Application of Metal-Organic Frameworks (MOFs) in Environmental Biosystems.

Metal-organic frameworks (MOFs) are crystalline materials that are formed by self-assembling organic linkers and metal ions with large specific areas and pore volumes. Their chemical tunability, structural diversity, and tailor-ability make them adaptive to decorate many substrate materials, such as biomass-derived carbon materials, and competitive in many environmental biosystems, such as biofuel cells, bioelectrocatalysts, microbial metal reduction, and fermentation systems. In this review, we surmised the recent progress of MOFs and MOF-derived materials and their applications in environmental biosystems. The behavior of MOFs and MOF-derived materials in different environmental biosystems and their influences on performance are described. The inherent mechanisms will guide the rational design of MOF-related materials and lead to a better understanding of their interaction with biocomponents.

Proteasome malfunction activates macroautophagy in the heart.

Protein quality control (PQC) senses and repairs misfolded/unfolded proteins and, if the repair fails, degrades the terminally misfolded polypeptides through an intricate collaboration between molecular chaperones and targeted proteolysis. Proteolysis of damaged proteins is performed primarily by the ubiquitin-proteasome system (UPS). Macroautophagy (commonly known as autophagy) may also play a role in PQC-associated proteolysis, especially when UPS function becomes inadequate. The development of a range of heart diseases, including bona fide cardiac proteinopathies and various forms of cardiac dysfunction has been linked to proteasome functional insufficiency (PFI). Both PFI and activation of autophagy have been observed in the heart of well-established mouse models of cardiac proteinopathy. A causal relationship between PFI and autophagic activation was suggested by a study using cultured cardiomyocytes but has not been established in the heart of intact animals. Taking advantage of an autophagy reporter, we demonstrated here that pharmacologically induced proteasome inhibition is sufficient to activate autophagy in cardiomyocytes in both intact animals and cell cultures, unveiling a potential cross-talk between the two major degradation pathways in cardiac PQC.

Autophagy and p62 in cardiac proteinopathy.

RATIONALE: Recent studies suggest an important role of autophagy in protection against αB-crystallin-based (CryAB(R120G)) desmin-related cardiomyopathies (DRC), but this has not been demonstrated in a different model of cardiac proteinopathy. Mechanisms underlying the response of cardiomyocytes to proteotoxic stress remain incompletely understood. OBJECTIVE: Our first objective was to determine whether and how the autophagic activity is changed in a mouse model of desminopathy. We also investigated the role of p62 in the protein quality control of cardiomyocytes. METHODS AND RESULTS: Using an autophagosome reporter and determining changes in LC3-II protein levels in response to lysosomal inhibition, we found significantly increased autophagic flux in mouse hearts with transgenic overexpression of a DRC-linked mutant desmin. Similarly, autophagic flux was increased in cultured neonatal rat ventricular myocytes (NRVMs) expressing a mutant desmin. Suppression of autophagy by 3-methyladenine increased, whereas enhancement of autophagy by rapamycin reduced the ability of a comparable level of mutant desmin overexpression to accumulate ubiquitinated proteins in NRVMs. Furthermore, p62 mRNA and protein expression was significantly up-regulated in cardiomyocytes by transgenic overexpression of the mutant desmin or CryAB(R120G) both in intact mice and in vitro. The p62 depletion impaired aggresome and autophagosome formation, exacerbated cell injury, and decreased cell viability in cultured NRVMs expressing the misfolded proteins. CONCLUSIONS: Autophagic flux is increased in desminopathic hearts, and as previously suggested in CryAB(R120G)-based DRC, this increased autophagic flux serves as an adaptive response to overexpression of misfolded proteins. The p62 is up-regulated in mouse proteinopathic hearts. The p62 promotes aggresome formation and autophagy activation and protects cardiomyocytes against proteotoxic stress.

Doxycycline attenuates protein aggregation in cardiomyocytes and improves survival of a mouse model of cardiac proteinopathy.

OBJECTIVES: The goal of this pre-clinical study was to assess the therapeutic efficacy of doxycycline (Doxy) for desmin-related cardiomyopathy (DRC) and to elucidate the potential mechanisms involved. BACKGROUND: DRC, exemplifying cardiac proteinopathy, is characterized by intrasarcoplasmic protein aggregation and cardiac insufficiency. No effective treatment for DRC is available presently. Doxy was shown to attenuate aberrant intranuclear aggregation and toxicity of misfolded proteins in noncardiac cells and animal models of other proteinopathies. METHODS: Mice and cultured neonatal rat cardiomyocytes with transgenic (TG) expression of a human DRC-linked missense mutation R120G of αB-crystallin (CryAB(R120G)) were used for testing the effect of Doxy. Doxy was administered via drinking water (6 mg/ml) initiated at 8 or 16 weeks of age. RESULTS: Doxy treatment initiated at 16 weeks of age significantly delayed the premature death of CryAB(R120G) TG mice, with a median lifespan of 30.4 weeks (placebo group, 25 weeks; p < 0.01). In another cohort of CryAB(R120G) TG mice, Doxy treatment initiated at 8 weeks of age significantly attenuated cardiac hypertrophy in 1 month. Further investigation revealed that Doxy significantly reduced the abundance of CryAB-positive microscopic aggregates, detergent-resistant CryAB oligomers, and total ubiquitinated proteins in CryAB(R120G) TG hearts. In cell culture, Doxy treatment dose-dependently suppressed the formation of both microscopic protein aggregates and detergent-resistant soluble CryAB(R120G) oligomers and reversed the up-regulation of p62 protein induced by adenovirus-mediated CryAB(R120G) expression. CONCLUSIONS: Doxy suppresses CryAB(R120G)-induced aberrant protein aggregation in cardiomyocytes and prolongs CryAB(R120G)-based DRC mouse survival.

Interplay between the ubiquitin-proteasome system and autophagy in proteinopathies.

Proteinopathies are a family of human disease caused by toxic aggregation-prone proteins and featured by the presence of protein aggregates in the affected cells. The ubiquitin-proteasome system (UPS) and autophagy are two major intracellular protein degradation pathways. The UPS mediates the targeted degradation of most normal proteins after performing their normal functions as well as the removal of abnormal, soluble proteins. Autophagy is mainly responsible for degradation of defective organelles and the bulk degradation of cytoplasm during starvation. The collaboration between the UPS and autophagy appears to be essential to protein quality control in the cell. UPS proteolytic function often becomes inadequate in proteinopathies which may lead to activation of autophagy, striving to remove abnormal proteins especially the aggregated forms. HADC6, p62, and FoxO3 may play an important role in mobilizing this proteolytic consortium. Benign measures to enhance proteasome function are currently lacking; however, enhancement of autophagy via pharmacological intervention and/or lifestyle change has shown great promise in alleviating bona fide proteinopathies in the cell and animal models. These pharmacological interventions are expected to be applied clinically to treat human proteinopathies in the near future.

[Cloning of ureI gene from Helicobacter pylori and its expression in E. coli].

OBJECTIVE: To clone the urea membrane channel gene (ureI) from Helicobacter pylori (Hp) for its expression in E. coli, and evaluate the expression conditions and immunological features of the fusion protein. METHODS: ureI gene cloned by PCR from Hp was inserted into the plasmid pET32a (+) to construct the recombinant plasmid pET32a/ureI, followed by identification by BglII and HindIII digestion and sequencing. E. coli BL-21+(DE3) was transformed with pET32a/ureI to obtain the engineered bacterium BL21+/UreI, which was cultured at different temperatures and induced with 1.0 mmol/L IPTG for expression of the recombinant protein. The expressed proteins were identified by SDS-PAGE and analyzed by Pro-gel analyzer 4.0. Western blotting was performed to evaluate the immunogenicity of the expressed protein. RESULTS: The cloned gene fragment was about 650 bp in length, and BglII and HindIII digestion of pET32a/ureI yielded a 650-bp band. Sequence analysis revealed that the cloned ureI gene contained 646 bp without reading frame alterations. Comparison against GenBank indicated a homology of 100% of the cloned gene with ureI gene of the corresponding Hp strains, and also one no less than 98.5% with ureI gene from other strains. The engineered E. coli BL21+/UreI could express recombinant UreI (rUreI) with His tag, and the target protein accounted for 20.2% of the total bacterial proteins after 1.0 mmol/L IPTG induction of the bacterium at 37 degrees C for 14 h. SDS-PAGE and Western blotting showed that the recombinant UreI protein was produced mainly in the inclusion bodies and fused with his-tag (rUreI/his), which could react with human anti-Hp and mAb to his tag but not with mAb to Hp UreB. CONCLUSIONS: We have successfully cloned ureI gene and constructed the prokaryotic expression plasmid for efficient rUreI expression, and the fusion protein rUreI/his expressed in the inclusion bodies can react specifically with both Hp antibody and his-tag antibody.