Improvement of polyhydroxybutyrate production by deletion of csrA in Escherichia coli

BACKGROUND: Poly-3-hydroxybutyrate (PHB) can be efficiently produced in recombinant Escherichia coli by the overexpression of an operon (NphaCAB) encoding PHB synthetase. Strain improvement is considered to be one of critical factors to lower the production cost of PHB in recombinant system. In this study, one of key regulators that affect the cell growth and PHB content was confirmed and analyzed. RESULT: S17-3, a mutant E. coli strain derived from S17-1, was found to be able to achieve high cell density when expressing NphaCAB with the plasmid pBhya-CAB. Whole genome sequencing of S17-3 revealed genetic alternations on the upstream regions of csrA, encoding a global regulator cross-talking between stress response, catabolite repression and other metabolic activities. Deletion of csrA or expression of mutant csrA resulted in improved cell density and PHB content. CONCLUSION: The impact of gene deletion of csrA was determined, dysfunction of the regulators improved the cell density of recombinant E. coli and PHB production, however, the detail mechanism needs to be further clarified.

Over-expression of miR-141-3p promotes malignant biological behaviors of ovarian cancer A2780 cells by down-regulating PTEN and activating PI3K/Akt signaling pathway / 中国肿瘤生物治疗杂志

@#Objective: To explore the effect of miR-141-3p on the proliferation, invasion and apoptosis of ovarian cancer cells via targeting PTEN and regulating PI3K/Akt pathway. Methods: Collecting twenty-eight cases pairs of ovarian cancerovarian cancer patients with tumor tissues and adjacent tissues were collected from patients, who from April 2014 to October 2017 were treated in the Department of Obstetrics and Gynecology. qPCR was applied to detect the expression of miR-141-3p in ovarian cancer tissues and cell lines. The relationship between miR-141-3p and PTEN was verified by dual-luciferase reporter gene assay. After over-expression or knockdown of miR-141 and PTEN genes, the cell viability, invasion and apoptosis of ovarian cancer A2780 cells were examined by CCK-8 assay, Transwell assay and Annexin V-FITC/PI double staining flow cytometry assay, respectively. Furthermore, the effect of miR-1413p on PTEN-PI3K/Akt signaling pathway was measured by WB. Results: miR-141-3p is was highly expressed in ovarian cancer tissues and cell lines (P<0.05 or P<0.01). The dual luciferase reporter gene assay confirmed that miR-141-3p targets PTEN was a target of miR-141-3p and downregulates its expression level was down-regulated (P<0.01). Compared with the control group, after knockdown of miR-141-3p, the proliferation ofA2780 cells was significantly inhibited after knockdown of miR-141-3p (at 48 h, 0.36±0.04 vs 0.82± 0.06, P<0.05), and the invasive ability of A2780 cells was significantly reduced (number of transmembrane cells: 215.32±16.04 vs 45.14±7.88, P<0.01), while the apoptotic rate was significantly increased ([1.85±0.26]% vs [9.29±0.65]%, P<0.01). Over-expression of PTEN significantly inhibited the expression of p-Akt and cell proliferation and invasion in A2780 cells (all P<0.01), inhibited cell proliferation and invasion (all P<0.01) and significantly promoted apoptosis (all P<0.01). However, simultaneous over-expression of miR141-3p or addition of IGF-1 wile over-expressing PTEN can offset the above effects. Conclusion: miR-141-3p facilitates the proliferation, invasion and decreases apoptosis of A2780 cells. The mechanism may be related to targeted regulation of PTEN and activation of PI3K/Akt pathway.