Optimization of multi-enzyme cascade process for the biosynthesis of benzylamine.

Benzylamine is a valuable intermediate in the synthesis of organic compounds such as curing agents and antifungal drugs. To improve the efficiency of benzylamine biosynthesis, we identified the enzymes involved in the multi-enzyme cascade, regulated the expression strength by using RBS engineering in Escherichia coli, and established a regeneration-recycling system for alanine. This is a cosubstrate, coupled to cascade reactions, which resulted in E. coli RARE-TP and can synthesize benzylamine using phenylalanine as a precursor. By optimizing the supply of cosubstrates alanine and ammonia, the yield of benzylamine produced by whole-cell catalysis was increased by 1.5-fold and 2.7-fold, respectively, and the final concentration reached 6.21 mM. In conclusion, we achieved conversion from l-phenylalanine to benzylamine and increased the yield through enzyme screening, expression regulation, and whole-cell catalytic system optimization. This demonstrated a green and sustainable benzylamine synthesis method, which provides a reference and additional information for benzylamine biosynthesis research.

High CTCF expression mediated by FGD5-AS1/miR-19a-3p axis is associated with immunosuppression and pancreatic cancer progression.

The most common reason for cancer-related death globally is predicted to be pancreatic cancer (PC), one of the deadliest cancers. The CCCTC-binding factor (CTCF) regulates the three-dimensional structure of chromatin, was reported to be highly regulated in various malignancies. However, the underlying biological functions and possible pathways via which CTCF promotes PC progression remain unclear. Herein, we examined the CTCF function in PC and discovered that CTCF expression in PC tissues was significantly raised compared to neighboring healthy tissues. Additionally, Kaplan-Meier survival analysis demonstrated a strong connection between elevated CTCF expression and poor patient prognosis. A study of the ROC curve (receiver operating characteristic) revealed an AUC value for CTCF of 0.968. Subsequent correlation analysis exhibited a strong relationship between immunosuppression and CTCF expression in PC. CTCF knockdown significantly inhibited the malignant biological process of PC in vitro and in vivo, suggesting that CTCF may be a potential PC treatment target. We also identified the FGD5 antisense RNA 1 (FGD5-AS1)/miR-19a-3p axis as a possible upstream mechanism for CTCF overexpression. In conclusion, our data suggest that ceRNA-mediated CTCF overexpression contributes to the suppression of anti-tumor immune responses in PC and could be a predictive biomarker and potential PC treatment target.

SNHG17 alters anaerobic glycolysis by resetting phosphorylation modification of PGK1 to foster pro-tumor macrophage formation in pancreatic ductal adenocarcinoma.

BACKGROUND: Within the tumor immune microenvironment (TME), tumor-associated macrophages (TAMs) are crucial in modulating polarization states to influence cancer development through metabolic reprogramming. While long non-coding RNAs (lncRNAs) have been shown to play a pivotal role in the progression of various cancers, the underlying mechanisms by which lncRNAs alter M2 polarization through macrophage metabolism remodeling remain unelucidated. METHODS: RNA sequencing was used to screen for differentially expressed lncRNAs in TAMs and normal tissue-resident macrophages (NTRMs) isolated from pancreatic ductal adenocarcinoma (PDAC) tissues, whilst RT-qPCR and FISH were employed to detect the expression level of SNHG17. Moreover, a series of in vivo and in vitro experiments were conducted to assess the functions of SNHG17 from TAMs in the polarization and glycolysis of M2-like macrophages and in the proliferation and metastasis of pancreatic cancer cells (PCs). Furthermore, Western blotting, RNA pull-down, mass spectrometry, RIP, and dual-luciferase assays were utilized to explore the underlying mechanism through which SNHG17 induces pro-tumor macrophage formation. RESULTS: SNHG17 was substantially enriched in TAMs and was positively correlated with a worse prognosis in PDAC. Meanwhile, functional assays determined that SNHG17 promoted the malignant progression of PCs by enhancing M2 macrophage polarization and anaerobic glycolysis. Mechanistically, SNHG17 could sponge miR-628-5p to release PGK1 mRNA and concurrently interact with the PGK1 protein, activating the pro-tumorigenic function of PGK1 by enhancing phosphorylation at the T168A site of PGK1 through ERK1/2 recruitment. Lastly, SNHG17 knockdown could reverse the polarization status of macrophages in PDAC. CONCLUSIONS: The present study illustrated the essential role of SNHG17 and its molecular mechanism in TAMs derived from PDAC, indicating that SNHG17 might be a viable target for PDAC immunotherapy.

KIN17 promotes tumor metastasis by activating EMT signaling in luminal-A breast cancer.

BACKGROUND: Breast cancer (BC), the most common cause of cancer death in women, overtook lung cancer as the leading cause of cancer worldwide in 2020. Although many studies have proposed KIN17 as a biomarker of tumorigenesis in different cancer types, its role in tumor metastasis, particularly in BC metastasis, has been underexplored. This study aimed to explore the role of KIN17 in BC metastasis. METHODS: Survival analyses was performed to identify the association between KIN17 expression and BC patient survival in silico. Using lentivirus constructs, we developed bidirectional KIN17 expression (KD, knockdown; OE, overexpression) cellular models of luminal-A (Lum-A) breast cancer MCF-7 cells. We performed in vitro wound healing, transwell with and without Matrigel assays, and in vivo tail-vein metastasis assay to evaluate the migration and invasion abilities of MCF-7 with stable KIN17 knockdown or overexpression. Western blotting was performed to compare the changes in protein expression. RESULTS: We found that KIN17 expression was associated with poor overall survival (OS), relapse-free survival (RFS), distant metastasis-free survival (DMFS) and post-progression survival (PPS), particularly in Lum-A breast cancer patients. Later, we found that KIN17 knockdown inhibited migration and invasion of MCF-7 cells via regulating EMT-associated signaling pathways in vitro and decreases metastatic spread of the disease in vivo. In contrast, KIN17 overexpression promoted migration and invasion of MCF-7 cells in vitro and increased the metastatic spread of the disease in vivo. CONCLUSIONS: Overall, our findings provide preliminary data which suggests KIN17 of importance to target in metastatic Lum-A patients.