Dynamic periplasmic chaperone reservoir facilitates biogenesis of outer membrane proteins.

Outer membrane protein (OMP) biogenesis is critical to bacterial physiology because the cellular envelope is vital to bacterial pathogenesis and antibiotic resistance. The process of OMP biogenesis has been studied in vivo, and each of its components has been studied in isolation in vitro. This work integrates parameters and observations from both in vivo and in vitro experiments into a holistic computational model termed "Outer Membrane Protein Biogenesis Model" (OMPBioM). We use OMPBioM to assess OMP biogenesis mathematically in a global manner. Using deterministic and stochastic methods, we are able to simulate OMP biogenesis under varying genetic conditions, each of which successfully replicates experimental observations. We observe that OMPs have a prolonged lifetime in the periplasm where an unfolded OMP makes, on average, hundreds of short-lived interactions with chaperones before folding into its native state. We find that some periplasmic chaperones function primarily as quality-control factors; this function complements the folding catalysis function of other chaperones. Additionally, the effective rate for the ß-barrel assembly machinery complex necessary for physiological folding was found to be higher than has currently been observed in vitro. Overall, we find a finely tuned balance between thermodynamic and kinetic parameters maximizes OMP folding flux and minimizes aggregation and unnecessary degradation. In sum, OMPBioM provides a global view of OMP biogenesis that yields unique insights into this essential pathway.

Cudraxanthone H Induces Growth Inhibition and Apoptosis in Oral Cancer Cells via NF-κB and PIN1 Pathways.

Cudraxanthone H (CH) is a natural compound isolated from a methanol extract of the root bark of Cudrania tricuspidata, a herbal plant also known as Moraceae. However, the effect of CH on human cancer cells has not been reported previously. The aim of this study was to investigate the anticancer effects and mechanism of action of CH on oral squamous cell carcinoma (OSCC) cells. CH exerted significant antiproliferative effects on OSCC cells in dose- and time-dependent manners. CH also induced apoptosis in OSCC cells, as evidenced by an increased percentage of cells in the sub-G1 phase of the cell cycle, annexin V-positive/propidium iodide-negative cells, and nuclear morphology. This antiproliferative effect of CH was associated with a marked reduction in the expression of cyclin D1 and cyclin E, with a concomitant induction of cyclin-dependent kinase inhibitor (CDKI) expression (p21 and p27). CH inhibited the phosphorylation and degradation of IκB-α and the nuclear translocation of NF-κB p65. Furthermore, CH treatment down-regulated PIN1 mRNA and protein expression in a dose-dependent manner. PIN1 overexpression by infection with adenovirus-PIN1 (Ad-PIN1) attenuated the CH-induced growth-inhibiting and apoptosis-inducing effects, blocked CH-enhanced CDKI expression and restored cyclin levels. In contrast, inhibiting PIN1 expression via juglone exerted the opposite effects. The present study is the first to demonstrate antiproliferative and apoptosis-inducing effects of CH, which exerts its effects by inhibiting NF-κB and PIN1. These data suggest that it might be a novel alternative chemotherapeutic agent for use in the treatment of oral cancer.

Functional replacement of the essential ESS1 in yeast by the plant parvulin DlPar13.

A functionally Pin1-like peptidyl-prolyl cis/trans isomerase (PPIase(1)) was isolated from proembryogenic masses (PEMs) of Digitalis lanata according to its enzymatic activity. Partial sequence analysis of the purified enzyme (DlPar13) revealed sequence homology to members of the parvulin family of PPIases. Similar to human Pin1 and yeast Ess1, it exhibits catalytic activity toward substrates containing (Thr(P)/Ser(P))-Pro peptide bonds and comparable inhibition kinetics with juglone. Unlike Pin1-type enzymes it lacks the phosphoserine or phosphothreonine binding WW domain. Western blotting with anti-DlPar13 serum recognized the endogenous form in nucleic and cytosolic fractions of the plant cells. Since the PIN1 homologue ESS1 is an essential gene, complementation experiments in yeast were performed. When overexpressed in Saccharomyces cerevisiae DlPar13 is almost as effective as hPin1 in rescuing the temperature-sensitive phenotype caused by a mutation in ESS1. In contrast, the human parvulin hPar14 is not able to rescue the lethal phenotype of this yeast strain at nonpermissive temperatures. These results suggest a function for DlPar13 rather similar to parvulins of the Pin1-type.