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Oxidative stress controls lncRNA-mediated sow granulosa cell functions in a FoxO1-dependent manner

Journal of Animal Science and Biotechnology volume 15, Article number: 171 (2024) Cite this article

Abstract

Background

Oxidative stress (OS) is involved in low female fertility by altering multi-omics such as the transcriptome, miRome, and lncRNome in follicular cells and follicular fluid. However, the mechanism by which OS affects multi-omics dynamics remains largely unknown. Here, we report that OS induces lncRNome dynamics in sow granulosa cells (sGCs), which is partially dependent on the transcription factor activity of its effector, FoxO1.

Results

A total of 2,283 putative FoxO recognition elements (FREs) were identified in the promoters of 394 lncRNAs, accounting for 91.20% (394/432) of the lncRNAs regulated by OS. ChIP and reporter assays showed that the effector FoxO1 mediated OS regulation of lncRNA transcription in a transcription factor activity-dependent manner. In sGCs, OS induces the transcription and function (e.g., apoptosis) of NORSF (non-coding RNA involved in sow fertility), a nuclear lncRNA involved in sGC function via FoxO1. Furthermore, FoxO1 has been identified as a transcriptional activator of NORSF in sGCs that interacts with the FRE motif of its promoter. Meanwhile, OS downregulates the transcription of CYP19A1, which encodes an essential enzyme for estrogen synthesis and 17β-estradiol (E2) release by sGCs via the FoxO1 and NORSF axis. Phenotypically, dysregulation of NORSF transcription caused by 2 novel adjacent transitions in the promoter leads to decreased sow fertility.

Conclusion

These results suggest a model of OS-stimulated lncRNome dynamics in sGCs and a new signaling pathway of OS that influences sGC function and sow fertility.

Introduction

Gene-environment interactions (GEI) determine all phenotypes of organisms, including sow reproductive traits, and their levels determine the economic benefits for the swine industry. Oxidative stress (OS) is a common mechanism underlying various internal and external environmental factors (e.g., heat stress, density, social rank, and feed mycotoxins) that result in low fertility [1,2,3]. Measures to reduce OS, such as adding antioxidant stress additives to the feed or reducing density, can improve sow fertility [1, 3,4,5]. OS leads to various abnormalities (e.g., apoptosis, autophagy, meiotic arrest, and oxidative damage) and multiple omics disorders in follicular cells, including sow granulosa cells (sGCs), oocytes, luteal cells of sow ovaries, while antioxidants can reverse these processes [6,7,8,9]. A more recent report showed that OS affects sGC functions and sow fertility through miR-23a, the first causal microRNA (miRNA) for sow fertility traits [10].

Interestingly, OS has also been shown to control sow ovarian function by interacting with non-coding RNAs (ncRNAs), including miRNAs and long ncRNAs (lncRNAs) [7, 11, 12]. Among the miRNAs, 55 were found to participate in the stimulation of OS on sGC apoptosis [7]. OS induces sGC apoptosis and oxidative damage via the miR-192 and Acvr2a axis [11]. On the other hand, several miRNAs (e.g., miR-183 cluster and let-7a) rescue OS-induced sGC apoptosis by interacting with FoxO1, a well-known sensor and effector of OS [13] and an apoptosis-inducing factor [14]. Among lncRNAs, SDNOR restrained by OS suppresses OS-induced sGC apoptosis and maintains the state and function of sGCs [12]. Additionally, NORHA, a lncRNA upregulated during sow follicular atresia, can synergistically induce FoxO1 expression with OS, thereby increasing the rate of sGC apoptosis [13]. Overall, little is known about the interactions between OS and ncRNAs in the regulation of sow ovarian function.

We previously demonstrated that OS can cause significant changes in lncRNAome dynamics in sGCs, and identified hundreds of differentially expressed lncRNAs (DElncRNAs) [12]. However, the mechanism by which OS influences the transcription of these DElncRNAs is still unclear. The effector of OS, FoxO1, is also a transcription factor (TF) that activates or inhibits target genes by interacting with the FoxO1 response elements (FREs) in their promoters [15, 16]. In this study, we aimed to explore the mechanism by which OS regulates DElncRNA transcription from the perspective of the TF activity of its effector FoxO1.

Materials and methods

Materials

The following reagents and kits were used: TRIzol (#RG-51001 A; Angle Gene, Nanjing, China), H2O2 (#MFCD00011333; Sinopharm, Shanghai, China), Lipofectamine 2000 (#11668500; Thermo Fisher Scientific, Waltham, MA, USA), trypsin (#25200072; Thermo Fisher Scientific), HiScript II Q Select RT SuperMix (#R232-01; Vazyme, Nanjing, China) for reverse transcription, AceQ qPCR SYBR Green Master Mix (#Q111-02; Vazyme), a DLR™ assay system for luciferase assay (#E1910; Promega, Madison, WI, USA) and a Detection Kit for Estrodiol (E2) (#E9096-E2; North Biotech, Beijing, China). DMEM and RPMI 1640 medium were ordered from Gibco (#C11330500BT; Carlsbad, CA, USA) and Hyclone (#SH30027.01; Logan, UT, USA). The anti-FoxO1 (1:1,000; #D194961) and anti-IgG (1:1,000; #D111024) antibodies were purchased from Sangon (Shanghai, China). The KGN cell line was obtained from Yili (#DCX-1125; Shanghai, China).

Sows

In total, 200 fresh ovaries were isolated from 100 newly slaughtered commercial sows (180 days old, weighing approximately 110 kg) that were healthy and sexually mature (Zhushun, Nanjing, China). The ear tissue blocks were collected from 310 healthy Yorkshire sows with fertility records (Kangle, Jintan, China).

Bioinformatics

The promoter sequences were downloaded from the NCBI database (https://www.ncbi.nlm.nih.gov/), and the TF-binding sites (TFBSs) were predicted using the JASPAR database (http://jaspar.genereg.net/). DAVID bioinformatics (https://david.ncifcrf.gov/) was used for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Data on tissue expression and quantitative trait loci (QTLs) were obtained from the NCBI and Animal QTLdb databases (https://www.animalgenome.org/), respectively. DNA–RNA triplexes in the FoxO1 promoter were detected using the Gaemons software (http://www.gaemons.net/LongTarget).

Constructs and small interfering RNAs (siRNAs)

Overexpression plasmids for NORSF (non-coding RNA involved in sow fertility) and FoxO1 have been previously constructed [13, 17]. Promoters harboring wild-type or mutated versions of the TFBSs were ligated into the pGL3-basic vector (#E1751, Promega), and the primers are given in Table S1. FoxO1- and NORSF-specific siRNAs (Table S2) were previously validated by our group [13, 17], and synthesized with Generay (Shanghai, China).

Cells and treatment

Fresh ovaries were cleaned three times with saline at 37 °C and 75% alcohol supplemented with gentamicin. Antral follicles of 3–5 mm were isolated, and the follicular fluid was suctioned using a UV-sterilized syringe and centrifuged at 1,000 r/min for 5 min. After discarding the supernatant, the cells were resuspended in PBS and centrifuged to collect sGCs. sGCs were cultured in DMEM with 15% FBS. KGN cells were cultured in RPMI 1640 with 15% FBS. After culture for 12 h, constructs and siRNAs were transfected with Lipofectamine 2000 according to the manufacturer’s instructions. After transfection for 12 h, cells were resuspended in DMEM without FBS, and treated with H2O2 at a final concentration of 150 µmol/L and for 2 h.

Reporter assays

KGN cells were co-treated with expression plasmids, reporter constructs, and pRL-TK for 24 h, and then collected for lysis and measurement. Luciferase activity for each sample was obtained using an enzyme-labeling instrument (Thermo Fisher Scientific) and normalized to the pRL-TK levels.

Chromatin immunoprecipitation (ChIP)

ChIP was conducted as described previously [10]. After crosslinking in formaldehyde, precipitating chromatin by centrifugation, and sonication, the samples were precleared with protein A/G agarose beads and incubated with anti-FoxO1 antibody. Primers (Table S3) and qPCR were used to amplify the input and bound DNA. The negative control was DNA enriched with the IgG antibody, and the input control was unprocessed DNA.

Quantitative real-time PCR (qPCR)

sGCs were collected 24 h post-transfection to extract total RNA using TRIzol reagent. Reverse transcription and qPCR were conducted with the corresponding kits and procedures [10]. The 2−ΔΔCT method was employed to calculate the transcription level, with the GAPDH and RPLP0 serving as the reference controls. The primers are given in Table S4.

Fluorescence-activated cell sorting (FACS)

After washing twice with pre-cooled PBS, sGCs were dyed with 5 µL of each Annexin V and Propidium Iodide (PI) for 10 min. The sGCs were then diluted with Binding Buffer and sorted by flow cytometry (Becton Dickinson, Franklin Lakes, NJ, USA). The sGCs count in each quadrant was read using FlowJo v7.6 software (TreeStar, Ashland, OR, USA).

SNV screening, genotyping, and association analysis

Identification of single nucleotide variations (SNVs) and genotyping were conducted using DNA sequencing. SAS v9.2 software (SAS Institute, Cary, NC, USA) was employed for association analysis with linear models in literature [18]. Fertility traits included the total number of piglets born (TNB), number of piglets born alive (NBA), number of healthy piglets (NHP), number of stillbirths (NSB), and litter weight (LW). The primers are given in Table S5.

Statistics

Two-tailed t-test and one-way analysis of variance in Prism v9.0 software (GraphPad, San Diego, CA, USA) were used to evaluate the data. Each experiment was independently conducted in triplicates. The level of significance was set at P < 0.05.

Results

OS stimulates the lncRNome of sGCs in a FoxO1-dependent manner

To understand whether FoxO1 participates as a TF in the OS-stimulated lncRNome dynamics of sGCs, we predicted FRE motifs in the promoters of 432 OS-stimulated DElncRNAs [12]. In total, 2,283 putative FRE motifs were identified, and 394 DElncRNAs were detected with at least one FRE motif, accounting for 91.20% (394/432) of the DElncRNAs stimulated by OS (Fig. 1A and Table S6). GO and KEGG analyses showed that 909 potential cis-target mRNAs of the 394 DElncRNAs were markedly enriched in pyruvate metabolism, fatty acid degradation, and regulation of actin cytoskeleton (Fig. 1B, S1, Tables S7 and S8). Additionally, 101 potential cis-target mRNAs were differentially expressed in OS-stimulated sGCs, with 24.62% (97/394) of DElncRNAs having at least one cis-target mRNAs regulated by OS (Fig. 1C and Table S9). Together, these data suggest that OS stimulates the lncRNome of sGCs, which may be related to the TF activity of its effector FoxO1.

Fig. 1
figure 1

OS stimulates lncRNome in a FoxO1-dependent manner. A Diagram depicting the putative FRE motifs in the promoters of DElncRNAs stimulated by OS. DElncRNAs were obtained from our previous study [12]. B KEGG analysis of cis-target mRNAs of DElncRNAs was conducted with DAVID bioinformatics (https://david.ncifcrf.gov/). C Diagram depicting cis-target mRNAs that are differentially expressed in OS-stimulated sGCs. Data for mRNAs in OS-stimulated sGCs were obtained from our previous report [7]. D Two DElncRNAs in both OS-stimulated sGCs and follicular atresia process. DElncRNAs in the follicular atresia process were obtained from our previous study [17]. E Diagram depicting the FRE motifs in the promoters of 2 DElncRNAs. F ChIP assays. ChIP-PCR was conducted with an anti-FoxO1 antibody in sGCs, and detected with agarose gel electrophoresis. G–I Reporter assays. The reporter constructs of promoter harboring FRE motifs (G) were transfected into sGCs, luciferase activities were measured (H), or re-treated with H2O2 and FoxO1-siRNA, luciferase activities were measured (I). Quantitative data are plotted as mean ± standard error. *P < 0.05. **P < 0.01

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OS and its effector FoxO1 have been shown to be related to the sow follicular atresia [10, 13]. Therefore, we conducted a joint analysis of the lncRNAome of sGCs stimulated by OS and the lncRNAome of sow follicular atresia [17] and found that of 19 DElncRNAs during follicular atresia, 2 (LOC100512907 and LOC102164325) were stimulated by OS (Fig. 1D). Interestingly, the promoters of these 2 DElncRNAs contained 2 FRE motifs each (Fig. 1E). Next, we investigated whether FoxO1 mediated the regulation of target lncRNAs in sGCs in response to OS by directly interacting with their promoters. As expected, FoxO1 physically interacted with the promoters of these 2 DElncRNAs, and this interaction was stimulated by OS (Fig. 1F). Furthermore, reporter assays showed that FoxO1 increased the promoter activities of these 2 DElncRNAs (Fig. 1G–H). Additionally, depleting FoxO1 restrained the increase in the promoter activities of these two DElncRNAs stimulated by OS (Fig. 1I), indicating that FoxO1 can mediate regulation of the promoter activities of these 2 DElncRNAs in response to OS. Taken together, our results support that OS controls the transcriptional activity of lncRNAs in sGCs in a FoxO1-dependent manner.

FoxO1 mediates OS induction of NORSF transcription and cell apoptosis

We noticed that LOC102164325 (also known as NORSF) is intensely involved in sow follicular atresia [17]. Thus, NORSF was selected as the target to analyze FoxO1 mediated role of OS in subsequent studies. As expected, a marked increase in NORSF transcription levels was observed in sGCs under OS triggered by H2O2 exposure (Fig. 2A). Furthermore, depleting FoxO1 in sGCs resulted in a remarkable decrease in NORSF transcription levels, whereas OS triggered by H2O2 exposure reversed this process (Fig. 2B). The above results suggest that the effector FoxO1 mediates the OS induction of lncRNA NORSF transcription in sGCs.

Fig. 2
figure 2

FoxO1 mediates OS induction of NORSF transcription and its role in sGC apoptosis. A NORSF transcription levels were determined in sGCs treated with H2O2. B NORSF transcription levels were determined in sGCs co-treated with H2O2 and FoxO1-siRNA. C Apoptosis rate was determined in sGCs co-treated with H2O2, FoxO1-siRNA, and pcDNA3.1-NORSF. D and E FoxO1-siRNA and pcDNA3.1-NORSF (D) or pcDNA3.1-FoxO1 and NORSF-siRNA (E) were co-transfected into sGCs, and the apoptosis rate was determined. Quantitative data are plotted as mean ± standard error. *P < 0.05. **P < 0.01. ***P < 0.001. ns, no significance

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OS is an essential factor for inducing apoptosis in sGCs [10, 13]. We first confirmed that OS triggered by H2O2 exposure induced sGC apoptosis (Fig. 2C). However, this process was reversed by depleting FoxO1 (Fig. 2C), confirming that OS induces apoptosis via its effector FoxO1 in sGCs. Similarly, depleting FoxO1 in sGCs resulted in a remarkable decrease in the apoptosis rate (Fig. 2E). Simultaneously, overexpression of NORSF partially increased the rate of apoptosis (Fig. 2D). The same results were observed for OS triggered by H2O2 exposure (Fig. 2C). Additionally, depleting NORSF transcript restrained the decrease in the apoptosis rate caused by the overexpression of FoxO1 (Fig. 2D). These data support the conclusion that OS and its effector, FoxO1, control sGC apoptosis through inducing NORSF transcription.

FoxO1 controls NORSF transcriptional activity via the FRE1 motif in its core promoter

We previously identified the NORSF core promoter [19]. Next, we predicted the potential TFBSs in this region and identified 149 TFBSs of 79 TFs with a relative score ≥ 0.90 (Fig. 3A; Table S10). These TFs were significantly enriched in endometrial cancer, acute myeloid leukemia, and FoxO signaling pathway (Fig. S2). Electronic tissue expression profiling showed that nine TFs, such as GATA4, FoxO1, and NFIX, were relatively abundant in sow ovarian tissue (Fig. 3B; Table S11). Additionally, four TFs, including FoxO1, NFIX, TCF7L1, and FoxO3, were observed to be stimulated by OS triggered by H2O2 exposure (Fig. 3C) using our previous transcriptome data from OS-stimulated sGCs [7]. Combined with previous ChIP assays, these data suggest that FoxO1 is a critical TF for NORSF in sGCs.

Fig. 3
figure 3

FoxO1 controls NORSF transcription activity via interaction with its promoter. A Diagram depicting the potential TFBSs in the core promoter of the NORSF gene. The core promoter was identified with our group and located at -139/-1 nt (unpublished), and only TFs with relative scores ≥ 0.95 were listed. B Electronic tissue expression profiling of TFs in swine. Data for tissue expression were obtained from the NCBI database. C The expression of TFs in OS-stimulated sGCs. Data for mRNAs in OS-stimulated sGCs were obtained from our previous report [7]. D Diagram depicting the reporter constructs of the NORSF core promoter. E and F Reporter assay. The reporter constructs and pcDNA3.1-FoxO1 were co-transfected into sGCs, and luciferase activities were measured. Quantitative data are plotted as mean ± standard error. *P < 0.05. ***P < 0.001. ns, no significance

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We have previously confirmed that the TF FoxO1 directly interacts with the promoter of NORSF gene to enhance its transcriptional activity (Fig. 1). Furthermore, 2 FRE motifs are located at -121/-117 nt (FRE1 motif) and -18/-11 nt (FRE2) (Fig. 3D). Therefore, we investigated whether FoxO1 induces NORSF transcriptional activity through specific motifs, and generated the reporter constructs of the NORSF core promoter harboring the FRE motifs (Fig. 3D). Compared to the empty vector, transfection of the FoxO1 overexpression vector gave a remarkable induction in the activity of NORSF core promoter (Fig. 3E). Furthermore, FoxO1 induced the activity of NORSF promoter via the FRE1 motif in the core promoter (Fig. 3F). Combined with previous ChIP assays, our data suggest that FoxO1 induces NORSF transcription in sGCs by physically interacting with the FRE1 motif of NORSF core promoter.

FoxO1 is a transcription activator of NORSF in sGCs

Next, we investigated the effect of FoxO1 on endogenous NORSF transcription in sGCs. Compared to the empty vector, transfection with the FoxO1 expression construct gave a remarkable increase in the levels of NORSF transcripts in sGCs (Fig. 4A). In contrast, silencing of FoxO1 gave a noteworthy reduction in the levels of NORSF transcripts in sGCs (Fig. 4B). Combined with the reporter assay and ChIP results, these data support that FoxO1 is a vital transcriptional activator of the lncRNA NORSF in sGCs.

Fig. 4
figure 4

FoxO1 induces NORSF transcription in sGCs. A and B NORSF transcript levels in FoxO1-overexpressing (A) or -depleting sGCs. C Diagram depicting the putative DNA–RNA triplex structures formed by NORSF transcripts and FoxO1 promoter. The DNA–RNA triplexes were predicted with a Gaemons. D and E FoxO1 transcription levels in NORSF-overexpressing or -depleting sGCs. Quantitative data are plotted as mean ± standard error. *P < 0.05. ns, no significance

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Our previous study showed that NORSF is a nuclear lncRNA in sGCs [17]. Nuclear lncRNAs usually control the transcription of targets by forming a DNA–RNA triplex with their promoters [20, 21]. Interestingly, 5 putative triplex target sites (TTSs) of the NORSF transcript were detected in the porcine FoxO1 promoter and five putative triplex-forming oligonucleotides (TFOs) were detected in the NORSF transcript (Fig. 4C), suggesting that nuclear NORSF may potentially bind to the FoxO1 promoter. However, we did not observe significant changes in FoxO1 mRNA levels in sGCs overexpressing NORSF transcript (Fig. 4D and E), indicating that NORSF has no feedback regulatory effect on the transcription activator FoxO1 transcription in sGCs.

The FoxO1 and NORSF axis mediates OS reduction of E2 release

NORSF is a vital modulator of E2 release from sGCs [17]. Next, we investigated whether the transcription activator FoxO1 regulates E2 release by sGCs. Compared to the cells transfected with the empty vector, transfection of the FoxO1 expression construct gave a remarkable reduction in E2 concentration in the culture medium of sGCs (Fig. 5A). In contrast, silencing of FoxO1 remarkably increased E2 concentration (Fig. 5B), indicating that FoxO1 restrains E2 release by sGCs. Furthermore, depleting the NORSF transcript restrained the decrease in E2 release caused by FoxO1 (Fig. 5A), while overexpression of the NORSF transcript restrained the increase in E2 release caused by depleting FoxO1 (Fig. 5B). These data suggest that NORSF mediates FoxO1 downregulation during E2 release from sGCs.

Fig. 5
figure 5

The FoxO1 and NORSF axis mediates OS reduction of E2 release by sGCs. A and B E2 concentration in the culture medium of sGCs co-transfecting with pcDNA3.1-FoxO1 and NORSF-siRNA (A) or FoxO1-siRNA and pcDNA3.1-NORSF (B). C and D CYP19A1 transcription levels in sGCs co-transfecting with pcDNA3.1-FoxO1 and NORSF-siRNA (C) or FoxO1-siRNA and pcDNA3.1-NORSF (D). E CYP19A1 transcription levels in sGCs co-treating with H2O2, FoxO1-siRNA, and pcDNA3.1-NORSF. Quantitative data are plotted as mean ± standard error. *P < 0.05. **P < 0.01. ***P < 0.001

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NORSF suppresses E2 release from sGCs by inhibiting the transcription of CYP19A1, a gene encoding an essential enzyme for estrogen synthesis [17]. Therefore investigated whether FoxO1 regulates CYP19A1 transcription in sGCs. Transfection of the FoxO1 expression construct gave a remarkable reduction in CYP19A1 transcription in sGCs compared to the cells transfected with the empty vector (Fig. 5C), whereas silencing of FoxO1 gave a remarkable induction in CYP19A1 transcription (Fig. 5D), indicating that FoxO1 reduces CYP19A1 transcription in sGCs. Furthermore, depleting the NORSF transcript restrained the decrease in CYP19A1 transcription levels caused by FoxO1 (Fig. 5C), while overexpression of the NORSF transcript restrained the increase in CYP19A1 transcription caused by depleting FoxO1 (Fig. 5D). These data suggest that NORSF and CYP19A1 axis mediates FoxO1 reduction of E2 release in sGCs.

Similarly, we also showed that, similar to its effector FoxO1, OS induces a decrease in CYP19A1 transcription, whereas inactivation of FoxO1 can reverse this process (Fig. 5E), indicating that OS restrains CYP19A1 transcription via FoxO1. Meanwhile, in OS triggered by H2O2 exposure, overexpression of NORSF restrained the increase in CYP19A1 transcription caused by depleting FoxO1 (Fig. 5E). These data suggest that OS restrains CYP19A1 transcription via the FoxO1 and NORSF axis.

NORSF is a candidate gene for sow fertility

Notably, 2 adjacent SNVs (C to T and G to A) were observed at -360/-359 nt in the promoter, not the core promoter, of Yorkshire sow NORSF gene (Fig. 6A). In the Yorkshire sow population, three genotypes were detected for both SNVs, with the genotype AA at SNV g.-360C > T being a rare genotype with a genotype frequency of less than 1% (2/310) (Fig. 6B–G). Furthermore, these 2 SNVs were low-to-moderate polymorphic loci, with PICs of 0.108 and 0.271, respectively (Table S12). However, neither of these SNVs was significantly associated with sow fertility traits, such as TNB, NBA, and LW (Fig. S3 and S4).

Fig. 6
figure 6

NORSF is a candidate gene for sow fertility traits. A Diagram depicting the 2 adjacent SNVs in the NORSF promoter. B–D Peak plot (B), genotype frequency (C), and allele frequency (D) of the SNV g.-360C > T. E–G Peak plot (E), genotype frequency (F), and allele frequency (G) of the SNV g.-359G > A. H and I The haplotype combinations frequency (H) and haplotype frequency (I) of the 2 adjacent SNVs. J–N Association analysis between the haplotype combinations and the TNB (J), NBA (K), NHP (L), NSB (M), and LW (N) traits. O Schematic showing the reporters of NORSF promoter with four haplotypes. P Reporter assay. The reporters were transfected into sGCs, and luciferase activities were measured. Quantitative data are plotted as the least squares mean ± standard error (J–N) or mean ± standard error (P). *P < 0.05. **P < 0.01

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Interestingly, 2 adjacent SNVs, g.-360C > T and g.-359G > A, exhibited linkage disequilibrium and formed four haplotypes: H1 (C-G), H2 (C-A), H3 (T-G), and H4 (T-A) in the Yorkshire sow population. Haplotypes H1 and H2 were major haplotypes with 74.84% and 19.03% frequency, whereas H4 was a rare genotype with a frequency of 1.29% (Fig. 6H–I). Furthermore, compared to sows with the haplotype combination H1H3, a remarkable increase in the average litter size of TNB, NBA, and NHP traits was found in sows with H1H2. The average litter size of the NHP trait was observed in sows with H1H1 (Fig. 6J–N), suggesting that NORSF is a candidate gene for sow fertility traits, and that H3 is an unfavorable haplotype of sow fertility.

Functional SNVs in the promoter primarily control the transcriptional activity of targets [10, 22]. To understand the impact of g.-360C > T and g.-359G > A on transcriptional activity, reporters of the NORSF promoter with the four haplotypes were generated (Fig. 6O). A reporter assay showed that these 2 SNVs significantly altered the activity of the NORSF promoter, with lower haplotype H2 activity and higher haplotype H3 activity, respectively (Fig. 6P). This is consistent with the function of NORSF in sGCs and the fertility phenotype of haplotype combinations. These data support that NORSF is a functional candidate gene for sow fertility traits, because as its 2 SNVs synergistically influence transcriptional activity and sow fertility.

Discussion

FoxO1 is a component of the forkhead box O (FoxO) subfamily of TFs that are known to respond to many biological stressors, including OS, and play crucial roles in both physiological and disease processes [23,24,25]. FoxO1 serves as an effector in response to OS, with an increased activity observed in OS-stimulated cells such as sGCs [7], MCF7 cells [26], and KGN cells [27]. On the other hand, FoxO1 also acts as a TF to transmit OS signals to the nucleus, activate or inhibit the transcription of downstream genes, and mediate the regulation of cellular functions in response to OS [28, 29]. However, the extent to which the TF activity of the effector FoxO1 plays a role in OS-stimulated transcription is unknown. In this study, using lncRNAs in sGCs as an example, we showed that more than 90% of OS-stimulated lncRNAs may be transcriptionally regulated by their effector and TF FoxO1. We demonstrated that FoxO1 mediates the stimulation of the transcription of lncRNAs (e.g., LOC100512907 and NORSF) in response to OS. In addition, we also noticed that multiple cis-target mRNAs of OS-stimulated lncRNAs have been proven to be modulated by OS [29, 30]. Interestingly, FoxO1 was the cis-target mRNA of OS-stimulated LOC102160118 in Sus scrofa Chr.3. In contrast, as an another cis-target mRNA of LOC102160118, SLC25A15 is an OS- and metabolism-related gene serves as a biomarker for predicting adverse events in patients with stomach adenocarcinoma [31]. 6-Pyruvoyltetrahydropterin synthase (PTS) and beta-carotene oxygenase 2 (BCO2), 2 cis-target mRNAs of NORSF, are involved in OS [30, 32]. Taken together, our findings suggest that the stimulation of lncRNome dynamics in response to OS depends on the transcription activity of the effector FoxO1.

OS and its effector FoxO1 are crucial endogenous environmental factor and TF that regulate cell state, function, metabolism, and structure [15, 33]. In the ovary, OS and its effector, FoxO1, have regulatory effects on multiple cell types, such as GC autophagic injuries, mitochondrial impairment, and apoptosis in humans, impaired viability, reduced proliferation, and cell cycle arrest in mice [34]. In addition, OS and FoxO1 also affect oocyte maturation and quality in mice, meiotic arrest in sows [35, 36], and luteal cell apoptosis and luteolysis in rats [37], and are involved in ovarian aging and diseases [35, 38]. In sGCs, OS and its effector FoxO1 are also modulators of cell viability, apoptosis, and autophagy [11, 12, 39]. OS induced by various concentrations (e.g., 100, 150, 200, 300, 500, and 1,000 µmol/L) of H2O2 and exposure times (e.g., 1.5, 2.0, 6.0, and 12.0 h) can induce sGC apoptosis and oxidative damage, whereas antioxidant factors can rescue sGC apoptosis [10,11,12,13, 40]. Furthermore, FoxO1 levels were increased in OS-stimulated sGCs and FoxO1 dysfunction rescued OS-induced sGC apoptosis, indicating that FoxO1 mediates OS-induced sGC apoptosis [13, 40]. In the present study, we demonstrated that FoxO1 mediated OS-induced sGC apoptosis. More importantly, we showed that OS restrained E2 release, a primary function of sGCs, and that FoxO1 mediated this process via NORSF and the estrogen synthesis pathway. Thus, our findings not only define a new function of OS and its effector FoxO1 in sGCs, but also reveal a new mechanism by which OS controls sGC functions and provides a new pathway for regulating sGC functions, the FoxO1/NORSF/CYP19A1 pathway.

OS affects sow fertility by interacting with transcripts related to sow fertility [10, 41, 42]. First, multiple OS-stimulated transcripts, including mRNAs (e.g., FZD4 and Acvr2a) and miRNAs (e.g., miR-192 and miR-370), are closely related to follicular atresia, which is detrimental for sow fertility [7, 11, 18, 42]. Interestingly, recent studies have demonstrated that several OS-stimulated lncRNAs such as lnc2300 and SDNOR contribute to sGC function [12, 43]. In sows, downregulated lnc2300 in atretic follicles inhibits sGC apoptosis by serving as a competing endogenous RNA of miR-365 and a cis-acting lncRNA of CYP11A1 [43]. SDNOR, a SMAD4-dependent lncRNA, contributes to multiple sGC functions such as cell proliferation, viability, cell cycle, and apoptosis [12, 41]. Importantly, SDNOR also acts as an antioxidant that inhibits the induction of sGC apoptosis via OS [12]. In addition, lncRNA NORHA and OS can synergistically induce apoptosis by co-targeting the miR-183-96-182 cluster in sGCs [13].

Second, multiple OS-stimulated transcripts were discovered to be localized within QTLs for sow fertility traits, such as VEGFA (QTL ID: 584), miR-182 (QTL ID: 24290), and lncRNA LOC100512907 (QTL ID: 7536), which are potential candidate genes for sow fertility traits. Of them, some transcripts such as TGF-β1, PRLR, and miR-27a have been demonstrated to be candidate genes for sow fertility traits [44, 45]. Several transcripts have been confirmed as the causal genes of sow fertility traits. For example, miR-23a, an miRNA located within a QTL (ID: 909) for sow fertility traits, has been identified as a causal gene for the TNB trait, because a point mutation in its promoter reduced the sGCs response to OS [10]. NORSF, a lncRNA activated by OS, increases during sow follicular atresia, inducing sGC apoptosis and restraining E2 release and is located in two QTLs (ID: 517 and 7462) for sow fertility traits [13, 17]. In this study, we further demonstrated that NORSF induced by OS is a causal gene for Yorkshire sow fertility traits (e.g., TNB, NBA, and NHP), as 2 novel SNVs in its promoter lead to transcriptional dysregulation and decreased sow fertility. However, further investigation is required to elucidate the mechanism underlying NORSF transcriptional dysregulation caused by these variants. Together, our findings support the conclusion that OS is deeply involved in fertility and provides a target for anti-OS therapies in sow reproductive regulation and a genetic marker (2 causal variants) for molecular breeding in swine.

Conclusions

In summary, we provided evidence that OS stimulates the lncRNome dynamics of sGCs in an effector FoxO1-dependent manner. FoxO1 acts as a TF to interact with the FRE motifs of target promoters. Furthermore, FoxO1 mediated OS-stimulated sGC apoptosis and E2 release via the FoxO1/NORSF/CYP19A1 pathway (Fig. S5). Additionally, NORSF induced by OS is a cause gene for Yorkshire sow fertility traits. Overall, our findings expand on the new functions of OS in female reproduction and elucidate the mechanism by which it regulates the transcription of downstream target, providing a new target for improving sow fertility through an anti-OS strategy.

Abbreviations

ChIP:

Chromatin immunoprecipitation

DElncRNA:

Differentially expressed lncRNA

E2:

Estrodiol

FRE:

FoxO1 response element

GEI:

Gene-environment interactions

LW:

Litter weight

miRNA:

MicroRNA

lncRNA:

Long non-coding RNA

NBA:

Number of piglets born alive

NSB:

Number of stillbirths

OS:

Oxidative stress

sGC:

Sow granulosa cell

PI:

Propidium iodide

qPCR:

Quantitative real-time PCR

QTLs:

Quantitative trait loci

SNV:

Single nucleotide variation

TFBS:

Transcription factor binding site

TNB:

Total number of piglets born

TF:

Transcription factor

References

  1. Zhao Y, Flowers WL, Saraiva A, Yeum KJ, Kim SW. Effect of social ranks and gestation housing systems on oxidative stress status, reproductive performance, and immune status of sows. J Anim Sci. 2013;91(12):5848–58. https://doi.org/10.2527/jas.2013-6388.

    Article  PubMed  CAS  Google Scholar 

  2. Papatsiros VG, Katsogiannou EG, Papakonstantinou GI, Michel A, Petrotos K, Athanasiou LV. Effects of phenolic phytogenic feed additives on certain oxidative damage biomarkers and the performance of primiparous sows exposed to heat stress under field conditions. Antioxidants (Basel). 2022;11(3):593. https://doi.org/10.3390/antiox11030593.

    Article  PubMed  CAS  Google Scholar 

  3. Huang S, Zhang L, Luo J, Wu D, Ma K, Chen Y, et al. Cysteamine and N-acetyl-cysteine alleviate placental oxidative stress and barrier function damage induced by deoxynivalenol. J Agric Food Chem. 2023;71(18):6846–58. https://doi.org/10.1021/acs.jafc.3c00399.

    Article  PubMed  CAS  Google Scholar 

  4. Huang S, Wu Z, Huang Z, Hao X, Zhang L, Hu C, et al. Maternal supply of cysteamine alleviates oxidative stress and enhances angiogenesis in porcine placenta. J Anim Sci Biotechnol. 2021;12:91. https://doi.org/10.1186/s40104-021-00609-8.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  5. Zhang P, Jiang G, Wang Y, Yan E, He L, Guo J, et al. Maternal consumption of L-malic acid enriched diets improves antioxidant capacity and glucose metabolism in offspring by regulating the gut microbiota. Redox Biol. 2023;67:102889. https://doi.org/10.1016/j.redox.2023.102889.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  6. Tatemoto H, Muto N, Sunagawa I, Shinjo A, Nakada T. Protection of porcine oocytes against cell damage caused by oxidative stress during in vitro maturation: role of superoxide dismutase activity in porcine follicular fluid. Biol Reprod. 2004;71(4):1150–7. https://doi.org/10.1095/biolreprod.104.029264.

    Article  PubMed  CAS  Google Scholar 

  7. Du X, Li Q, Cao Q, Wang S, Liu H, Li Q. Integrated analysis of miRNA-mRNA interaction network in porcine granulosa cells undergoing oxidative stress. Oxid Med Cell Longev. 2019;2019:1041583. https://doi.org/10.1155/2019/1041583.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  8. Zhang X, Chen Y, Li H, Chen B, Liu Z, Wu G, et al. Sulforaphane acts through NFR2L2 to prevent hypoxia-induced apoptosis in porcine granulosa cells via activating antioxidant defenses and mitophagy. J Agric Food Chem. 2022;70(26):8097–110. https://doi.org/10.1021/acs.jafc.2c01978.

    Article  PubMed  CAS  Google Scholar 

  9. Mierzejewski K, Kurzynska A, Gerwel Z, Golubska M, Stryinski R, Bogacka I. PPARβ/δ ligands regulate oxidative status and inflammatory response in inflamed corpus luteum-an in vitro study. Int J Mol Sci. 2023;24(5):4993. https://doi.org/10.3390/ijms24054993.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  10. Wang S, Li Y, Zeng Q, Yang L, Du X, Li Q. A mutation in endogenous saRNA miR-23a influences granulosa cells response to oxidative stress. Antioxidants (Basel). 2022;11(6):1174. https://doi.org/10.3390/antiox11061174.

    Article  CAS  Google Scholar 

  11. Zhang J, Ren Q, Chen J, Lv L, Wang J, Shen M, et al. Downregulation of miR-192 alleviates oxidative stress-induced porcine granulosa cell injury by directly targeting ACVR2A. Cells. 2022;11(15):2362. https://doi.org/10.3390/cells11152362.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  12. Huo Y, Li Q, Yang L, Li X, Sun C, Liu Y, et al. SDNOR, a novel antioxidative lncRNA, is essential for maintaining the normal state and function of porcine follicular granulosa cells. Antioxidants (Basel). 2023;12(4):799. https://doi.org/10.3390/antiox12040799.

    Article  CAS  Google Scholar 

  13. Yao W, Pan Z, Du X, Zhang J, Liu H, Li Q. NORHA, a novel follicular atresia-related lncRNA, promotes porcine granulosa cell apoptosis via the miR-183-96-182 cluster and FoxO1 axis. J Anim Sci Biotechnol. 2021;12:103. https://doi.org/10.1186/s40104-021-00626-7.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  14. Zhang JQ, Wang XW, Chen JF, Ren QL, Wang J, Gao BW, et al. Grape seed procyanidin B2 protects porcine ovarian granulosa cells against oxidative stress-induced apoptosis by upregulating let-7a expression. Oxid Med Cell Longev. 2019;2019:1076512. https://doi.org/10.1155/2019/1076512.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  15. Rodriguez-Colman MJ, Dansen TB, Burgering B. FoxO transcription factors as mediators of stress adaptation. Nat Rev Mol Cell Biol. 2024;25(1):46–64. https://doi.org/10.1038/s41580-023-00649-0.

    Article  PubMed  CAS  Google Scholar 

  16. Zhang X, Evans TD, Chen S, Sergin I, Stitham J, Jeong SJ, et al. Loss of macrophage mTORC2 drives atherosclerosis via FoxO1 and IL-1β signaling. Circ Res. 2023;133(3):200–19. https://doi.org/10.1161/CIRCRESAHA.122.321542.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  17. Wang M, Wang Y, Yang L, Du X, Li Q. Nuclear lncRNA NORSF reduces E2 release in granulosa cells by sponging the endogenous small activating RNA miR-339. BMC Biol. 2023;21:221. https://doi.org/10.1186/s12915-023-01731-x.

  18. Du X, Liu L, Wu W, Li P, Pan Z, Zhang L, et al. SMARCA2 is regulated by NORFA-miR-29c, a novel pathway that controls granulosa cell apoptosis and is related to female fertility. J Cell Sci. 2020;133(23):jcs249961. https://doi.org/10.1242/jcs.249961.

  19. Wang M, Zhang J, Sheng W, Wu W, Du X, Li Q. A variant in long noncoding RNA NORSF affects granulosa cells response to transcription factor RFX7. J Cell Physiol. 2024;239:e31414. Online Version. https://doi.org/10.1002/jcp.31414.

  20. Zhang X, Jiang Q, Li J, Zhang S, Cao Y, Xia X, et al. KCNQ1OT1 promotes genome-wide transposon repression by guiding RNA-DNA triplexes and HP1 binding. Nat Cell Biol. 2022;24(11):1617–29. https://doi.org/10.1038/s41556-022-01008-5.

    Article  PubMed  CAS  Google Scholar 

  21. Ali T, Rogala S, Krause NM, Bains JK, Melissari MT, Wahrisch S, et al. Fendrr synergizes with Wnt signalling to regulate fibrosis related genes during lung development via its RNA:dsDNA triplex element. Nucleic Acids Res. 2023;51(12):6227–37. https://doi.org/10.1093/nar/gkad395.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  22. Miao Y, Zhao Y, Wan S, Mei Q, Wang H, Fu C, et al. Integrated analysis of genome-wide association studies and 3D epigenomic characteristics reveal the BMP2 gene regulating loin muscle depth in Yorkshire pigs. PLoS Genet. 2023;19(6):e1010820. https://doi.org/10.1371/journal.pgen.1010820.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  23. Chan JD, Scheffler CM, Munoz I, Sek K, Lee JN, Huang YK, et al. FoxO1 enhances CAR T cell stemness, metabolic fitness and efficacy. Nature. 2024;629(8010):201–10. https://doi.org/10.1038/s41586-024-07242-1.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  24. Doan AE, Mueller KP, Chen AY, Rouin GT, Chen Y, Daniel B, et al. FoxO1 is a master regulator of memory programming in CAR T cells. Nature. 2024;629(8010):211–8. https://doi.org/10.1038/s41586-024-07300-8.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  25. Zhang X, Ge L, Jin G, Liu Y, Yu Q, Chen W, et al. Cold-induced FoxO1 nuclear transport aids cold survival and tissue storage. Nat Commun. 2024;15:2859. https://doi.org/10.1038/s41467-024-47095-w.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  26. Jose E, March-Steinman W, Wilson BA, Shanks L, Parkinson C, Alvarado-Cruz I, et al. Temporal coordination of the transcription factor response to H2O2stress. Nat Commun. 2024;15:3440. https://doi.org/10.1038/s41467-024-47095-w.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  27. Zhang M, Zhang Q, Hu Y, Xu L, Jiang Y, Zhang C, et al. miR-181a increases FoxO1 acetylation and promotes granulosa cell apoptosis via SIRT1 downregulation. Cell Death Dis. 2017;8(10):e3088. https://doi.org/10.1038/cddis.2017.467.

    Article  PubMed  PubMed Central  Google Scholar 

  28. Li N, Liu B, Xiong R, Li G, Wang B, Geng Q. HDAC3 deficiency protects against acute lung injury by maintaining epithelial barrier integrity through preserving mitochondrial quality control. Redox Biol. 2023;63:102746. https://doi.org/10.1016/j.redox.2023.102746.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  29. Chen Z, Wang R, Zhu Y, Huang Z, Yang X, Li Q, et al. A novel circular RNA, circSQTM1, protects the endothelial function in atherosclerosis. Free Radic Biol Med. 2023;209(Pt 2):301–19. https://doi.org/10.1016/j.freeradbiomed.2023.10.398.

    Article  PubMed  CAS  Google Scholar 

  30. Wu L, Lu P, Guo X, Song K, Lyu Y, Bothwell J, et al. β-Carotene oxygenase 2 deficiency-triggered mitochondrial oxidative stress promotes low-grade inflammation and metabolic dysfunction. Free Radic Biol Med. 2021;164:271–84. https://doi.org/10.1016/j.jnutbio.2020.108542.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  31. Dong Y, Yuan Q, Ren J, Li H, Guo H, Guan H, et al. Identification and characterization of a novel molecular classification incorporating oxidative stress and metabolism-related genes for stomach adenocarcinoma in the framework of predictive, preventive, and personalized medicine. Front Endocrinol (Lausanne). 2023;14:1090906. https://doi.org/10.3389/fendo.2023.1090906.

    Article  PubMed  Google Scholar 

  32. Hara S, Fukumura S, Ichinose H. Reversible S-glutathionylation of human 6-pyruvoyl tetrahydropterin synthase protects its enzymatic activity. J Biol Chem. 2019;294(4):1420–7. https://doi.org/10.1074/jbc.RA118.005280.

    Article  PubMed  CAS  Google Scholar 

  33. Howes AC, Perisic O, Williams RL. Structural insights into the activation of ataxia-telangiectasia mutated by oxidative stress. Sci Adv. 2023;9(39):eadi8291. https://doi.org/10.1126/sciadv.adi8291.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  34. Mu H, Cai S, Wang X, Li H, Zhang L, Li H, et al. RNA binding protein IGF2BP1 meditates oxidative stress-induced granulosa cell dysfunction by regulating MDM2 mRNA stability in an m6A-dependent manner. Redox Biol. 2022;57:102492. https://doi.org/10.1016/j.redox.2022.102492.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  35. Zhang Y, Bai J, Cui Z, Li Y, Gao Q, Miao Y, et al. Polyamine metabolite spermidine rejuvenates oocyte quality by enhancing mitophagy during female reproductive aging. Nat Aging. 2023;3(11):1372–86. https://doi.org/10.1038/s43587-023-00498-8.

    Article  PubMed  CAS  Google Scholar 

  36. Li X, Duan J, Wang S, Cheng J, Chen H, Zhang Z, et al. Isorhamnetin protects porcine oocytes from zearalenone-induced reproductive toxicity through the PI3K/Akt signaling pathway. J Anim Sci Biotechnol. 2023;14:22. https://doi.org/10.1186/s40104-022-00809-w.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  37. Tang Z, Chen J, Zhang Z, Bi J, Xu R, Lin Q, et al. HIF-1α activation promotes luteolysis by enhancing ROS levels in the corpus luteum of pseudopregnant rats. Oxid Med Cell Longev. 2021;2021:1764929. https://doi.org/10.1155/2021/1764929.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  38. Srnovrsnik T, Virant-Klun I, Pinter B. Heavy metals and essential elements in association with oxidative stress in women with polycystic ovary syndrome-a systematic review. Antioxidants (Basel). 2023;12(7):1398. https://doi.org/10.3390/antiox12071398.

    Article  CAS  Google Scholar 

  39. Zhang JQ, Ren QL, Chen JF, Gao BW, Wang XW, Zhang ZJ, et al. Autophagy contributes to oxidative stress-induced apoptosis in porcine granulosa cells. Reprod Sci. 2021;28(8):2147–60. https://doi.org/10.1007/s43032-020-00340-1.

    Article  PubMed  CAS  Google Scholar 

  40. Kong C, Su J, Wang Q, Liu K, Fu R, Sui S. Signaling pathways of Periplaneta americana peptide resist H2O2-induced apoptosis in pig-ovary granulosa cells through FoxO1. Theriogenology. 2022;183:108–19. https://doi.org/10.1016/j.theriogenology.2022.02.004.

    Article  PubMed  CAS  Google Scholar 

  41. Du X, Li Q, Yang L, Liu L, Cao Q, Li Q. SMAD4 activates wnt signaling pathway to inhibit granulosa cell apoptosis. Cell Death Dis. 2020;11(5):373. https://doi.org/10.1038/s41419-020-2578-x.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  42. Zhou X, He Y, Pan X, Quan H, He B, Li Y, et al. DNMT1-mediated lncRNA IFFD controls the follicular development via targeting GLI1 by sponging miR-370. Cell Death Differ. 2023;30(2):576–88. https://doi.org/10.1038/s41418-022-01103-y.

    Article  PubMed  CAS  Google Scholar 

  43. Wang M, Wang Y, Yao W, Du X, Li Q. Lnc2300 is a cis-acting long noncoding RNA of CYP11A1 in ovarian granulosa cells. J Cell Physiol. 2022;237(11):4238–50. https://doi.org/10.1002/jcp.30872.

    Article  PubMed  CAS  Google Scholar 

  44. Pang P, Li Z, Hu H, Wang L, Sun H, Mei S, et al. Genetic effect and combined genotype effect of ESR, FSH β, CTNNAL1 and miR-27a loci on litter size in a large white population. Anim Biotechnol. 2019;30(4):287–92. https://doi.org/10.1080/10495398.2018.1486322.

    Article  PubMed  CAS  Google Scholar 

  45. Wang L, Du X, Li Q, Wu W, Pan Z, Li Q. miR-2337 induces TGF-β1 production in granulosa cells by acting as an endogenous small activating RNA. Cell Death Discov. 2021;7(1):253. https://doi.org/10.1038/s41420-021-00644-4.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

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Funding

This work was supported by the National Key R&D Program of China (2022YFD1600903) and the National Natural Science Foundation of China (32272842).

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Authors and Affiliations

  1. College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, 210095, China

    Wenmin Sheng, Miaomiao Wang, Yuqi Li, Zhenyu Sun, Xing Du & Qifa Li

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  1. Wenmin Sheng
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  2. Miaomiao Wang
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  3. Yuqi Li
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Contributions

Conception and design: QL. Collection and assembly of data: WS and MW. Data analysis and interpretation: WS, YL, ZS and XD. Manuscript writing: WS and QL. Final approval of the manuscript: all authors.

Corresponding author

Correspondence to Qifa Li.

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Ethics approval and consent to participate

All sow experiments were approved by the Animal Ethics Committee at Nanjing Agriculture University, China.

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The authors declare no conflict of interest.

Supplementary Information

Additional file 1: Table S1

Primers designed for reporter vector construction. Table S2 Oligonucleotide sequences. Table S3 Primers designed for ChIP. Table S4 Primers designed for qPCR. Table S5 Primers designed for genotyping. Table S6 The putative FRE motifs in the promoters of the OS-stimulated DElncRNAs. Table S7 Cis-target mRNAs of OS-stimulated DElncRNAs with the promoters containing FRE motifs. Table S8 GO terms and KEGG pathway analysis of cis-target mRNAs. Table S9 OS-stimulated cis-target mRNAs. Table S10 The putative TFBSs in the NORSF core promoter. Table S11 Tissue expression of the putative TFs in the NORSF core promoter. Table S12 The polymorphism of 2 SNVs in NORSF promoter.

Additional file 2: Fig. S1

GO analysis of cis-target mRNAs of OS-stimulated DElncRNAs. Fig. S2 GO terms and KEGG pathway analysis of the putative TFs in the NORSF core promoter. Fig. S3 Association analysis between SNV g.-360C > T and sow fertility traits. Fig. S4 Association analysis between SNV g.-359G > A and sow fertility traits. Fig. S5 Working model.

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Sheng, W., Wang, M., Li, Y. et al. Oxidative stress controls lncRNA-mediated sow granulosa cell functions in a FoxO1-dependent manner. J Animal Sci Biotechnol 15, 171 (2024). https://doi.org/10.1186/s40104-024-01120-6

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Journal of Animal Science and Biotechnology

ISSN: 2049-1891