Methotrexate affects brain development by regulating serum-glucocorticoid inducible protein kinase 3 (SGK3) signal pathway in brain organoids.
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ABSTRACT:
PROVIDER: PRJNA1092099 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR28464500_1.fastq.gz | Fastqsanger.gz | |||
| SRR28464500_2.fastq.gz | Fastqsanger.gz | |||
| SRR28464501_1.fastq.gz | Fastqsanger.gz | |||
| SRR28464501_2.fastq.gz | Fastqsanger.gz | |||
| SRR28464502_1.fastq.gz | Fastqsanger.gz |
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Project description:Excessive glucocorticoid exposure is associated with significant metabolic side effects, partly attributed to the transcriptional regulation of adipogenesis by the Glucocorticoid receptor (GR). However, emerging evidence suggests the involvement of alternative pathways beyond GR activation, which are inadequately explored. Through quantitative phosphoproteomics analysis, we identified a significant enrichment of rapidly phosphorylated proteins harboring the RxxS motif following glucocorticoid exposure. Notably, a subset of these proteins undergoes phosphorylation at the RxxS motif by Serum- and glucocorticoid-induced kinase 3 (SGK3) upon glucocorticoid stimulation in preadipocytes. SGK3 emerges as a critical mediator of glucocorticoid-induced preadipocyte differentiation, supported by genetic ablation and pharmacological inhibition experiments that demonstrate impaired adipogenesis both in vitro and in vivo. Mechanistically, upon glucocorticoid stimulation, SGK3 translocates to the nucleus and directly interacts with the core ATPase subunit of the BAF chromatin remodeling complex, BRG1, leading to phosphorylation at threonine 428 and serine 1417 residues. This phosphorylation event prevents proteasomal degradation of BRG1, facilitating chromatin remodeling essential for adipogenesis. Furthermore, Sgk3 knockout mice exhibit resistance to obesity induced by either glucocorticoids or a high-fat diet (HFD). These findings illuminate novel roles of glucocorticoids beyond GR-mediated transcriptional control, highlighting SGK3 as a promising therapeutic target for mitigating metabolic side effects associated with glucocorticoid therapy.
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Project description:bulk sequencing outputs of Caco2 cells after exposure to permeability modifying and permeability rescuing agents. To identify the molecular drivers for methotrexate-induced barrier dysfunction, we conducted RNA sequencing on Caco2 spheroids treated with methotrexate and lactoferrin. Given our observation that barrier function was compromised as early as 4-6 hours after exposure to methotrexate, we isolated RNA from spheroids 4-hour post-treatment to capture the transcriptional events responsible for initiating the processes.
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Project description:Deregulation of the PI3K/Akt pathway is a common feature in many types of cancer. Recent data has demonstrated that prolonged inhibition of breast cancer cell lines with PI3K and Akt inhibitors leads to up-regulation and increased activity of the AGC kinase Serum and glucocorticoid activated kinase 3, SGK3, which in turn substitutes for Akt and leads to activation of mTORC1. SGK3 and Akt exhibit similar substrate specificity for Ser/Thr residues in the RxRxxS/T motif. However, SGK3 can be regulated by both PI3K class I and class III and possesses a unique PX domain, enabling it to bind to PI(3)P lipids in endosomes. Still, the known SGK3 substrates are either shared or have not been tested whether Akt can phosphorylate them. In the present study, we undertook genetic and pharmacologic phosphoproteomic screens to search for selective SGK3 substrates. We identified a significant number of potential in vivo SGK3 selective substrates, including the endosomal proteins STX7, STX12, RFIP4 and WDR44. In vitro phosphorylation studies demonstrated that these substrates were directly phosphorylated at the sites identified in vivo by SGK3 but, unlike all other SGK3 substrates reported, not by Akt. We provide evidence that endogenous STX7 and STX12 are physiological SGK3 substrates whose phosphorylation is potently stimulated by IGF1. Our data pinpoint that STX7 and STX12 are the first selective SGK3 substrates in vivo and this phosphorylation enhances their interaction with the respective SNARE complexes. Finally, we suggest that STX7 and STX12 phosphorylation can be utilised as in vivo biomarkers for SGK3 activity.
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