuated weight gain. We identified succinate as its key metabolite, activating succinate receptor 1 (SUCNR1) to mimic P. copri’s effects. The macrophage-derived IL-6-STAT3 pathway, triggered by forkhead box M1 (FOXM1)-mediated Il-6 transcription, was essential for this P. copri- or succinate-inducible iWAT browning. Human data showed positive correlations between WAT TBX1/UCP1 expression and fecal P. copri/succinate levels. These findings propose P. copri-succinate signaling as a meaningful anti-obesity strategy.

Results

Individuals with obesity have aberrantly low abundance of P. copri in their fecal microbiomes

To explore potential relationships of gut microbes with obesity, we performed microbial metagenomics in individuals without obesity [Body mass index (BMI) < 28 kg/m², n = 15] and individuals with obesity (BMI ≥ 28 kg/m², n = 19) to investigate obesity-associated changes in fecal microbiota. The detailed information for the cohorts is presented in Supplementary Table 1. Linear discriminant analysis effect size (LEfSe) data and the heat maps revealed the total differences at phylum, order, family, genus, and species levels between the non-obesity and obesity groups (Fig. 1a, Supplementary Fig. 1a–e). We analyzed bacterial species that upregulated or downregulated in the two groups (Fig. 1b, c). Notably, among taxa with relative higher proportions in the fecal microbes, P. copri showed the most significant reduction in patients with obesity, with concomitant but less pronounced decrease in Coprococcus eutactus and Bifidobacterium dentium (Fig. 1d). We subsequently collected 92 samples from people with different BMI (detailed in Supplementary Data 1) and analyzed their fecal P. copri abundance by qPCR using specific primers²². In contrast to the non-obesity group, the abundance of P. copri was significantly lower in the people with obesity (Fig. 1e). Furthermore, inverse correlations were observed between P. copri abundance and several adiposity indices, including BMI, waist circumference, body fat percentage, and visceral fat area (Fig. 1f–i). These data indicated that people with obesity have aberrantly low abundance of P. copri in their fecal microbiomes.

Fig. 1: Individuals with obesity have low abundance of P. copri in their fecal microbiomes. — **Fig. 1: Individuals with obesity have low abundance of *P. copri* in their fecal microbiomes.**

Supplementation of HFD-induced adiposity model mice with P. copri alleviates adiposity and promotes iWAT browning

In light of the observed inverse association between P. copri abundance and obesity, we investigated this bacterium’s potential role in lipid metabolism. In a classic HFD murine model, male and female mice received P. copri (10⁸ CFU/kg BW) every other day for 8 weeks showed attenuated body weight gain, reduced fat deposition, and higher rectal temperature, whereas no effect was observed at the 10⁹ CFU/kg BW dosage (Supplementary Figs. 2–3). Given this dose-dependent response, coupled with our focus on white adipose browning and reported sexual dimorphism in Ucp1 and Tbx1 expression^23,24 (higher in females as shown in Supplementary Fig. 4), we subsequently administered live or pasteurized P. copri (10⁸ CFU/kg BW) exclusively to female HFD-fed mice for 8 weeks (denoted as HFD P. copri and HFD P. copri-pasteurized groups, respectively) (Fig. 2a).

qPCR analysis of feces and cecal contents after 6 weeks confirmed successful colonization, with P. copri abundance being significantly higher in live-bacterium-treated mice compared to pasteurized or saline controls (Fig. 2b, c). Mice receiving P. copri demonstrated significant resistance to HFD-induced metabolic alterations, including attenuated weight gain, reduced fat accumulation, decreased iWAT mass, and smaller adipocyte size after 8 weeks of treatment, without affecting lean mass or feed intake (Fig. 2d–l, Supplementary Fig. 5a). While P. copri administration showed a modest but non-significant reduction in epididymal WAT (eWAT) weight and adipocyte area, it had no observable effect on interscapular BAT (iBAT) mass or microstructure (Supplementary Fig. 5b–i). Notably, the P. copri-treated group exhibited significantly reduced hepatic lipid deposition and improved glucose tolerance and insulin sensitivity compared to HFD or P. copri-pasteurized Control (Supplementary Fig. 6). These results hinted that the prevention of iWAT expansion plays a pivotal role in P. copri-mediated protection against HFD-induced weight gain.

Decreased lipogenesis, increased lipolysis, and enhanced thermogenesis all could lead to reduction of WAT weight²⁵. At week 7 post-treatment, we first monitored murine core body temperature and respiratory metabolism. Mice administered P. copri exhibited significantly elevated rectal temperatures compared to both the P. copri-pasteurized or HFD Control groups (Fig. 2m). Respiratory metabolic analysis demonstrated substantial increases in O₂ consumption, CO₂ production, respiratory exchange ratio (RER) (VCO₂/VO₂), and heat production in the P. copri-treated mice (Fig. 2n–q). These collective changes indicate enhanced basal metabolic activity. Subsequent molecular analysis of iWAT, the most P. copri-responsive adipose depot, demonstrated no significant alterations in lipogenic markers PPARγ or AP2²⁶ protein expression, though Ap2 mRNA was downregulated (Fig. 2r–t). For lipolytic markers, expression of Atgl, Hsl and Cpt1²⁷ was unchanged across groups (Fig. 2r). Notably, the thermogenic marker Ucp1 and the beiging gene Tbx1³ were significantly elevated at both the mRNA and protein levels in the P. copri-treated iWAT (Fig. 2r-t), with immunofluorescence confirming pronounced UCP1 induction (Fig. 2u). These data established that P. copri prevents iWAT expansion primarily through Ucp1/Tbx1-mediated thermogenesis rather than lipogenic or lipolytic pathway modulation.

The primary P. copri metabolite, succinate, recapitulates the P. copri promotion of iWAT browning

The modulation of host metabolism by a specific bacterium is predominantly dependent on its metabolites, which act as potent signal transmitters for conveying information to target tissues^12,13,14. Our findings revealed that solely live, not pasteurized, P. copri alleviated HFD-induced adiposity and reduced WAT weight (Fig. 2d–j), indicating that its beneficial effects may rely on the production of bioactive metabolites. Consequently, targeted metabolomic analysis of media from standard P. copri cultures identified succinate as the primary metabolite (Fig. 3a). Furthermore, oral gavage of P. copri significantly elevated fecal and cecal succinate levels relative to the HFD control group (Fig. 3b, c). Collectively, these data confirmed succinate as the predominant metabolite of P. copri.

To determine whether succinate mediates P. copri-induced iWAT browning, we provided the HFD-fed mice with sodium succinate (SS) at 0.1 or 0.25 g/kg BW for 8 weeks (Fig. 3d). Mice received 0.25 g/kg BW SS exhibited significantly reduced weight gain, lower fat percentage, decreased iWAT weights, and smaller adipocyte sizes compared to HFD Control, with lower eWAT weight but no change in iBAT proportion or feed intake (Fig. 3e-m, Supplementary Fig. 7a-i). Given the role of P. copri in iWAT browning, its primary metabolite likely enhances thermogenesis. Supporting this hypothesis, the SS group showed elevated rectal temperature, CO₂ production, RER, and heat generation versus HFD Control (Fig. 3n-r). Furthermore, SS induced substantial upregulation of Ucp1/Tbx1 mRNA and protein levels, and higher UCP1-stained immunofluorescence intensity (Fig. 3s–v). Similar to P. copri, SS (0.25 g/kg BW) decreased hepatic TG accumulation and improved glucose homeostasis (Supplementary Fig. 8). For in vitro validation, primary iWAT-derived cells (iWATDCs)²⁸ were treated with P. copri cultures (P. copri sup) or SS at serum-equivalent concentrations from P. copri-treated mice. Both treatments elevated UCP1/TBX1 expression, decreased Oil red O-stained lipid droplets, and increased maximal oxygen consumption rate (SeaHorse assay) (Supplementary Fig. 9). These results established succinate as a bioactive molecule through which P. copri promotes iWAT browning.

SUCNR1 is required for P. copri or succinate promotion of iWAT browning

As a signaling molecule, succinate engages diverse cell types via its cognate receptor SUCNR1²⁹, prompting investigation into its necessity for P. copri- or SS-induced iWAT browning. Comparative analysis of Sucnr1 mRNA levels across murine adipose depots (iWAT, eWAT, iBAT) identified iWAT as the predominant site of expression (Fig. 4a). Quantitative assessment via RT-qPCR and WB revealed significant SUCNR1 upregulation at both mRNA and protein levels in iWAT from P. copri- or SS-treated mice versus HFD controls (Fig. 4b–d). For functional validation in iWATDCs, SUCNR1 inhibition using compound 4c abolished Ucp1 and Tbx1 induction by P. copri supernatant or SS (Supplementary Fig. 10). To further determine the importance of SUCNR1, in vivo confirmation employed Sucnr1^-/- female mice³⁰ subjected to oral P. copri or SS administration under HFD conditions (Fig. 4e). Body weight changes, fat mass, iWAT proportion, and adipocyte size remained comparable across control and treatment groups in Sucnr1^-/- mice (Fig. 4f–k). Moreover, rectal temperature decreased in supplemented groups, while O₂ consumption, CO₂ production, RER, and heat production showed no significant differences versus controls (Fig. 4l–p). Neither P. copri nor SS affected Tbx1 or Ucp1 mRNA levels nor UCP1 protein abundance in iWAT (Fig. 4q–s). ‌Collectively, these results demonstrated that SUCNR1 is essential for‌ P. copri ‌and succinate-mediated iWAT browning.

The IL-6-STAT3 pathway plays a crucial role for iWAT browning in response to P. copri or succinate

Given that succinate-SUCNR1 signaling is known to trigger inflammatory responses²⁹; we next examined serum levels of inflammatory factors, inflammatory gene expression in iWAT, and the macrophage proportion in iWAT. Among the tested inflammatory factors, serum IL-6 but not TNF-α levels were significantly increased in WT mice exposed to P. copri or SS (Fig. 5a, b).

Consistent with this, Il-6 gene transcript, IL-6 protein and STAT3 phosphorylation levels in iWAT also increased in both treatment groups (Fig. 5c-e). To trace the primary source of IL-6, in vitro confirmation was performed by adding P. copri sup or SS into cultures of RAW264.7 murine mononuclear macrophage leukemia cells³¹ or 3T3-L1 cells (a pre-adipocyte cell line that could differentiate into mature adipocytes)³². Analysis by RT-qPCR and WB revealed that P. copri sup- or SS-treated RAW264.7 cells exhibited significantly elevated Il-6 mRNA and protein levels, along with increased STAT3 phosphorylation accompanied by increased SUCNR1 expression (Supplementary Fig. 11a–c). Conversely, no alteration of Sucnr1, IL-6, p-STAT3, or UCP1 levels was observed in P. copri sup- or SS-exposed 3T3-L1 cells (Supplementary Fig. 11d–f). Moreover, we performed flow cytometry to quantify F4/80⁺/CD11b⁺/CD86⁺ macrophages within iWATDCs, as adipose tissue macrophages are established IL-6 secretors³³. Notably, both P. copri and SS treatments profoundly expanded the F4/80⁺/CD11b⁺ macrophage proportion and upregulated CD86 mean fluorescence intensity (MFI) in iWATDCs of WT mice (Fig. 5f, g).

To functionally define whether macrophages are essential for the beneficial in vivo effects of P. copri or succinate, macrophage populations were selectively ablated through tail vein injection of clodronate liposomes (CL), an established macrophage-depleting method validated in prior studies^34,35. Cohorts of CL-treated mice received either P. copri or SS for 8 weeks under HFD conditions (Supplementary Fig. 12a). It showed that the proportion of F4/80⁺ macrophages in circulation and iWAT was depleted by ~95% with CL (Supplementary Fig. 12b, c). P. copri or succinate did not alter the body weight gain, the body temperature, the white fat percentage, or thermogenic gene expression following macrophage depletion (Supplementary Fig. 12d-n). These findings collectively demonstrated that intact macrophage populations are indispensable for P. copri-derived succinate to exert its thermogenic effects.

The above findings and the direct IL-6-triggered p-STAT3 activation of Ucp1 expression through promoter binding²⁸ prompted a focus on IL-6. ChIP assays revealed that P. copri sup or SS could enhance p-STAT3 binding at both the Ucp1 and Tbx1 promoter regions in iWATDCs (Fig. 5h, i). To evaluate the necessity of IL-6-STAT3 signaling, we assessed iWAT browning in HFD-fed Il-6-KO mice exposed to P. copri or SS (Fig. 5j). Strikingly, Il-6-KO mice in both treatments exhibited no change in body weight, iWAT index, rectal temperature, O₂ consumption, heat production, and iWAT UCP1 and p-STAT3 contents, but increased iWAT adipocyte size, reduced CO₂ production and lower RER (Fig. 5k-z), indicating that neither P. copri nor SS could induce iWAT browning in the absence of IL-6. However, both treatments still activated SUCNR1 in iWAT of Il-6-KO mice (Fig. 5x-z), suggesting SUCNR1 operates upstream of IL-6. These results supported a role for the macrophage-derived IL-6-STAT3 pathway in P. copri- or succinate-mediated iWAT browning.

The transcription factor, FOXM1, positively regulates Il-6 expression

Given the essential role of ‌macrophage-derived IL-6/STAT3 signaling‌ in P. copri or SS-induced iWAT browning, we performed bulk RNA sequencing on iWAT from SS-treated and HFD control mice to identify transcription factors (TFs) regulating P. copri-/SS-inducible Il-6 expression. Principal component analysis (PCA) revealed clear separation of the HFD and SS groups (Fig. 6a). We identified 270 differentially expressed genes (edgeR; |log2FoldChange | >1; P value < 0.05; Fig. 6b) that including several TFs, among which, Foxm1 showed the highest up-regulation in SS-treated iWAT (Fig. 6c). Based on reported roles of ‌FOXM1 in macrophage migration and pro-inflammatory gene regulation³⁶ and its co-upregulation with Il-6 in SS-treated iWAT (Fig. 5c, Fig. 6c), we assessed Foxm1 expression across models. iWAT tissue‌, ‌primary iWATDCs‌, and ‌RAW264.7 macrophages‌ showed elevated Foxm1 mRNA/ protein after P. copri or SS exposure, but did not in 3T3-L1 adipocytes (Fig. 6d–f, Supplementary Fig. 13). This induction was ‌abolished by the SUCNR1 antagonist 4c in iWATDCs‌ (Fig. 6e). Crucially, lentiviral Foxm1 knockdown (via shRNA) in iWATDCs blocked P. copri sup/SS-induced upregulation of Foxm1, Il-6, Ucp1, Tbx1, and their protein products, as well as abrogated the activation of p-STAT3 (Fig. 6g, h). P. copri or SS failed to induce IL-6 when SUCNR1 was deficient or blocked (Supplementary Fig. 14a–d). However, Foxm1 remained inducible by P. copri or SS in Il-6-KO iWAT and iWATDCs but did not in iWAT from Sucnr1-KO mice (Supplementary Fig. 14e–j). These results suggested that ‌FOXM1 acts downstream of SUCNR1 but upstream of IL-6 in iWAT recognition and response to P. copri or SS‌.

Fig. 6: FOXM1 is an important transcription factor that positively regulates Il-6 expression. — **Fig. 6: FOXM1 is an important transcription factor that positively regulates *Il-6* expression.**

To verify whether FOXM1 could directly bind to the Il-6 promoter to regulate its transcription, we subsequently identified two 9 bp predicted binding sites (BS1, −699 to −707; BS2, −954 to −962) in the Il-6 promoter (Fig. 6i) based on previous literature³⁷ and the promoter prediction result from JASPAR. ChIP assays revealed that FOXM1 binding at both the Il-6 BS1 and BS2 sites was significantly enhanced in RAW264.7 cells (Fig. 6j, k), while being increased significantly at the BS1 and moderately at the BS2 region in iWATDCs, following P. copri sup or SS exposure (Fig. 6l, m). Furthermore, dual luciferase trans-activation assays showed that co-expression of pcDNA-FOXM1 could enhance luciferase expression driven by the Il-6 promoter compared to signal induced by either pcDNA-FOXM1 or the Il-6-LUC reporter alone (Fig. 6n), supporting the likelihood that FOXM1 could directly activate Il-6 transcription. Alternatively, deletion of either BS1, BS2, or both FOXM1 binding sites resulted in significantly lower luciferase expression under co-expression with pcDNA-FOXM1 compared to that driven by the WT Il-6 promoter (Fig. 6o, p). These results collectively demonstrated that FOXM1 acts as a direct regulator of Il-6 transcription by binding to either or both of the predicted binding sites (BS1 and BS2) within the Il-6 promoter.

Subcutaneous WAT TBX1 and UCP1 levels are positively correlated with P. copri abundance and succinate content in human individuals

In view of the P. copri or succinate promotion of iWAT browning, we explored links between thermogenic gene expression in human abdominal subcutaneous WAT and P. copri abundance or succinate contents. HPLC analysis showed significantly lower fecal and plasma succinate concentrations in individuals with obesity versus individuals without obesity (Fig. 7a, Supplementary Fig. 15a), aligning with reduced fecal P. copri abundance in the individuals with obesity (Fig. 1e). Notably, negative associations‌ were observed for fecal and plasma succinate levels with BMI (Fig. 7b, Supplementary Fig. 15b). Positive correlations‌ emerged between fecal P. copri abundance or succinate content and WAT UCP1/TBX1expression (Fig. 7c–f). Additionally, UCP1 and TBX1 protein levels were markedly lower in WAT from individuals with obesity compared to individuals without obesity (Fig. 7g). Given SUCNR1’s critical role in succinate-mediated WAT browning, we found reduced SUCNR1 mRNA and protein expression in WAT of individuals with obesity, with SUCNR1 mRNA positively correlating with plasma succinate concentrations (Supplementary Fig. 15c–e). Collectively, these data implicated reduced P. copri abundance, lowered succinate availability, and decreased SUCNR1 expression as factors in obesity pathogenesis, driven primarily by impaired WAT thermogenesis.

Fig. 7: WAT TBX1 and UCP1 levels positively correlate with the fecal P. copri abundance and succinate content in human individuals. — **Fig. 7: WAT *TBX1* and *UCP1* levels positively correlate with the fecal *P. copri* abundance and succinate content in human individuals.**

Discussion

This study identified decreased fecal P. copri abundance in people with obesity, aligning with reports linking Prevotella spp. to non-Westernized and non-obesity populations^38,39, but contrasting with pediatric studies showing positive P. copri-obesity correlations¹⁷. Despite being prevalent gut symbionts, Prevotella (especially P. copri) remains understudied possibly due to technical hurdles in bacterial cultivation and inconsistent findings on their host health impacts^20,40,41,42. We here demonstrated that P. copri at a relative low dosage (2–4 × 10⁶ CFU/mouse) could directly promote iWAT browning and ameliorate adiposity. This contrasts with prior correlational studies between P. copri and obesity or associated disorders^17,18, and opposes reports of P. copri (1 × 10⁹ CFU/mouse) increasing fat deposition in germ-free mice¹⁹. This discrepancy may stem from host species differences, dosage effects [higher doses (2–4 × 10⁷ CFU/mouse) failed to reduce fat deposition in our preliminary trials] and strain heterogeneity since phylogenetic analyses reveal P. copri comprises four human subspecies⁴¹, and the lack of standardized strains may explain inter-study variability.

The characterization of microbial-derived metabolites provides critical insights into the intricate interplay between gut microbiota and host metabolism. Our study identified succinate as a bioactive metabolite produced by P. copri. While Kovatcheva-Datchary et al.²⁰ observed elevated cecal succinate levels in P. copri-colonized mice, they ruled out its role in glucose homeostasis improvement. In contrast, we demonstrate that succinate functionally mimics P. copri’s ability to induce iWAT browning. As a metabolite intersecting host TCA cycle and microbial metabolic pathways⁴³, succinate exhibits context-dependent effects on host physiology largely depending on its origin, concentration, and target tissues/organs^{43,44,45,46,47,48}. Succinate has been shown to improve glycemic control via intestinal gluconeogenesis activation⁴³ and enhance thermogenic respiration in brown adipocytes through mitochondrial SLC25A10-dependent mechanisms⁴⁴. Paradoxically, succinate promotes inflammation^45,46 and is elevated in diabetic and metabolic syndrome rodent models^47,48. Our data reveal succinate’s role in iWAT browning via TBX1/UCP1 upregulation, countering HFD-induced adiposity. Clinically relevant correlations were observed between P. copri or succinate levels and subcutaneous WAT thermogenic markers in humans. Notably, in established adiposity models, P. copri inoculation or succinate administration attenuated weight gain (though statistically non-significant) while boosting iWAT thermogenic genes (Supplementary Fig. 16). These findings hinted the application potential of low dose P. copri or succinate in preventing and combating obesity.

In association with the P. copri and succinate promotion of iWAT browning, they also improve glucose homeostasis with lower insulin secretion (Supplementary Fig. 17) under HFD conditions. This context-dependent regulatory pattern of P. copri on glucose metabolism in our study seems to contrast sharply with Pedersen et al.’s findings indicating P. copri-mediated exacerbation of HFD-induced insulin resistance and glucose intolerance²¹. This discrepancy may arise from divergent administration regimens, whereby they administered P. copri at a higher dosage (5 × 10⁸ CFU/mouse twice weekly) compared to our protocol [2–4 × 10⁶ CFU/mouse (10⁸ CFU/kg BW) every two days]. However, Kovatcheva-Datchary et al.²⁰ paradoxically demonstrated that daily oral gavage of P. copri (10⁸ CFU/mouse) enhanced glucose homeostasis under high-fiber conditions. In fact, our investigation reveals a biphasic dose-response relationship: ‌lower dosages‌ (2–4 × 10⁶ CFU/mouse for P. copri or 0.25 g/kg succinate) significantly improve glucose tolerance and mitigate HFD-induced weight gain, whereas ‌higher dosages‌ (2–4 × 10⁷ CFU/mouse or 0.5 g/kg succinate) fail to confer metabolic protection at most circumstances. These findings further underscore the complex duality of P. copri’s metabolic effects, emphasizing the interplay between bacterial dosage, dietary context, and host metabolic outcomes, which align with its reported beneficial or detrimental actions in host health^{19,20,21,40,41,42}.

Subsequent investigations established that the P. copri-derived succinate- induced enhancement of iWAT browning exhibited at least partial dependence on SUCNR1 (also termed GPR91). This succinate receptor demonstrates expression in WAT, in both white adipocytes and WAT-associated macrophages^49,50. Our data revealed that the SUCNR1 in macrophages primarily mediates P. copri or succinate promotion of iWAT browning. One previous study showed that adipocyte-specific deletion of Sucnr1 inhibits WAT deposition in the context of a chow diet but, conversely, induces obesity and hepatic steatosis under HFD conditions⁵¹. McCreath et al.⁵² revealed dichotomous effects of complete Sucnr1-KO on obesity, showing protective effects against HFD-induced obesity during the initial period, but not at later stages. Collectively, these historical observations, in conjunction with our present findings, underscore the multifaceted regulatory role of SUCNR1 in lipid homeostasis, which depends largely on genetic manipulation strategies, dietary compositions, and animal’s developmental stages.

The role of succinate-SUCNR1 signaling in mediating P. copri’s metabolic effects motivated our systematic investigation of downstream mediators. Existing literature documents that SUCNR1 activation by accumulated succinate enhances proinflammatory cytokine production across multiple tissues^45,53. We discovered systemic and tissue-specific IL-6 elevation, and expanded F4/80⁺/CD11b⁺ macrophage population in iWAT. Il-6-KO models and macrophage-depleted (CL-treated) mice further confirmed macrophage- derived IL-6 as the principal effector. Notably, P. copri or SS also triggered mild inflammation activation in eWATs, but that increment was too weak to increase thermogenic gene expression (Supplementary Fig. 18). However, in livers, lower inflammation was found in the P. copri and succinate exposure group according to the flow cytometry and RT-qPCR results (Supplementary Fig. 19).

These tissue-specific reactions further demonstrate the complexity of the P. copri or succinate roles in host metabolism, which need intensive investigation in future studies. As a pleiotropic cytokine with both pro- and anti-inflammatory properties, the role of IL-6 in obesity and metabolic diseases remains controversial^54,55. IL-6 has been shown to trigger inflammation in adipose tissue, consequently antagonizing insulin activity and promoting metabolic abnormalities⁵⁴, whereas IL-6 and IL-6Rα could trigger alternative activation of macrophages and improve glucose homeostasis in mice with diet-induced obesity⁵⁵. In terms of thermogenesis, Il-6 gene delivery has been demonstrated to induce UCP1 expression in BAT as one possible mechanism leading to body weight reduction in rats⁵⁶. Phosphorylated STAT3 could bind to specific DNA response elements in the target gene promoter to modulate its transcription^54,56. We previously confirmed that p-STAT3^Tyr705 functions as a TF to induce Ucp1 expression in iWAT²⁸. Surprisingly, herein we also found P. copri or succinate exposure enhanced p-STAT3^Tyr705 interaction with the Tbx1 promoter. These results establish the macrophage IL-6-STAT3 axis as the core pathway for P. copri- or succinate-induced iWAT browning.

Strikingly, our current results demonstrate that FOXM1 acts downstream of SUCNR1 and directly binds to the Il-6 promoter at two different potential binding sites. As an important Fox family protein, FOXM1 has been shown to be involved in the migration and functional differentiation of immune cells to regulate several physiological and pathological processes^36,57,58. For example, macrophage-specific deletion of Foxm1 leads to decreased expression of proinflammatory genes, such as iNOS, Cox-2, Il-1β, and Il-6³⁶. Decreased FOXM1 expression in a mouse model of diabetes inhibits neutrophil and macrophage recruitment, resulting in delayed wound healing⁵⁷. This study thus represents a report of FOXM1 TF function in directly activating Il-6 expression, a key step in P. copri- or succinate-induced thermogenesis in iWAT.

In summary, the present investigation reveals that decreased abundance of P. copri in patients with obesity is strongly linked to lower TBX1 and UCP1 expression levels in WAT. P. copri exposure effectively promotes iWAT browning and counteracts adiposity via its metabolite, succinate, in a manner largely dependent on activation of the macrophage SUCNR1-FOXM1-IL-6- STAT3 axis (Fig. 8). These findings expand the scope of our understanding of the mechanisms of host-P. copri interplay, and suggest that P. copri and succinate may serve as promising interventions to combat diet-related obesity and associated disorders.

Fig. 8: The model diagram of how Prevotella copri promotes white adipose browning and ameliorates adiposity. — **Fig. 8: The model diagram of how *Prevotella copri* promotes white adipose browning and ameliorates adiposity.**

Methods

Human cohort analysis

Abdominal subcutaneous WAT was collected from patients who underwent bariatric surgery or cholecystectomy, and stored at −80 °C for detection of SUCNR1 and thermogenic gene expression. The BMI, body fat index, and visceral fat area of the participants were tested by a body composition analyzer (InBody770, Seoul, Korea). The waist circumference was measured by using a soft measuring tape.

For microbial metagenomic analysis, and fecal and plasma succinate content examination, 15 individuals without obesity (BMI < 28 kg/m², 4 males and 11 females) and 19 individuals with obesity (BMI ≥ 28 kg/m², 6 males and 13 females) were recruited (Supplementary Table 1). A total of 92 participants were recruited to quantify the relative abundance of fecal P. copri using qPCR (Supplementary Data 1), and 40 individuals participate in measuring the relationship between P. copri abundance and WAT UCP1/TBX1 mRNA (Supplementary Table 2).

Metagenomic sequencing of fecal microbiomes and data analysis

Total genomic DNA was extracted from 0.2 g of stool material via the PF Mag-Bind Stool DNA Kit (Omega Biotek, Norcross, GA, USA) following the manufacturer’s instructions. The DNA was fragmented to approximately 350 bp using Covaris M220 (Gene Company Limited, China) for paired-end library construction with NEXTFLEX Rapid DNA-Seq (Bioo Scientific, Austin, TX, USA). Paired-end sequencing was performed on an Illumina NovaSeq 6000 (Illumina Inc., San Diego, CA, USA) at Majorbio Bio-Pharm Technology Co., Ltd. (Shanghai, China) via a NovaSeq X Series 25B Reagent Kit according to the manufacturer’s instructions (www.illumina.com). The data were analyzed on the free online platform of the Majorbio Cloud Platform (www.majorbio.com).

P. copri cultivation and examination of organic acids in the bacterial culture supernatant

P. copri (strain DSM18205) from Deutsche Sammlung von Mikroorganismen und Zellkulturen (Braunschweig, Germany) was cultivated in KVLB (kanamycin-vancomycin laked blood) medium under anaerobic conditions at 37 °C. P. copri cultivated for 48 h was harvested in log phase, centrifuged at 1000 × g for 10 min, and washed twice with PBS. The precipitate was resuspended with PBS to a concentration of 10⁷ CFU/mL.

To detect the potential metabolites of P. copri, targeted metabolomics was carried out by using the bacterial culture supernatant (Sci-tech innovation quality inspection Co., Ltd, Qingdao, China). Briefly, 200 μL bacterial culture supernatant was mixed with 200 μL of ice-cold methanol and put under vortex movement for 1 min. The mixture was sonicated for 30 min and then incubated at 60 °C for 60 min, which was then centrifuged at 12,000 × g at 4 °C for 20 min. The supernatants were collected and analyzed via an Agilent 1260 HPLC system coupled to an Agilent 6120 mass spectrometer (California, USA). Chromatographic separation was performed at 30 °C with an Agilent AQ column (4.6 × 250 mm, 5 μm).

Animal experiments

The animals used in this study were C57BL/6 J mice. Il-6-KO mice were purchased from the Jackson Laboratory (Bar Harbor, ME, USA). Sucnr1-KO mice³⁰ were a gift from Professor Gang Shu of South China Agricultural University, China. The mice were housed in a controlled environment at Shandong Agricultural University and maintained under a standard 12:12 h light cycle with temperature kept at 21 °C–24 °C, and humidity at 40–70%. The animals had ad libitum access to feed and water and were fed a standard chow diet or a HFD with 60% fat energy (D12492, Research Diets, New Jersey, USA). P. copri (live and 65 °C pasteurized bacteria at a dosage of 10⁸ CFU/kg BW) and SS (0.1 or 0.25 g/kg BW) were administered intragastrically every two days during the trial periods, whereas normal saline was used as a control. The BW and feed intake of the mice were recorded weekly. Finally, after fasting for 12 h, blood was collected, and serum was prepared. The iWAT, iBAT, eWAT, and liver were harvested and weighed.

qPCR quantification of P. copri in human feces and mouse cecal contents and feces

Fecal or cecal DNA was extracted via a stool DNA extraction kit (Tiangen Biochemical Technology Co., Ltd., Beijing, China). The primers used for PCR-based amplification were as follows: Forward-TCGCTGACATGAGC GATAAC, Reverse-CCGTTGGCACTACCTTCATT, as previously described²². Quantitative PCR was performed via ChamQ SYBR Color qPCR Master Mix (Vazyme Biotech Co., Ltd., Nanjing, China) on an ABI Prism 7500 PCR system (Applied Biosystems, USA) at 95 °C for 1 min, followed by 40 cycles of 95 °C for 10 s and 60 °C for 30 s. A standard curve was constructed with various bacterial counts as described in the literature²².

Measurement of fecal, cecal and plasma succinate levels

Feces or ceal contents were mixed with extraction solvent containing 1 μM chlorpropamide, which was further dissolved in ice-cold 80% methanol (v/v) and homogenized with a KZ-II homogenizer. Plasma was mixed with ice-cold methanol and put under vortex movement for 1 min. The mixture was sonicated for 30 min and then incubated at 60 °C for 60 min, which was then centrifuged at 12,000 × g at 4 °C for 20 min. The measurement of succinate was performed via a liquid chromatography‒tandem mass spectrometry system composed of a HPLC system (Kyoto, Japan) coupled to a Sciex 4500 triple quadrupole linear ion trap mass spectrometer (AB SCIEX, Framingham, MA, USA). The liquid chromatography system was equipped with a XDB-C18 column (4.6 × 250 mm, 5 µm). The mobile phase comprised A (0.1% phosphoric acid in pure water) and B (acetonitrile) with a flow rate of 1 mL/min.

Body composition analysis of mice

The body composition of the mice was scanned and imaged via dual-energy X-ray absorptiometry (DXA) (MEDIKORS, Korea), the fat and lean indices were calculated via the system’s software. Each mouse was scanned and analyzed at least three times.

Glucose Tolerance Test (GTT) and Insulin Tolerance Test (ITT)

For GTT, mice in different groups were fasted for 12 h, and the baseline blood glucose was measured via tail vein by using a glucometer (time=0). Then the mice were intragastrically administered with glucose (2.0 g/kg BW), and the blood glucose concentrations were measured at 15, 30, 60, 90, and 120 min post-injection.

For ITT, mice were fasted for 6 h and the baseline blood glucose was measured (time=0). After injecting with insulin (0.75 U/kg BW) intraperitoneally, the blood glucose concentrations were monitored at 30, 60, 90, and 120 min.

Respiratory metabolic assay

Mice were individually placed in a Comprehensive Lab Animal Monitoring System (Coulumbus Instruments, USA) with 12 h of light and 12 h of darkness. The animals were allowed to eat and drink freely and were allowed to adapt for 24 h before formal testing. Data were collected every 5 min for another 24 h. O₂ consumption (VO₂), CO₂ production (VCO₂), and heat production were measured by the system, and the RER were calculated by VCO₂ /VO₂. The data were analyzed via ANCOVA (SPSS statistics 26.0).

Rectal temperature measurement

A digital thermometer equipped with a copper thermocouple probe was used to measure the rectal temperature. The probe was inserted 1.5 cm into the anal ducts of the mice.

Determination of serum IL-6 and TNF-α levels

Whole blood from the eyeballs of the mice was collected, put aside for 30 min, and subsequently centrifuged at 2000 g for 10 min to isolate the serum. The serum levels of IL-6 and TNF-α were quantified via commercial ELISA kits (Mlbio, Shanghai, China).

Measurement of serum indices

The serum ALT and AST activities, as well as the triglyceride (TG) and total cholesterol (TCH) contents, were assessed via commercial kits (Nanjing Jiancheng Bioengineering Institute, China).

Hematoxylin‒eosin (H&E) staining and immunofluorescence staining of UCP1

The tissue samples were fixed in 4% formaldehyde solution for more than 24 h. Subsequently, the tissues were embedded in paraffin and dissected into 5 μm sections for H&E staining.

For immunofluorescence staining, the dewaxed iWAT sections were treated with Triton X-100 and blocked in PBS supplemented with 5% fetal bovine serum (FBS). The sections were subsequently incubated overnight at 4 °C with a 1:100 diluted UCP1 antibody (sc-6529, Sacta Cruz, USA), followed by incubation with a fluorescent-labeled secondary antibody (1:500) and DAPI (1 μg/mL). Images were acquired via a confocal microscope (LSM880, Zeiss, Germany).

Isolation, cultivation, and differentiation of iWATDCs

Adipose tissue from the mouse inguinal region was digested with collagenase I (Solarbio Science & Technology Co., Ltd., Beijing, China) to isolate iWATDCs, a mixed cell type that containing adherent mesenchymal stem cells, macrophages, and a small quantity of fibroblasts, following previously established protocols⁵⁹. The isolated cells were cultured to subconfluence in low-glucose Dulbecco’s modified Eagle’s medium (DMEM) (HyClone, USA) supplemented with 15% FBS (Tianhang Biotechnology Co., Ltd., Hangzhou, China) and 100 units/mL penicillin/100 μg/mL streptomycin (Solarbio Science & Technology Co., Ltd., Beijing, China). The sub-confluent iWATDCs were induced to differentiate into adipocytes as previously described²⁸. SS (Macklin Biochemical Technology Co., Ltd., Shanghai, China) and P. copri culture supernatant, in the absence or presence of 0.1 μM 4c (Wuhe Biotechnology Co., Ltd., Shenyang, China), were added during the differentiation period.

Oil red O (ORO) staining of iWATDC-differentiated adipocytes

For ORO staining, iWATDC-differentiated adipocytes were rinsed with PBS and fixed in a 4% formaldehyde solution for 1 h. The cells were subsequently stained with a 0.6% ORO solution for 40 min, followed by hematoxylin staining for 1 min to facilitate the observation of cell morphology.

SeaHorse assay

Oxygen consumption rates (OCRs) were assessed using an Agilent Seahorse XFe24 Analyser (California, USA). iWATDCs were prepared and seeded in XF24 V28 cell culture microplates (Agilent, 100882-004) for 6 days’ adipogenic induction with AS in the presence or absence of P. copri sup or SS (0.1 mM). XFe24 sensor cartridges were hydrated according to the manufacturer’s protocol. Oligomycin, FCCP, and rotenone + antimycin A (R/A) (Agilent, 103015-100) were used at final concentrations of 1.5 µM, 1 µM, and 0.5 µM, respectively. The final OCR results were standardized to total cell number in each well.

Flow cytometry

iWATDCs were collected from iWAT, hepatic cells were were collected from Liver, rinsed twice and resuspended in PBS supplemented with 2% FBS to achieve a concentration of 1×10⁶ ~ 10⁷ cells/mL. The cells were subsequently stained with antibodies against specific cell surface markers, including PerCP-Cyanine 5.5-conjugated CD45.2 (45-0454-82, Thermo Fisher Scientific, USA), phycoerythrin (PE)-conjugated F4/80 (12-4801-82, Thermo Fisher Scientific, USA), APC-conjugated CD11b (17-0112-82, Thermo Fisher Scientific, USA), and eFluor™ 450-conjugated CD86 (48-0862-82, Thermo Fisher Scientific, USA). The staining process was carried out in the dark at 4 °C for 30 min. Flow cytometry analysis was performed using a BD FACSCanto™ II instrument (BD Biosciences, New Jersey, USA). Data analysis was performed via FlowJo software (Tree Star, Inc., Ashland, USA).

Cultivation and treatments of RAW264.7 cells

RAW264.7 cells were cultured in high-glucose DMEM supplemented with 10% FBS, along with 100 units/mL penicillin and 100 μg/mL streptomycin. The cells were maintained at 37 °C in a humidified atmosphere containing 5% CO₂. The sub-confluent cells were treated with SS and P. copri culture supernatant for 4 days, respectively.

Cultivation and adipogenic induction of 3T3L1 cell lines

3T3-L1 cells were cultured in high-glucose DMEM supplemented with 10% calf serum (Tianhang Biotechnology Co., Ltd., Hangzhou, China), along with 100 units/mL penicillin and 100 μg/mL streptomycin. The cells were maintained at 37 °C in a humidified atmosphere containing 5% CO₂.

For the adipogenic induction of 3T3-L1 differentiation, the cells were treated with differentiation-inducing medium containing 10 µg/mL insulin, 1 µM DEX, and 0.5 mM IBMX for 2 days. The induction medium was subsequently replaced with maintenance medium containing 10 µg/mL insulin for 2 days, as previously described⁶⁰.The induction hormones were subsequently removed, and the cells were cultured in standard growth medium until they reached full differentiation into mature adipocytes.

Macrophage depletion through intravenous injection of clodronate liposomes (CL)

Macrophage populations were selectively ablated through tail vein injection of clodronate liposomes (CL), an established macrophage-depleting agent validated in prior studies^34,35. In brief, CL (Yeasen Biotechnology, Shanghai, China) were injected into mice by tail vein twice a week (200 µL/time). The ablation efficiency was tested by flow cytometry measuring the proportion of F4/80⁺/Cd11b⁺ cells in iWAT and blood. After validation of about 95% macrophages were ablated, cohorts of CL-treated mice received either P. copri (10⁸ CFU/kg BW) or SS (0.25 g/kg BW) for another 8 weeks under HFD conditions.

RNA extraction and real-time quantitative PCR (RT-qPCR)

RNA extraction from both cell and tissue samples was carried out via RNAiso Plus Reagent (Takara, Japan). The extracted RNA was subsequently reverse transcribed into cDNA via a HiScript III All-in-one RT SuperMix Perfect for qPCR Kit (Vazyme Biotech Co., Ltd., Nanjing, China). RT-qPCR was performed via ChamQ Universal SYBR qPCR Master Mix (Vazyme Biotech Co., Ltd., Nanjing, China) on an ABI Prism 7500 PCR system (Applied Biosystems, USA), with an initial denaturation step at 95 °C for 30 s, followed by 40 cycles of denaturation at 95 °C for 15 s and annealing/extension at 60 °C for 30 s. The primer sequences utilized are detailed in Supplementary Data 2. Melting curve analysis was employed to validate the specificity of the primers. The relative mRNA expression levels of target genes were quantified via the 2^−ΔΔCt method and normalized to the expression level of the housekeeping gene Rpl13α in mice samples and gene 18S rRNA in human samples.

Protein extraction and western blotting

Protein was isolated from cells or tissues using RIPA lysis buffer supplemented with phosphatase and protease inhibitors (Roche, USA). Protein samples were separated by SDS‒PAGE and then transferred onto polyvinylidene fluoride (PVDF) membranes, which were incubated at room temperature for 15 min with TBST containing 5% non-fat milk to block nonspecific binding. The membranes were subsequently subjected to overnight incubation at 4 °C with specific primary antibodies, followed by incubation with corresponding HRP-conjugated secondary antibodies. The primary antibodies used in this study included UCP1 (sc-293418, Santa Cruz, USA), TBX1 (bs-21501R, Bioss, China), PPARγ (sc-7273, Santa Cruz, USA), AP2 (sc-271529, Santa Cruz, USA), SUCNR1 (AY4281, Abways, China), IL-6 (12912S, CST, USA), p-STAT3^Tyr705 (9145S, CST, USA), STAT3 (4904S, CST, USA), FOXM1 (sc-376471x, Santa Cruz, USA), β-actin (AB0035, Abways, China), and GAPDH (AB0038, Abways, China). The protein bands were visualized through chemiluminescence via an enhanced chemiluminescence (ECL) kit (Beyotime, China).

Transcriptome sequencing and data analysis

iWATs from control and SS-treated mice were collected for transcriptome sequencing. Total RNA was extracted via TRIzol reagent (Thermo Fisher Scientific, USA), and 4 µg of RNA was used to construct biologically triplicate sequencing libraries. Amplified libraries were sequenced on an Illumina HiSeq 2000 platform. Fastp (0.22.0) software was used to filter the sequencing data to get high quality sequence (Clean Data). The reference genome and gene annotation files were downloaded from genome website. The filtered reads were mapping to the reference genome using HISAT2 (v2.1.0). HTSeg (v0.9.1) statistics was used to compare the Read Count values on each gene as the original expression of the gene, and then Fragments Per Kilo bases per Million fragments (FPKM) values were used to standardize the expression. Then difference expression of genes was analyzed by edgeR with screened conditions as follows: expression difference multiple |log2FoldChange | >1, significant P value < 0.05.

Lentivirus packaging of shRNA and lentivirus infection

HEK293T cells were cultured in high-glucose DMEM supplemented with 10% FBS in a humidified atmosphere containing 5% CO₂ at 37 °C. Lentivirus production was carried out by using HEK293T cells. Briefly, HEK293T cells were transfected with a PSIH1-H1-CopGFP vector encoding shRNAs against Foxm1 (shRNA sequences are listed in Supplementary Table 3), along with the packaging plasmids pMD2. G, pMDL, and pRSV-Rev. The culture medium was replaced after 6 h, and the viral particles were harvested at 48 and 72 h post-transfection, respectively. iWATDCs were infected with lentivirus at 5 MOI in the presence of 8 µg/mL polybrene (Beyotime, China).

Chromatin immunoprecipitation (ChIP)

Sub-confluent iWATDCs or RAW264.7 cells were treated with P. copri supernatant and SS for 3 days, respectively. 1% (v/v) formaldehyde was used to cross-link the DNA and proteins. Subsequently, the chromatin DNA was sonicated into shorter pieces (200 ~ 1,000 bp) by using a Bioruptor Plus (Diagenode, Denville, NJ, USA) over 20 cycles of 30 s on and 30 s off. The DNA‒protein complexes were treated with protein A/G beads blocked with bovine serum albumin, followed by overnight incubation with specific antibodies for p-STAT3 or FOXM1, or control IgG. The cross-linked complexes were released by treatment with 5 M NaCl at 65 °C for 4 h. DNA was purified via a phenol‒chloroform extraction process. The eluted DNA was used as a template for qPCR. The input control was the supernatant collected before immunoprecipitation. The primer sequences utilized to amplify the Ucp1, Tbx1, and Il-6 promoter sequences are listed in Supplementary Data 2.

Dual luciferase assay

For the luciferase reporter assay, pGL3-basic luciferase reporter plasmids and the internal control renilla luciferase control vector were transfected into HEK293T cells via Lipofectamine 3000 (Thermo Fisher Scientific, USA). A Mut Express® II Fast Mutagenesis Kit V2 (Vazyme Biotech Co., Ltd., Nanjing, China) was used to construct mutated plasmids within the Il-6 promoter deleted of BS1, BS2, or the both regions. The primer sequences used for homologous recombination are provided in Supplementary Table 4. HEK293T cells were cultured in 96-well plates and grown to 70% confluence prior to transfection. The FOXM1 expression plasmid and different pGL3-basic-Il-6 promoter-luciferase reporter vectors were cotransfected into the cells for 24 h. The firefly and renilla luciferase activities of the cells were subsequently measured via a Duo-LiteTM Luciferase Assay System (Vazyme Biotech Co., Ltd., Nanjing, China), following the manufacturer’s instructions. The activity of the pGL3-basic-Il-6 promoter-luciferase reporter, normalized to that of the pRL-TK Rluc reporter, was compared between cells transfected with the FOXM1 expression plasmid or the control vector.

Statistical analysis

All data are expressed as mean ± SEM. Statistical analyses was performed via SPSS statistics 26.0 (SPSS, USA). Data analyses, such as bar charts and line charts were performed via GraphPad Prism 10.1.2 (GraphPad, USA). Parametric or non-parametric statistical tests were appropriately applied after Shapiro-Wilk test for the normal data distribution. For comparison between the two groups, an unpaired two-tailed t-test was used when samples were normally distributed, and the two-tailed Mann-Whitney U test was used when samples were not normally distributed. For comparisons among more than two groups, one-way ANOVA, followed by LSD (with equal variances) or Dunnett’s T3 (with different variances) post hoc test was used when samples were normally distributed, and Kruskal-Wallis test followed by Dunn’s post hoc test was used when samples were not normally distributed. All non-animal experiments were repeated independently three times with similar results. At least 3 independent samples were included in each experiment. Significance was established for P < 0.05, n.s indicates non-significant.

Ethics statement

For the human cohort, all participants were recruited from the Third People’s Hospital of Chengdu (Chengdu, China) and provided informed consent, which included a detailed explanation of the study’s purpose, procedures, and potential risks and benefits. We have obtained consent from all participants for the publication of identifiable information and the use of their data in the analysis and publication of the research findings. The study was approved by the Institutional Ethics Review Board of the Third People’s Hospital of Chengdu (Approval No: S-62 [2022]) and was carried out following the Chinese ethical guidelines for human genome/gene research.

For the animal experiments, ethical approval for all experimental procedures was obtained from the Animal Care and Use Committee of Shandong Agricultural University (Approval No: SDAU-2021-129).

Reporting summary

Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article.

Data availability

The metagenomic sequencing data generated in this study were deposited in the NCBI SRA database (accession code PRJNA1189811). The transcriptome sequencing data generated in this study were deposited in the NCBI SRA database (accession code PRJNA1189824). All data supporting the article are included in the main article, Supplementary Information, Supplementary Data 1, Supplementary Data 2, and the Source Data file. Source data are provided with this paper. The Source data file has been deposited in the Figshare repository [https://doi.org/10.6084/m9.figshare.30675941].

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Acknowledgements

We thank Gang Shu for gifting the Sucnr1-KO mice, Lifen Gao for gifting the RAW264.7 cell line, Wencheng Zhang and Jingyuan Li for assistance with respiratory metabolic assays, Wenchu Qian, Tiantian Zeng, and Yizhuo Chen for organizing reference materials. This work was supported by the National Natural Science Foundation of China (No. 32330101, 82202007, 82170887, H.L.), the National Key Research and Development Program of China (No. 2021YFD1300405, H.L.), the Natural Science Foundation of Shandong Province, China (No. ZR2023MC085, H.L., H.F.L), and the Third People’s Hospital of Chengdu Clinical Research Program (CSY-YN-01-2023-011, 076, 077, H.F.L). The graphical abstract was created with Figdraw.com (Copyright Certificate ID: TTSUP444f8).

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These authors contributed equally: Shuang Liu, Haifang Li.

Authors and Affiliations

College of Animal Science and Technology, Shandong Agricultural University, Tai’an, People’s Republic of China
Shuang Liu, Min Liu, Hongchao Jiao, Xiaojuan Wang, Jingpeng Zhao & Hai Lin
College of Life Sciences, Shandong Agricultural University, Tai’an, People’s Republic of China
Haifang Li, Rui Li, Yaxin Chang, Zhengang Wang, Yawen Sun, Mengying Li & Bin Li
Center of Gastrointestinal and Minimally Invasive Surgery, Department of General Surgery, the Third People’s Hospital of Chengdu, Chengdu, People’s Republic of China
Senlin Wang, Tongtong Zhang, Zhonghui Feng & Junbo Xu
College of Chemistry and Materials Science, Shandong Agricultural University, Tai’an, People’s Republic of China
Yunlei Zhou

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Shuang Liu
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Haifang Li
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Rui Li
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Yaxin Chang
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Senlin Wang
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Min Liu
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Hongchao Jiao
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Xiaojuan Wang
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Jingpeng Zhao
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Junbo Xu
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Contributions

Conceptualization: H.F.L., H.L., Z.H.F., J.B.X., and S.L.; data curation: S.L. and H.F.L.; formal analysis: S.L., H.F.L., R.L, Y.X.C., S.L.W., Z.G.W., Y.W.S., and M.Y.L.; funding acquisition: H.L., H.F.L., J.B.X., and Z.H.F.; investigation and resources: M.L., Y.L.Z., X.J.W., H.C.J., J.P.Z., B.L., and T.T.Z.; supervision: H.F.L., H.L., J.B.X., and Z.H.F.; writing – original draft: S.L., and H.F.L.; writing – review & editing, all authors.

Corresponding authors

Correspondence to Haifang Li, Zhonghui Feng, Junbo Xu or Hai Lin.

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Liu, S., Li, H., Li, R. et al. Prevotella copri promotes white adipose browning and ameliorates adiposity. Nat Commun 17, 680 (2026). https://doi.org/10.1038/s41467-025-67327-x

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Received: 18 November 2024
Accepted: 27 November 2025
Published: 13 December 2025
Version of record: 20 January 2026
DOI: https://doi.org/10.1038/s41467-025-67327-x