The Mechano-Ubiquitinome of Articular Cartilage: Differential Ubiquitination and Activation of a Group of ER-Associated DUBs and ER Stress Regulators.

Understanding how connective tissue cells respond to mechanical stimulation is important to human health and disease processes in musculoskeletal diseases. Injury to articular cartilage is a key risk factor in predisposition to tissue damage and degenerative osteoarthritis. Recently, we have discovered that mechanical injury to connective tissues including murine and porcine articular cartilage causes a significant increase in lysine-63 polyubiquitination. Here, we identified the ubiquitin signature that is unique to injured articular cartilage tissue upon mechanical injury (the "mechano-ubiquitinome"). A total of 463 ubiquitinated peptides were identified, with an enrichment of ubiquitinated peptides of proteins involved in protein processing in the endoplasmic reticulum (ER), also known as the ER-associated degradation response, including YOD1, BRCC3, ATXN3, and USP5 as well as the ER stress regulators, RAD23B, VCP/p97, and Ubiquilin 1. Enrichment of these proteins suggested an injury-induced ER stress response and, for instance, ER stress markers DDIT3/CHOP and BIP/GRP78 were upregulated following cartilage injury on the protein and gene expression levels. Similar ER stress induction was also observed in response to tail fin injury in zebrafish larvae, suggesting a generic response to tissue injury. Furthermore, a rapid increase in global DUB activity following injury and significant activity in human osteoarthritic cartilage was observed using DUB-specific activity probes. Combined, these results implicate the involvement of ubiquitination events and activation of a set of DUBs and ER stress regulators in cellular responses to cartilage tissue injury and in osteoarthritic cartilage tissues. This link through the ER-associated degradation pathway makes this protein set attractive for further investigation in in vivo models of tissue injury and for targeting in osteoarthritis and related musculoskeletal diseases.

Biodegradation of crude oil by immobilized Exiguobacterium sp. AO-11 and shelf life evaluation.

Exiguobacterium sp. AO-11 was immobilized on bio-cord at 109 CFU g-1 carrier for the removal of crude oil from marine environments. To prepare a ready-to-use bioremediation product, the shelf life of the immobilized cells was calculated. Approximately 90% of 0.25% (v/v) crude oil removal was achieved within 9 days when the starved state of immobilized cells was used. The oil removal activity of the immobilized cells was maintained in the presence of oil dispersant (89%) and at pH values of 7-9. Meanwhile, pH, oil concentration and salinity affected the oil removal efficacy. The immobilized cells could be reused for at least 5 cycles. The Arrhenius equation describing the relationship between the rate of reaction and temperature was validated as a useful model of the kinetics of retention of activity by an immobilized biocatalyst. It was estimated that the immobilized cells could be stored in a non-vacuum bag containing phosphate buffer (pH 7.0) at 30 °C for 39 days to retain the cells at 107 CFU g-1 carrier and more than 50% degradation activity. These results indicated the potential of using bio-cord-immobilized crude oil-degrading Exiguobacterium sp. AO-11 as a bioremediation product in a marine environment.