Mechanisms of Alveolar Type 2 Epithelial Cell Death During Acute Lung Injury.

Diffuse alveolar epithelial cell (AEC) death occurs extensively during acute lung injury (ALI). Due to the limited proliferative capacity of alveolar type 1 epithelial (AT1) cells, the differentiation and regenerative capacity of alveolar type 2 epithelial (AT2) cells are required to restore the barrier function of AECs. However, during lung injury, AT1 cells are particularly susceptible to injury, and ATII cells die in the presence of severe or certain types of injury. This disruption ultimately results in a hindrance to the ability of AT2 cells to proliferate and differentiate into AT1 cells in time to repair the extensively damaged AECs. Therefore, understanding the mechanism of injury death of AT2 cells may be beneficial to reverse the above situation. This article reviews the main death modes of AT2 cells, including apoptosis, necrosis, necroptosis, pyroptosis, autophagic cell death, and ferroptosis. It compares the various forms of death, showing that various cell injury death modes have unique action mechanisms and partially overlapping pathways. Studying the mechanism of AT2 cell death is helpful in screening and analyzing the target pathway of AEC barrier function recovery. It opens up new ideas and strategies for preventing and treating ALI.

A novel point mutation in RpoB improves osmotolerance and succinic acid production in Escherichia coli.

BACKGROUND: Escherichia coli suffer from osmotic stress during succinic acid (SA) production, which reduces the performance of this microbial factory. RESULTS: Here, we report that a point mutation leading to a single amino acid change (D654Y) within the ß-subunit of DNA-dependent RNA polymerase (RpoB) significantly improved the osmotolerance of E. coli. Importation of the D654Y mutation of RpoB into the parental strain, Suc-T110, increased cell growth and SA production by more than 40% compared to that of the control under high glucose osmolality. The transcriptome profile, determined by RNA-sequencing, showed two distinct stress responses elicited by the mutated RpoB that counterbalanced the osmotic stress. Under non-stressed conditions, genes involved in the synthesis and transport of compatible solutes such as glycine-betaine, glutamate or proline were upregulated even without osmotic stimulation, suggesting a "pre-defense" mechanism maybe formed in the rpoB mutant. Under osmotic stressed conditions, genes encoding diverse sugar transporters, which should be down-regulated in the presence of high osmotic pressure, were derepressed in the rpoB mutant. Additional genetic experiments showed that enhancing the expression of the mal regulon, especially for genes that encode the glycoporin LamB and maltose transporter, contributed to the osmotolerance phenotype. CONCLUSIONS: The D654Y single amino acid substitution in RpoB rendered E. coli cells resistant to osmotic stress, probably due to improved cell growth and viability via enhanced sugar uptake under stressed conditions, and activated a potential "pre-defense" mechanism under non-stressed conditions. The findings of this work will be useful for bacterial host improvement to enhance its resistance to osmotic stress and facilitate bio-based organic acids production.

Osmotolerance in Escherichia coli Is Improved by Activation of Copper Efflux Genes or Supplementation with Sulfur-Containing Amino Acids.

Improvement in the osmotolerance of Escherichia coli is essential for the production of high titers of various bioproducts. In this work, a cusS mutation that was identified in the previously constructed high-succinate-producing E. coli strain HX024 was investigated for its effect on osmotolerance. CusS is part of the two-component system CusSR that protects cells from Ag(I) and Cu(I) toxicity. Changing cusS from strain HX024 back to its original sequence led to a 24% decrease in cell mass and succinate titer under osmotic stress (12% glucose). When cultivated with a high initial glucose concentration (12%), introduction of the cusS mutation into parental strain Suc-T110 led to a 21% increase in cell mass and a 40% increase in succinate titer. When the medium was supplemented with 30 g/liter disodium succinate, the cusS mutation led to a 120% increase in cell mass and a 492% increase in succinate titer. Introducing the cusS mutation into the wild-type strain ATCC 8739 led to increases in cell mass of 87% with 20% glucose and 36% using 30 g/liter disodium succinate. The cusS mutation increased the expression of cusCFBA, and gene expression levels were found to be positively related to osmotolerance abilities. Because high osmotic stress has been associated with deleterious accumulation of Cu(I) in the periplasm, activation of CusCFBA may alleviate this effect by transporting Cu(I) out of the cells. This hypothesis was confirmed by supplementing sulfur-containing amino acids that can chelate Cu(I). Adding methionine or cysteine to the medium increased the osmotolerance of E. coli under anaerobic conditions.IMPORTANCE In this work, an activating Cus copper efflux system was found to increase the osmotolerance of E. coli In addition, new osmoprotectants were identified. Supplementation with methionine or cysteine led to an increase in osmotolerance of E. coli under anaerobic conditions. These new strategies for improving osmotolerance will be useful for improving the production of chemicals in industrial bioprocesses.

Improving Succinate Productivity by Engineering a Cyanobacterial CO2 Concentrating System (CCM) in Escherichia coli.

Biologically fixation of CO2 has great potential as a significant carbon source for biosynthesis, which is also a major way to reduce CO2 accumulation in atmosphere. Phosphoenolpyruvate (PEP) carboxylation is the key step of anaerobic succinate production in Escherichia coli. In this reaction, one mole CO2 is assimilated with PEP to form oxaloacetate by PEP carboxykinase (PCK). The preferred substrate of PCK is CO2 , which is very limited in cytoplasm. In this study, the carbon concentration mechanism (CCM) of cyanobacteria was introduced into Escherichia coli to enhance the intracellular inorganic carbon concentration for improving carboxylation velocity. Overexpression of the bicarbonate transporter (BT) or carbonic anhydrase (CA) gene from Synechococcus sp. PCC7002 led to a 22 or 35% increase in succinate titer at 36 h, respectively. The carboxylation rate of PCK increased from 2.46 to 3.92 µmol min-1 mg-1 protein by overexpression of the CA gene. In addition, co-overexpression of BT and CA genes had a synergetic effect, leading to a 44% increase in succinate titer at 36 h. This work is the first attempt to increase carbon fixation involved in microbial biosynthesis by engineering a biological CO2 delivery system, which provides new direction and strategies for improving industrial fermentations based on biological CO2 assimilation pathways.