基于氘示踪剂和热扩散技术的栓皮栎水分运输速率与效率研究

利用同位素示踪和热扩散技术研究了不同胁迫处理栓皮栎的水分运输和储存差异。结果表明,注射氘同位素后,充分灌溉、轻度和重度胁迫处理的最大氘同位素比率分别升高到586.67‰、997.33‰和1364.89‰,处理间差异显著。轻度和重度胁迫处理的示踪速率分别为0.10 m/h和0.07 m/h,显著低于充分灌溉处理,但半减期和残留期显著高于充分灌溉。轻度和重度胁迫处理的枝条栓塞程度(PLC)比充分灌溉显著增加,液流通量、水势和蒸腾速率则显著减小。统计分析表明蒸腾作用强弱决定树体水分运输速率,PLC的增加和枝条水势的降低阻碍木质部水分运输。半减期和残留期内,轻度和重度胁迫处理的累积液流量显著高于充分灌水处理,运载相同体积的示踪剂,胁迫处理栓皮栎需要的水量增加,表明胁迫环境下受到PLC、蒸腾以及与树体储水交换的影响,水分运输效率下降。栓皮栎通过栓塞和储水交换来降低水分运输速率和效率,调控水分的收支平衡来适应干旱的环境。     

Haloalkylphosphorus Hydrolases Purified from Sphingomonas sp. Strain TDK1 and Sphingobium sp. Strain TCM1

Phosphotriesterases catalyze the first step of organophosphorus triester degradation. The bacterial phosphotriesterases purified and characterized to date hydrolyze mainly aryl dialkyl phosphates, such as parathion, paraoxon, and chlorpyrifos. In this study, we purified and cloned two novel phosphotriesterases from Sphingomonas sp. strain TDK1 and Sphingobium sp. strain TCM1 that hydrolyze tri(haloalkyl)phosphates, and we named these enzymes haloalkylphosphorus hydrolases (TDK-HAD and TCM-HAD, respectively). Both HADs are monomeric proteins with molecular masses of 59.6 (TDK-HAD) and 58.4 kDa (TCM-HAD). The enzyme activities were affected by the addition of divalent cations, and inductively coupled plasma mass spectrometry analysis suggested that zinc is a native cofactor for HADs. These enzymes hydrolyzed not only chlorinated organophosphates but also a brominated organophosphate [tris(2,3-dibromopropyl) phosphate], as well as triaryl phosphates (tricresyl and triphenyl phosphates). Paraoxon-methyl and paraoxon were efficiently degraded by TCM-HAD, whereas TDK-HAD showed weak activity toward these substrates. Dichlorvos was degraded only by TCM-HAD. The enzymes displayed weak or no activity against trialkyl phosphates and organophosphorothioates. The TCM-HAD and TDK-HAD genes were cloned and found to encode proteins of 583 and 574 amino acid residues, respectively. The primary structures of TCM-HAD and TDK-HAD were very similar, and the enzymes also shared sequence similarity with fenitrothion hydrolase (FedA) of Burkholderia sp. strain NF100 and organophosphorus hydrolase (OphB) of Burkholderia sp. strain JBA3. However, the substrate specificities and quaternary structures of the HADs were largely different from those of FedA and OphB. These results show that HADs from sphingomonads are novel members of the bacterial phosphotriesterase family.     

Cloning and expression of carp acetylcholinesterase gene in Pichia pastoris and characterization of the recombinant enzyme

The gene encoding acetylcholinesterase (AChE) was cloned from common carp muscle tissue. The full-length cDNA was 2368 bp that contains a coding region of 1902 bp, corresponding to a protein of 634 amino acids. The deduced amino acid sequence showed a significant homology with those of ichthyic AChEs and several common features among them, including T peptide encoded by exon T in the C-terminus. Three yeast expression vectors were constructed and introduced into the yeast Pichia pastoris. The transformant harboring carp AChE gene lacking exon T most effectively produced AChE activity extracellularly. The replacement of the native signal sequence with the yeast α-factor prepro signal sequence rather decreased the production. A decrease in cultivation temperature from 30 to 15 °C increased the activity production 32.8-fold. The purified recombinant AChE lacking T peptide, eluted as a single peak with a molecular mass of about 230 kDa on the gel filtration chromatography, exhibited the specific activity of 4970 U/mg. On the SDS–PAGE, three proteins with molecular masses of 73, 54, and 22 kDa were observed. These proteins were N-glycosylated, and their N-terminal sequence showed that the latter two were produced from the former probably by proteolytic cleavage at the C-terminal region. Thus, the recombinant AChE is homotrimer of three identical subunits with 73 kDa. The optimal temperature and pH of the recombinant were comparable to those of the native enzyme purified previously, but the values of kinetic parameters and the sensitivities to substrate inhibition and inhibitors were considerably different between them.     

Hypoxic glioma-derived exosomes promote M2-like macrophage polarization by enhancing autophagy induction

Exosomes participate in intercellular communication and glioma microenvironment modulation, but the exact mechanisms by which glioma-derived exosomes (GDEs) promote the generation of the immunosuppressive microenvironment are still unclear. Here, we investigated the effects of GDEs on autophagy, the polarization of tumor-associated macrophages (TAMs), and glioma progression. Compared with normoxic glioma-derived exosomes (N-GDEs), hypoxic glioma-derived exosomes (H-GDEs) markedly facilitated autophagy and M2-like macrophage polarization, which subsequently promoted glioma proliferation and migration in vitro and in vivo. Western blot and qRT-PCR analyses indicated that interleukin 6 (IL-6) and miR-155-3p were highly expressed in H-GDEs. Further experiments showed that IL-6 and miR-155-3p induced M2-like macrophage polarization via the IL-6-pSTAT3-miR-155-3p-autophagy-pSTAT3 positive feedback loop, which promotes glioma progression. Our study clarifies a mechanism by which hypoxia and glioma influence autophagy and M2-like macrophage polarization via exosomes, which could advance the formation of the immunosuppressive microenvironment. Our findings suggest that IL-6 and miR-155-3p may be novel biomarkers for diagnosing glioma and that treatments targeting autophagy and the STAT3 pathway may contribute to antitumor immunotherapy.Subject terms: Cancer microenvironment, Autophagy     

Condition factor dependency of burst swimming ability between wild and hatchery-reared chum salmon fry (Oncorhynchus keta)

Ichthyological Research - Burst swimming velocity (Uburst) was compared between wild and hatchery-reared chum salmon fry. In the hatchery-reared fry, Uburst was significantly correlated with the...