Melanin is a pigment produced by organisms throughout all domains of life. Due to its unique physicochemical properties, biocompatibility, and biostability, there has been an increasing interest in the use of melanin for broad applications. In the vast majority of studies, melanin has been either chemically synthesized or isolated from animals, which has restricted its use to small-scale applications. Using bacteria as biocatalysts is a promising and economical alternative for the large-scale production of biomaterials. In this study, we engineered the marine bacterium Vibrio natriegens, one of the fastest-growing organisms, to synthesize melanin by expressing a heterologous tyrosinase gene and demonstrated that melanin production was much faster than in previously reported heterologous systems. The melanin of V. natriegens was characterized as a polymer derived from dihydroxyindole-2-carboxylic acid (DHICA) and, similarly to synthetic melanin, exhibited several characteristic and useful features. Electron microscopy analysis demonstrated that melanin produced from V. natriegens formed nanoparticles that were assembled as “melanin ghost” structures, and the photoprotective properties of these particles were validated by their protection of cells from UV irradiation. Using a novel electrochemical reverse engineering method, we observed that melanization conferred redox activity to V. natriegens. Moreover, melanized bacteria were able to quickly adsorb the organic compound trinitrotoluene (TNT). Overall, the genetic tractability, rapid division time, and ease of culture provide a set of attractive properties that compare favorably to current E. coli production strains and warrant the further development of this chassis as a microbial factory for natural product biosynthesis. IMPORTANCE Melanins are macromolecules that are ubiquitous in nature and impart a large variety of biological functions, including structure, coloration, radiation resistance, free radical scavenging, and thermoregulation. Currently, in the majority of investigations, melanins are either chemically synthesized or extracted from animals, which presents significant challenges for large-scale production. Bacteria have been used as biocatalysts to synthesize a variety of biomaterials due to their fast growth and amenability to genetic engineering using synthetic biology tools. In this study, we engineered the extremely fast-growing bacterium V. natriegens to synthesize melanin nanoparticles by expressing a heterologous tyrosinase gene with inducible promoters. Characterization of the melanin produced from V. natriegens-produced tyrosinase revealed that it exhibited physical and chemical properties similar to those of natural and chemically synthesized melanins, including nanoparticle structure, protection against UV damage, and adsorption of toxic compounds. We anticipate that producing and controlling melanin structures at the nanoscale in this bacterial system with synthetic biology tools will enable the design and rapid production of novel biomaterials for multiple applications.

Here, we report on the development of a genetic system for Marinobacter sp. strain CP1, previously isolated from the Biocathode MCL community and shown to oxidize iron and grow as a cathodic biofilm. Sequence analysis of the small and large subunits of the 16S rRNA gene of CP1, as well as comparison of select conserved proteins, indicate that it is most closely related to Marinobacter adhaerens HP15 and Marinobacter sp. ES.042. In silico DNA–DNA hybridization using the genome-to-genome distance calculator (GGDC) predicts CP1 to be a new species of Marinobacter described here as Marinobacter atlanticus. CP1 is competent for transformation with plasmid DNA using conjugation with Escherichia coli donor strain WM3064 and constitutive expression of green fluorescent protein (GFP) is stable in the absence of antibiotic selection. Targeted double deletion mutagenesis of homologs for the M. aquaeoli fatty acyl-CoA reductase (acrB) and fatty aldehyde reductase (farA) genes resulted in a loss of production of wax esters; however, single deletion mutants for either gene resulted in an increase in total wax esters recovered. Genetic tools presented here for CP1 will enable further exploration of wax ester synthesis for biotechnological applications, as well as furthering our efforts to understand the role of CP1 within the Biocathode MCL community.

Contributions of mechanical signals to airway remodeling during asthma are poorly understood. Transient receptor potential vanilloid 4 (TRPV4), a mechanosensitive ion channel, has been implicated in cardiac and pulmonary fibrosis; however, its role in asthma remains elusive. Employing a -induced asthma model, we report here that TRPV4-knockout mice were protected from-induced airway remodeling. Furthermore, lung fibroblasts that were isolated from TRPV4-knockout mice showed diminished differentiation potential compared with wild-type mice. Fibroblasts from asthmatic lung exhibited increased TRPV4 activity and enhanced differentiation potential compared with normal human lung fibroblasts. Of interest, TGF-β1 treatment enhanced TRPV4 activation in a PI3K-dependent manner in normal human lung fibroblasts Mechanistically, TRPV4 modulated matrix remodeling in the lung 2 distinct but dependent pathways: one enhances matrix deposition by fibrotic gene activation, whereas the other slows down matrix degradation by increased plasminogen activator inhibitor 1. Of importance, both pathways are regulated by Rho/myocardin-related transcription factor-A and contribute to fibroblast differentiation and matrix remodeling in the lung. Thus, our results support a unique role for TRPV4 in -induced airway remodeling and warrant further studies in humans for it to be used as a novel therapeutic target in the treatment of asthma.-Gombedza, F., Kondeti, V., Al-Azzam, N., Koppes, S., Duah, E., Patil, P., Hexter, M., Phillips, D., Thodeti, C. K., Paruchuri, S. Mechanosensitive transient receptor potential vanilloid 4 regulates-induced airway remodeling 2 distinct pathways modulating matrix synthesis and degradation.

Recent reports have shown that Gram-positive bacteria actively secrete spherical nanometer-sized proteoliposome membrane vesicles (MVs) into their surroundings. Though MVs are implicated in a broad range of biological functions, few studies have been conducted to examine their potential as delivery vehicles of antimicrobials. Here, we investigate the natural ability of Lactobacillus acidophilus MVs to carry and deliver bacteriocin peptides to the opportunistic pathogen, Lactobacillus delbrueckii. We demonstrate that upon treatment with lactacin B-inducing peptide, the proteome of the secreted MVs is enriched in putative bacteriocins encoded by the lab operon. Further, we show that purified MVs inhibit growth and compromise membrane integrity in L. delbrueckii, which is confirmed by confocal microscopy imaging and spectrophotometry. These results show that L. acidophilus MVs serve as conduits for antimicrobials to competing cells in the environment, suggesting a potential role for MVs in complex communities such as the gut microbiome. With the potential for controlling their payload through microbial engineering, MVs produced by L. acidophilus may be an interesting platform for effecting change in complex microbial communities or aiding in the development of new biomedical therapeutics.