Improving the productivity of Candida glycerinogenes in the fermentation of ethanol from non-detoxified sugarcane bagasse hydrolysate by a hexose transporter mutant.

AIMS: In this study, we attempted to increase the productivity of Candida glycerinogenes yeast for ethanol production from non-detoxified sugarcane bagasse hydrolysates (NDSBH) by identifying the hexose transporter in this yeast that makes a high contribution to glucose consumption, and by adding additional copies of this transporter and enhancing its membrane localisation stability (MLS). METHODS AND RESULTS: Based on the knockout and overexpression of key hexose transporter genes and the characterisation of their promoter properties, we found that Cghxt4 and Cghxt6 play major roles in the early and late stages of fermentation, respectively, with Cghxt4 contributing most to glucose consumption. Next, subcellular localisation analysis revealed that a common mutation of two ubiquitination sites (K9 and K538) in Cghxt4 improved its MLS. Finally, we overexpressed this Cghxt4 mutant (Cghxt4.2A) using a strong promoter, PCgGAP , which resulted in a significant increase in the ethanol productivity of C. glycerinogenes in the NDSBH medium. Specifically, the recombinant strain showed 18 and 25% higher ethanol productivity than the control in two kinds of YP-NDSBH medium (YP-NDSBH1G160 and YP-NDSBH2G160 ), respectively. CONCLUSIONS: The hexose transporter mutant Cghxt4.2A (Cghxt4K9A,K538A ) with multiple copies and high MLS was able to significantly increase the ethanol productivity of C. glycerinogenes in NDSBH. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results provide a promising strategy for constructing efficient strains for ethanol production.

Enhanced 1,3-propanediol production in Klebsiella pneumoniae by a combined strategy of strengthening the TCA cycle and weakening the glucose effect.

AIMS: This study aimed to strengthen the reducing equivalent generation in Klebsiella pneumoniae for improving 1,3-propanediol (PDO) production. METHODS AND RESULTS: Disruption of the arcA gene activated the transcription levels of the TCA cycle genes and thus increased the NADH/NAD+ ratio by 54·2%, leading to the improved PDO titre and yield per cell from 16·1 g l-1 and 4·0 g gDCW-1 to 18·8 g l-1 and 6·4 g gDCW-1 respectively. Further ldhA gene deletion eliminated lactate accumulation and promoted the PDO titre to 19·9 g l-1 . Finally, the glucose effect was weakened by deleting the crr gene to enhance the co-utilization of glucose and glycerol, resulting in the increased PDO production to 23·8 g l-1 with the glycerol conversion rate of 59·5%. The PDO titre in bioreactor was promoted from 61·2 to 78·1 g l-1 . CONCLUSIONS: Deletions of the arcA and the crr genes showed positive effects on the TCA cycle activity and the co-utilization of glucose and glycerol, leading to the strengthened reducing equivalent generation and the improved PDO titre by 47·8% in shaker. The PDO titre in the bioreactor was enhanced to 78·1 g l-1 . SIGNIFICANCE AND IMPACT OF THE STUDY: This study provided novel information on generating reducing equivalent for the PDO biosynthesis by strengthening the TCA cycle and weakening the glucose effect in K. pneumoniae.

MASTL is a potential poor prognostic indicator in ER+ breast cancer.

OBJECTIVE: In this study, we aimed to explore prognostic value of MASTL (microtubule-associated serine/threonine kinase-like) in breast cancer patients on the basis of ER status and molecular subtypes. MATERIALS AND METHODS: The raw microarray data (GDS5666) of 4T1 derived bone-aggressive explant and primary tumor explant were reanalyzed to identify the dysregulated genes. To pool previous annotated genomic data that assessed the association between MASTL expression and metastatic relapse (MR) risk, MR-free survival, any event (AE, defined as any relapse or death) risk, and AE-free survival in breast cancer patients, a meta-analysis was performed by bc-GenExMiner 4.0. RESULTS: MASTL is a significantly upregulated gene in 4T1 bone-aggressive explant compared to primary tumor explant. Univariate Cox analysis showed that high MASTL expression is associated with a higher risk of MR (HR: 1.43, 95%CI: 1.28-1.60; p<0.001) and a higher risk of AE (HR: 1.27, 95%CI: 1.18-1.37; p<0.001) in ER+ breast cancer. Also, high MASTL expression also predicts a worse MR-free survival (HR: 1.74, 95%CI: 1.40-2.17; p<0.001) and a worse AE-free survival (HR: 1.42, 95%CI: 1.23-1.63; p<0.001) in ER+ breast cancer. However, the associations were not observed in ER- patients. The following NPI adjusted analyses confirmed the results of univariate Cox analysis. In Single Sample Predictors (SSPs) and Subtype Clustering Models (SCMs) subtypes, high MASTL expression is associated with increased risk of AE and predicts a poor AE-free survival in ER+ subgroups. CONCLUSIONS: MASTL might be a valuable indicator of MR risk and AE risk in ER+ patients, but not in ER- patients.