RESUMO
Signaling pathways that regulate blood-tissue barriers are important for studying the biology of various blood-tissue barriers. This information, if deciphered and better understood, will provide better therapeutic management of diseases particularly in organs that are sealed by the corresponding blood-tissue barriers from systemic circulation, such as the brain and the testis. These barriers block the access of antibiotics and/or chemotherapeutical agents across the corresponding barriers. Studies in the last decade using the blood-testis barrier (BTB) in rats have demonstrated the presence of several signaling pathways that are crucial to modulate BTB function. Herein, we critically evaluate these findings and provide hypothetical models regarding the underlying mechanisms by which these signaling molecules/pathways modulate BTB dynamics. This information should be carefully evaluated to examine their applicability in other tissue barriers which shall benefit future functional studies in the field. This article is part of a Special Issue entitled: Gap Junction Proteins edited by Jean Claude Herve.
Assuntos
Barreira Hematotesticular/metabolismo , Conexinas/metabolismo , Junções Comunicantes/metabolismo , Modelos Cardiovasculares , Transdução de Sinais/fisiologia , Animais , Humanos , MasculinoRESUMO
Certain plant-associating bacteria produce ice nucleation proteins (INPs) which allow the crystallization of water at high subzero temperatures. Many of these microbes are considered plant pathogens since the formed ice can damage tissues, allowing access to nutrients. Intriguingly, certain plants that host these bacteria synthesize antifreeze proteins (AFPs). Once freezing has occurred, plant AFPs likely function to inhibit the growth of large damaging ice crystals. However, we postulated that such AFPs might also serve as defensive mechanisms against bacterial-mediated ice nucleation. Recombinant AFP derived from the perennial ryegrass Lolium perenne (LpAFP) was combined with INP preparations originating from the grass epiphyte, Pseudomonas syringae. The presence of INPs had no effect on AFP activity, including thermal hysteresis and ice recrystallization inhibition. Strikingly, the ice nucleation point of the INP was depressed up to 1.9°C in the presence of LpAFP, but a recombinant fish AFP did not lower the INP-imposed freezing point. Assays with mutant LpAFPs and the visualization of bacterially-displayed fluorescent plant AFP suggest that INP and LpAFP can interact. Thus, we postulate that in addition to controlling ice growth, plant AFPs may also function as a defensive strategy against the damaging effects of ice-nucleating bacteria.
Assuntos
Proteínas Anticongelantes/química , Proteínas da Membrana Bacteriana Externa/química , Lolium/química , Proteínas de Plantas/química , Pseudomonas syringae/química , Proteínas Recombinantes de Fusão/química , Proteínas Anticongelantes/genética , Proteínas Anticongelantes/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Bioensaio , Cristalização , Congelamento , Expressão Gênica , Gelo , Cinética , Lolium/genética , Lolium/metabolismo , Lolium/microbiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Pseudomonas syringae/genética , Pseudomonas syringae/crescimento & desenvolvimento , Pseudomonas syringae/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , TemperaturaRESUMO
To produce food-grade ice nucleators, a 3.77 kb ice nucleation gene (iceE) isolated from Pantoea agglomerans (Erwinia herbicola) was introduced into the Gram-positive microorganism Bacillus amyloliquefaciens for the first time. The differential scanning calorimetry (DSC) results indicated that recombined strain B9-INP was an effective ice nucleator for controlling the supercooling point of distilled water at low concentrations. In the presence of B9-INP cells, model food systems, including sucrose solution and sodium chloride solution, different pH solutions froze at a relatively high subzero temperature, thus increasing the supercooling point by 5.8~16.7 °C. Moreover, B9-INP also facilitated model and real food systems to freeze at -6 °C. This recombinant strain not only improved the freezing temperature of food systems but also shortened the total freezing time, thus saving energy and reducing consumption. The results suggest that B9-INP has great application potential in the frozen food industry.
RESUMO
In nature, frost can form at a few degrees below 0 °C. However, this process requires the assembly of tens of thousands of ice-like water molecules that align together to initiate freezing at these relatively high temperatures. Water ordering on this scale is mediated by the ice nucleation proteins (INPs) of common environmental bacteria like Pseudomonas syringae and Pseudomonas borealis. However, individually, these 100 kDa proteins are too small to organize enough water molecules for frost formation, and it is not known how giant, megadalton-sized multimers, which are crucial for ice nucleation at high sub-zero temperatures, form. The ability of multimers to self-assemble was suggested when the transfer of an INP gene into Escherichia coli led to efficient ice nucleation. Here, we demonstrate that a positively charged subdomain at the C-terminal end of the central ß-solenoid of the INP is crucial for multimerization. Truncation, relocation, or change of the charge of this subdomain caused a catastrophic loss of ice nucleation ability. Cryo-electron tomography of the recombinant E. coli showed that the INP multimers form fibres that are ~5 nm across and up to 200 nm long. A model of these fibres as an overlapping series of antiparallel dimers can account for all their known properties and suggests a route to making cell-free ice nucleators for biotechnological applications.
Assuntos
Escherichia coli , Gelo , Congelamento , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , ÁguaRESUMO
Medicago sativa (also known as alfalfa), a forage legume, is widely cultivated due to its high yield and high-value hay crop production. Infectious diseases are a major threat to the crops, owing to huge economic losses to the agriculture industry, worldwide. The protein-protein interactions (PPIs) between the pathogens and their hosts play a critical role in understanding the molecular basis of pathogenesis. Pseudomonas syringae pv. syringae ALF3 suppresses the plant's innate immune response by secreting type III effector proteins into the host cell, causing bacterial stem blight in alfalfa. The alfalfa-P. syringae system has little information available for PPIs. Thus, to understand the infection mechanism, we elucidated the genome-scale host-pathogen interactions (HPIs) between alfalfa and P. syringae using two computational approaches: interolog-based and domain-based method. A total of â¼14 M putative PPIs were predicted between 50,629 alfalfa proteins and 2,932 P. syringae proteins by combining these approaches. Additionally, â¼0.7 M consensus PPIs were also predicted. The functional analysis revealed that P. syringae proteins are highly involved in nucleotide binding activity (GO:0000166), intracellular organelle (GO:0043229), and translation (GO:0006412) while alfalfa proteins are involved in cellular response to chemical stimulus (GO:0070887), oxidoreductase activity (GO:0016614), and Golgi apparatus (GO:0005794). According to subcellular localization predictions, most of the pathogen proteins targeted host proteins within the cytoplasm and nucleus. In addition, we discovered a slew of new virulence effectors in the predicted HPIs. The current research describes an integrated approach for deciphering genome-scale host-pathogen PPIs between alfalfa and P. syringae, allowing the researchers to better understand the pathogen's infection mechanism and develop pathogen-resistant lines.
RESUMO
Actin cytoskeleton is crucial to support spermatogenesis in the mammalian testis. However, the molecular mechanism(s) underlying changes of actin cytoskeletal organization in response to cellular events that take place across the seminiferous epithelium (e.g., self-renewal of spermatogonial stem cells, germ cell differentiation, meosis, spermiogenesis, spermiation) at specific stages of the epithelial cycle of spermatogenesis remain largely unexplored. This, at least in part, is due to the lack of suitable study models to identify the crucial regulatory proteins and to investigate how these proteins work in concert to support actin dynamics. Much of the information on the role of actin binding proteins in the literature, namely the actin bundling proteins, actin nucleation proteins and motor proteins, are either findings based on genetic models or morphological analyses. While this information is helpful to delineate the function of these proteins to support spermatogenesis, they are not helpful to identify the regulatory signaling proteins, the signaling pathways and the cascade of events to modulate actin cytoskeleton dynamics. Recent studies based on the use of toxicant models, both in vitro and in vivo, however, have bridged this gap by identifying putative regulatory and signaling proteins of actin cytoskeleton. Herein, we summarize and critically evaluate these findings. We also provide a hypothetical model by which actin cytoskeletal dynamics in Sertoli cells are regulated, which in turn supports spermatid transport across the seminiferous epithelium, and at the blood-testis barrier (BTB) during the epithelial cycle of spermatogenesis.
Assuntos
Citoesqueleto de Actina/efeitos dos fármacos , Proteínas do Citoesqueleto/metabolismo , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Proteínas dos Microfilamentos/metabolismo , Modelos Biológicos , Espermatogênese/efeitos dos fármacos , Animais , HumanosRESUMO
This study aims to use ice nucleation proteins (INPs) as a novel approach to improve the efficiency of freeze drying process and investigate the related mechanism of ice morphology. Our results show that INPs can significantly improve freeze drying efficiency with increased primary drying rate under the increase of INP concentration from 0 to 10-2mg/mL. Moreover, such improvement was more significant at higher subzero freezing temperatures with the addition of INPs, when the control samples were unable to freeze. Those improvements further lead to reduced total drying time, which suggests an estimated total energy saving of 28.5% by INPs. Our ice morphology results indicate the ability of INPs to alter ice morphology with lamellar ice structure and larger crystal size, which both show linear relationships with primary drying rate. The results further suggest that these ice morphology characteristics induced by INPs are very likely to facilitate the water vapor flow and improve the sublimation rate. Additionally, the increase of freeze drying efficiency can also be achieved by INPs in other food systems like coffee and milk with elevated primary drying rate. The results of this study suggest great potential of using INPs to improve the efficiency of freeze drying process for a wide range of food products and other related applications. This study also provides new insights into the relationship between process efficiency and ice morphology.
Assuntos
Proteínas da Membrana Bacteriana Externa , Dessecação , Manipulação de Alimentos/métodos , Liofilização/métodos , Congelamento , Gelo/análise , Água/química , Animais , Café , Eficiência , Humanos , Leite , VaporRESUMO
Freeze concentration is a separation process with high success in product quality. The remaining challenge is to achieve high efficiency with low cost. This study aims to evaluate the potential of using ice nucleation proteins (INPs) as an effective method to improve the efficiency of block freeze concentration while also exploring the related mechanism of ice morphology. Our results show that INPs are able to significantly improve the efficiency of block freeze concentration in a desalination model. Using this experimental system, we estimate that approximately 50% of the energy cost can be saved by the inclusion of INPs in desalination cycles while still meeting the EPA standard of drinking water (<500 ppm). Our investigative tools for ice morphology include optical microscopy and X-ray computed tomography imaging analysis. Their use indicates that INPs promote the development of a lamellar structured ice matrix with larger hydraulic diameters, which facilitates brine drainage and contains less brine entrapment as compared to control samples. These results suggest great potential for applying INPs to develop an energy-saving freeze concentration method via the alteration of ice morphology.