Genetic Determinant of Silicibacter sp. TM1040 Motility

Silicibacter sp. TM1040 is a member of the Roseobacter clade. Roseobacters play an important role in sulfur cycling in the ocean by degrading dimethylsulfoniopropionate (DMSP). Roseobacters are found in communities associated with most marine habits, especially with marine algae. Therefore, the ability to sense, move towards and maintain the interaction is an important physiological trait for the symbiosis between roseobacters and dinoflagellate. Previous work from our laboratory demonstrated that TM1040 is chemotaxis towards DMSP and DMSP catabolites, and motility of TM1040 is important for growth of P. piscicida. In contrast to enteric bacteria, little is known about the genes regulating motility in roseobacter species. This study, revealed similarities between the genes associated with motility in TM1040 and those from other α-proteobacteria species, but most importantly, it identified three new regulators that maybe involved in regulating the motility of TM1040

CULTURED HAIR FOLLICLE CELLS FOR THE TREATMENT OF VITILIGO

Objective: The objective of this study was to develop a method to isolate cells from hair follicles and cultured them in a hydrogel.Methods: Different cell types obtained from hair follicles were investigated and mixed with three formulations of Lutrol® F-127-based hydrogels. The percentages of the cell attachment and viability were observed within 48 h.Results: The results showed that three cell types, including keratinocyte, dermal papilla, and melanocyte cells, were obtained, as shown by the expression of their corresponding genes. All formulations of the hydrogels supported cell attachment and viability. Interestingly, more than 60% cell attachment and viability were found in lutrol hydrogels supplemented with either fetal bovine serum (FBS) or heat-activated human serum.Conclusion: Higher cell attachment and viability were observed when the hair follicle cells were cultured in the hydrogel with FBS than the hydrogel with human serum. However, the lutrol gel formulation with human serum was more appropriate to be used in the future clinical study, as this formulation contained no animal-derived component

Proteus mirabilis ZapA Metalloprotease Degrades a Broad Spectrum of Substrates, Including Antimicrobial Peptides

The 54-kDa extracellular metalloprotease ZapA is an important virulence factor of uropathogenic Proteus mirabilis. While ZapA has the ability to degrade host immunoglobulins (Igs), the dramatic attenuation of virulence in ZapA mutants suggests that this enzyme may have a broader spectrum of activity. This hypothesis was tested by in vitro assays with purified ZapA and an array of purified protein or peptide substrates. The data reveal that many proteins found in the urinary tract are substrates of ZapA proteolysis, including complement (C1q and C3), cell matrix (collagen, fibronectin, and laminin), and cytoskeletal proteins (actin and tubulin). Proteolysis of IgA and IgG was significantly enhanced by conditions that denatured the Igs. It was discovered that the antimicrobial peptides human β-defensin 1 (hBD1) and LL-37 are readily cleaved by the enzyme. To the best of our knowledge, this is the first report of a bacterial protease capable of cleaving hBD1, a component of the human renal tubule innate immune response. Proteolysis of hBD1 resulted in ca. six peptides, while proteolysis of LL-37 resulted in at least nine products. Matrix-assisted laser desorption ionization-time of flight mass spectrometry analysis of the molecular masses of the reaction products indicated that ZapA preferred no distinct peptide bond. The antimicrobial activity of hBD1 and LL-37 was significantly reduced following ZapA treatment, suggesting that proteolysis results in inactivation of these peptides. The data suggest that a function of ZapA during urinary tract infections is the proteolysis of antimicrobial peptides associated with the innate immune response

Genetic Determinants of Silicibacter sp. TM1040 Motility▿ †

Silicibacter sp. TM1040 is a member of the marine Roseobacter clade of Alphaproteobacteria that forms symbioses with unicellular eukaryotic phytoplankton, such as dinoflagellates. The symbiosis is complex and involves a series of steps that physiologically change highly motile bacteria into cells that readily form biofilms on the surface of the host. The initial phases of symbiosis require bacterial motility and chemotaxis that drive the swimming bacteria toward their planktonic host. Cells lacking wild-type motility fail to establish biofilms on host cells and do not produce effective symbioses, emphasizing the importance of understanding the molecular mechanisms controlling flagellar biosynthesis and the biphasic “swim-or-stick” switch. In the present study, we used a combination of bioinformatic and genetic approaches to identify the genes critical for swimming of Silicibacter sp. TM1040. More than 40 open reading frames with homology to known flagellar structural and regulatory genes were identified, most of which are organized into approximately eight operons comprising a 35.4-kb locus, with surprising similarity to the fla2 locus of Rhodobacter sphaeroides. The genome has homologs of CckA, CtrA, FlbT, and FlaF, proteins that in Caulobacter crescentus regulate flagellum biosynthesis. In addition, we uncovered three novel genes, flaB, flaC, and flaD, which encode flagellar regulatory proteins whose functions are likely to involve regulation of motor function (FlaD) and modulation of the swim-or-stick switch (FlaC). The data support the conclusion that Silicibacter sp. TM1040 uses components found in other Alphaproteobacteria, as well as novel molecular mechanisms, to regulate the expression of the genes required for motility and biofilm formation. These unique molecular mechanisms may enhance the symbiosis and survival of Roseobacter clade bacteria in the marine environment