Arabidopsis class A S-acyl transferases modify the pollen receptors LIP1 and PRK1 to regulate pollen tube guidance.

Protein S-acylation catalyzed by protein S-acyl transferases (PATs) is a reversible lipid modification regulating protein targeting, stability, and interaction profiles. PATs are encoded by large gene families in plants, and many proteins including receptor-like cytoplasmic kinases (RLCKs) and receptor-like kinases (RLKs) are subject to S-acylation. However, few PATs have been assigned substrates, and few S-acylated proteins have known upstream enzymes. We report that Arabidopsis (Arabidopsis thaliana) class A PATs redundantly mediate pollen tube guidance and participate in the S-acylation of POLLEN RECEPTOR KINASE1 (PRK1) and LOST IN POLLEN TUBE GUIDANCE1 (LIP1), a critical RLK or RLCK for pollen tube guidance, respectively. PAT1, PAT2, PAT3, PAT4, and PAT8, collectively named PENTAPAT for simplicity, are enriched in pollen and show similar subcellular distribution. Functional loss of PENTAPAT reduces seed set due to male gametophytic defects. Specifically, pentapat pollen tubes are compromised in directional growth. We determine that PRK1 and LIP1 interact with PENTAPAT, and their S-acylation is reduced in pentapat pollen. The plasma membrane (PM) association of LIP1 is reduced in pentapat pollen, whereas point mutations reducing PRK1 S-acylation affect its affinity with its interacting proteins. Our results suggest a key role of S-acylation in pollen tube guidance through modulating PM receptor complexes.

The canonical α-SNAP is essential for gametophytic development in Arabidopsis.

The development of male and female gametophytes is a pre-requisite for successful reproduction of angiosperms. Factors mediating vesicular trafficking are among the key regulators controlling gametophytic development. Fusion between vesicles and target membranes requires the assembly of a fusogenic soluble N-ethylmaleimide sensitive factor attachment protein receptors (SNAREs) complex, whose disassembly in turn ensures the recycle of individual SNARE components. The disassembly of post-fusion SNARE complexes is controlled by the AAA+ ATPase N-ethylmaleimide-sensitive factor (Sec18/NSF) and soluble NSF attachment protein (Sec17/α-SNAP) in yeast and metazoans. Although non-canonical α-SNAPs have been functionally characterized in soybeans, the biological function of canonical α-SNAPs has yet to be demonstrated in plants. We report here that the canonical α-SNAP in Arabidopsis is essential for male and female gametophytic development. Functional loss of the canonical α-SNAP in Arabidopsis results in gametophytic lethality by arresting the first mitosis during gametogenesis. We further show that Arabidopsis α-SNAP encodes two isoforms due to alternative splicing. Both isoforms interact with the Arabidopsis homolog of NSF whereas have distinct subcellular localizations. The presence of similar alternative splicing of human α-SNAP indicates that functional distinction of two α-SNAP isoforms is evolutionarily conserved.

Implementing a completely autotrophic nitrogen removal over nitrite process using a novel umbrella basalt fiber carrier.

The completely autotrophic nitrogen removal over nitrite (CANON) process is significantly hindered by prolonged start-up periods and unstable nitrogen removal efficiency. In this study, a novel umbrella basalt fiber (BF) carrier with good biological affinity and adsorption performance was used to initiate the CANON process. The CANON process was initiated on day 64 in a sequencing batch reactor equipped with umbrella BF carriers. During this period, the influent NH4+-N concentration gradually increased from 100 to 200 mg·L-1, and the dissolved oxygen was controlled below 0.8 mg L-1. Consequently, an average ammonia nitrogen removal efficiency (ARE) and total nitrogen removal efficiency (TNRE) of ∼90 and 80% were achieved, respectively. After 130 days, ARE and TNRE remained stable at 92 and 81.1%, respectively. This indicates a reliable method for achieving rapid start-up and stable operation of the CANON process. Moreover, Candidatus Kuenenia and Candidatus Brocadia were identified as dominant anammox genera on the carrier. Nitrosomonas was the predominant genus among ammonia-oxidizing bacteria. Spatial differences were observed in the microbial population of umbrella BF carriers. This arrangement facilitated autotrophic nitrogen removal in a single reactor. This study indicates that the novel umbrella BF carrier is a highly suitable biocarrier for the CANON process.

Stress response and tolerance mechanisms of NaHCO3 exposure based on biochemical assays and multi-omics approach in the liver of crucian carp (Carassius auratus).

The development and utilization of saline-alkaline water, an important backup resource, has received widespread attention. However, the underuse of saline-alkaline water, threatened by the single species of saline-alkaline aquaculture, seriously affects the development of the fishery economy. In this work, a 30-day NaHCO3 stress experimental study combined with analyses of untargeted metabolomics, transcriptome, and biochemical approaches was conducted on crucian carp to provide a better understanding of the saline-alkaline stress response mechanism in freshwater fish. This work revealed the relationships among the biochemical parameters, endogenous differentially expressed metabolites (DEMs), and differentially expressed genes (DEGs) in the crucian carp livers. The biochemical analysis showed that NaHCO3 exposure changed the levels of several physiological parameters associated with the liver, including antioxidant enzymes (SOD, CAT, GSH-Px), MDA, AKP, and CPS. According to the metabolomics study, 90 DEMs are involved in various metabolic pathways such as ketone synthesis and degradation metabolism, glycerophospholipid metabolism, arachidonic acid metabolism, and linoleic acid metabolism. In addition, transcriptomics data analysis showed that a total of 301 DEGs were screened between the control group and the high NaHCO3 concentration group, of which 129 up-regulated genes and 172 down-regulated genes. Overall, NaHCO3 exposure could cause lipid metabolism disorders and induce energy metabolism imbalance in the crucian carp liver. Simultaneously, crucian carp might regulate its saline-alkaline resistance mechanism by enhancing the synthesis of glycerophospholipid metabolism, ketone bodies, and degradation metabolism, at the same time increasing the vitality of antioxidant enzymes (SOD, CAT, GSH-Px) and nonspecific immune enzyme (AKP). Herein, all results will provide new insights into the molecular mechanisms underlying the stress responses and tolerance to saline-alkaline exposure in crucian carp.

Arabidopsis ADP-RIBOSYLATION FACTOR-A1s mediate tapetum-controlled pollen development.

Propagation of angiosperms mostly relies on sexual reproduction, in which gametophytic development is a pre-requisite. Male gametophytic development requires both gametophytic and sporophytic factors, most importantly early secretion and late programmed cell death of the tapetum. In addition to transcriptional factors, proteins at endomembrane compartments, such as receptor-like kinases and vacuolar proteases, control tapetal function. The cellular machinery that regulates their distribution is beginning to be revealed. We report here that ADP-RIBOSYLATION FACTOR-A1s (ArfA1s) are critical for tapetum-controlled pollen development. All six ArfA1s in the Arabidopsis genome are expressed during anther development, among which ArfA1b is specific to the tapetum and developing microspores. Although the ArfA1b loss-of-function mutant showed no pollen defects, probably due to redundancy, interference with ArfA1s by a dominant negative approach in the tapetum resulted in tapetal dysfunction and pollen abortion. We further showed that all six ArfA1s are associated with the Golgi and the trans-Golgi network/early endosome, suggesting that they have roles in regulating post-Golgi trafficking to the plasma membrane or to vacuoles. Indeed, we demonstrated that the expression of ArfA1bDN interfered with the targeting of proteins critical for tapetal development. The results presented here demonstrate a key role of ArfA1s in tapetum-controlled pollen development by mediating protein targeting through post-Golgi trafficking routes.

Rapid cultivation of anammox sludge based on Ca-alginate cell beads.

Current gel entrapment technology has certain advantages for the enrichment of anammox sludge. In this study, the optimal preparation conditions and cultivation equipment of Ca-alginate cell beads for the culturing anammox sludge were proposed. The preparation parameters of the Ca-alginate cell beads were as follows: 3% sodium alginate, 4% CaCl2, VSA:Vcell = 1:1, a drop height of 9 cm, stirring speed of 300 rpm, and cross-linking time of 24 h. The prepared cell beads were regular spheres with a uniform size and hard texture. Throughout the 9 days of cultivation, the number of anammox bacteria in the Ca-alginate cell beads was 4.3 times that of the initial sludge, and the color of the cell beads changed from yellowish-brown to reddish-brown. Scanning electron microscopy (SEM) analysis showed that the SA gel beads had a good microporous structure. The fluorescence in situ hybridization (FISH) results illustrated that the bacteria were mostly dispersed inside the Ca-alginate cell beads. Additionally, the qPCR results implied that only a relatively small amount of anammox biomass (2.74×106 copies/gel-bead) was required to quickly start the anammox process. The anammox bacteria in the Ca-alginate cell beads grew with a fast growth rate in a short period and exhibited high activity due to diffusion limitations. In addition, the anammox bacteria cultivated in the Ca-alginate cell beads could adapt to the increase in substrate concentration in a short period. The optimal incubation time of this gel entrapment method for anammox sludge was no more than 17 days under the experimental conditions of this work. Therefore, this simple and practicable gel entrapment method may serve as a suitable pre-culture means for the rapid enrichment of anammox bacteria.

COPII Components Sar1b and Sar1c Play Distinct Yet Interchangeable Roles in Pollen Development.

The development of pollen is a prerequisite for double fertilization in angiosperms. Coat protein complex II (COPII) mediates anterograde transport of vesicles from the endoplasmic reticulum to the Golgi. Components of the COPII complex have been reported to regulate either sporophytic or gametophytic control of pollen development. The Arabidopsis (Arabidopsis thaliana) genome encodes five Sar1 isoforms, the small GTPases essential for COPII formation. By using a dominant negative approach, Sar1 isoforms were proposed to have distinct cargo specificity despite their sequence similarity. Here, we examined the functions of three Sar1 isoforms through analysis of transfer DNA insertion mutants and CRISPR/Cas9-generated mutants. We report that functional loss of Sar1b caused malfunction of tapetum, leading to male sterility. Ectopic expression of Sar1c could compensate for Sar1b loss of function in sporophytic control of pollen development, suggesting that they are interchangeable. Functional distinction between Sar1b and Sar1c may have resulted from their different gene transcription levels based on expression analyses. On the other hand, Sar1b and Sar1c redundantly mediate male gametophytic development such that the sar1b;sar1c microspores aborted at anther developmental stage 10. This study uncovers the role of Sar1 isoforms in both sporophytic and gametophytic control of pollen development. It also suggests that distinct functions of Sar1 isoforms may be caused by their distinct transcription programs.

A biocompatible electrode/exoelectrogens interface augments bidirectional electron transfer and bioelectrochemical reactions.

Bidirectional electron transfer is about that exoelectrogens produce bioelectricity via extracellular electron transfer at anode and drive cytoplasmic biochemical reactions via extracellular electron uptake at cathode. The key factor to determine above bioelectrochemical performances is the electron transfer efficiency under biocompatible abiotic/biotic interface. Here, a graphene/polyaniline (GO/PANI) nanocomposite electrode specially interfacing exoelectrogens (Shewanella loihica) and augmenting bidirectional electron transfer was conducted by in-situ electrochemical modification on carbon paper (CP). Impressively, the GO/PANI@CP electrode tremendously improved the performance of exoelectrogens at anode for wastewater treatment and bioelectricity generation (about 54 folds increase of power density compared to blank CP electrode). The bacteria on electrode surface not only showed fast electron release but also exhibited high electricity density of extracellular electron uptake through the proposed direct electron transfer pathway. Thus, the cathode applications of microbial electrosynthesis and bio-denitrification were developed via GO/PANI@CP electrode, which assisted the close contact between microbial outer-membrane cytochromes and nanocomposite electrode for efficient nitrate removal (0.333 mM/h). Overall, nanocomposite modified electrode with biocompatible interfaces has great potential to enhance bioelectrochemical reactions with exoelectrogens.

Arabidopsis Sar1 isoforms play redundant roles in female gametophytic development.

KEY MESSAGE: Functional loss of Arabidopsis Sar1b with that of either Sar1a or Sar1c inhibits mitosis of functional megaspores, leading to defective embryo sac formation and reduced fertility. Vesicular trafficking among diverse endomembrane compartments is critical for eukaryotic cells. Anterograde trafficking from endoplasmic reticulum (ER) to the Golgi apparatus is mediated by coat protein complex II (COPII) vesicles. Among five cytosolic components of COPII, secretion-associated Ras-related GTPase 1 (Sar1) mediates the assembly and disassembly of the COPII coat. Five genes in Arabidopsis encode Sar1 isoforms, whose different cargo specificities and redundancy were both reported. We show here that Arabidopsis Sar1a, Sar1b, and Sar1c mediate the development of female gametophytes (FGs), in which Sar1b plays a major role, whereas Sar1a and Sar1c play a minor role. We determined that female transmission of sar1a;sar1b or sar1c;sar1b was significantly reduced due to defective mitosis of functional megaspores. Half of ovules in sar1a;sar1b/+ or sar1c;sar1b/+ plants failed to attract pollen tubes, leading to fertilization failure. The homozygous sar1a;sar1b or sar1c;sar1b double mutant was obtained by introducing either UBQ10:GFP-Sar1b or UBQ10:GFP-Sar1c, supporting their redundant function in FG development.

Signaling network controlling ROP-mediated tip growth in Arabidopsis and beyond.

Cell polarity operates across a broad range of spatial and temporal scales and is essential for specific biological functions of polarized cells. Tip growth is a special type of polarization in which a single and unique polarization site is established and maintained, as for the growth of root hairs and pollen tubes in plants. Extensive studies in past decades have demonstrated that the spatiotemporal localization and activity of Rho of Plants (ROPs), the only class of Rho GTPases in plants, are critical for tip growth. ROPs are switched on or off by different factors to initiate dynamic intracellular activities, leading to tip growth. Recent studies have also uncovered several feedback modules for ROP signaling. In this review, we summarize recent progress on ROP signaling in tip growth, focusing on molecular mechanisms that underlie the dynamic distribution and activity of ROPs in Arabidopsis. We also highlight feedback modules that control ROP-mediated tip growth and provide a perspective for building a complex ROP signaling network. Finally, we provide an evolutionary perspective for ROP-mediated tip growth in Physcomitrella patens and during plant-rhizobia interaction.

Arabidopsis RAN GTPases are critical for mitosis during male and female gametogenesis.

The development of male and female gametophytes is a prerequisite for successful propagation of angiosperms. The small GTPases RAN play fundamental roles in numerous cellular processes. Although RAN GTPases have been characterized in plants, their roles in cellular processes are far from understood. We report here that RAN GTPases in Arabidopsis are critical for gametophytic development. RAN1 loss-of-function showed no defects in gametophytic development likely due to redundancy. However, the expression of a dominant negative or constitutively active RAN1 resulted in gametophytic lethality. Genetic interference of RAN GTPases caused the arrest of pollen mitosis I and of mitosis of functional megaspores, implying a key role of properly regulated RAN activity in mitosis during gametophytic development.

Chitin degradation and electricity generation by Aeromonas hydrophila in microbial fuel cells.

Chitin is one of the most abundant biopolymers in nature and the main composition of shrimp and crab shells (usually as food wastes). Thus it is essential to investigate the potential of degrading chitin for energy recovery. This study investigated the anaerobic degradation of chitin by Aeromonas hydrophila, a chitinolytic and popular electroactive bacterium, in both fermentation and microbial fuel cell (MFC) systems. The primary chitin metabolites produced in MFC were succinate, lactate, acetate, formate, and ethanol. The total metabolite concentration from chitin degradation increased seven-fold in MFC compared to the fermentation system, as well as additional electricity generation. Moreover, A. hydrophila degraded GlcNAc (the intermediate of chitin hydrolysis) significantly faster (0.97 and 0.94 mM C/d/mM-GlcNAc) than chitin (0.13 and 0.03 mM C/d/mM-GlcNAc) in MFC and fermentation systems, indicating that extracellular hydrolysis of chitin was the rate-limiting step and this step could be accelerated in MFC. Furthermore, more chemicals produced by the addition of exogenous mediators in MFC. This study proves that the chitin could be degraded effectively by an electroactive bacterium in MFC, and our results suggest that this bioelectrochemical system might be useful for the degradation of recalcitrant biomass to recover energy.

Silver nanoparticles formation by extracellular polymeric substances (EPS) from electroactive bacteria.

Microbial extracellular polymeric substances (EPS) excreted from microorganisms were a complex natural biological polymer mixture of proteins and polysaccharides, which played an important roles in the transport of metals, such as Ag(+). Electroactive bacteria, is an important class of environmental microorganisms, which can use iron or manganese mineral as terminal electron acceptors to generate energy for biosynthesis and cell maintenance. In this work, the EPS extracted of three electroactive bacteria (Shewanella oneidensis, Aeromonas hydrophila, and Pseudomonas putida) were used for reducing Ag(+) and forming silver nanoparticles (AgNPs). Results showed that all the three microbial EPS could reduce Ag(+) to AgNPs. The formed AgNPs were characterized in depth by the UV-visible spectroscopy, X-ray diffraction, X-ray photoelectron spectroscopy, and high-resolution transmission electron microscopy. The main components in the EPS from the three electroactive bacteria were analyzed. The presence of cytochrome c in these EPS was confirmed, and they were found to contribute to the reduction of Ag(+) to AgNPs. The results indicated that the EPS of electroactive bacteria could act as a reductant for AgNPs synthesis and could provide new information to understand the fate of metals and their metal nanoparticles in the natural environments.

Redox properties of extracellular polymeric substances (EPS) from electroactive bacteria.

Although the capacity for electroactive bacteria to convert environmental metallic minerals and organic pollutants is well known, the role of the redox properties of microbial extracellular polymeric substances (EPS) in this process is poorly understood. In this work, the redox properties of EPS from two widely present electroactive bacterial strains (Shewanella oneidensis and Pseudomonas putida) were explored. Electrochemical analysis demonstrates that the EPS extracted from the two strains exhibited redox properties. Spectroelectrochemical and protein electrophoresis analyses indicate that the extracted EPS from S. oneidensis and P. putida contained heme-binding proteins, which were identified as the possible redox components in the EPS. The results of heme-mediated behavior of EPS may provide an insight into the important roles of EPS in electroactive bacteria to maximize their redox capability for biogeochemical cycling, environmental bioremediation and wastewater treatment.

Iron reduction in the DAMO/Shewanella oneidensis MR-1 coculture system and the fate of Fe(II).

Dissimilatory iron reduction and anaerobic methane oxidation processes play important roles in the global iron and carbon cycle, respectively. This study explored the ferrihydrite reduction process with methane as a carbon source in a coculture system of denitrifying anaerobic methane oxidation (DAMO) microbes enriched in laboratory and Shewanella oneidensis MR-1, and then characterized the reduced products. Ferrihydrite reduction was also studied in the DAMO and Shewanella systems alone. The ferrihydrite was reduced slightly (<13.3%) in the separate systems, but greatly (42.0-88.3%) in the coculture system. Isotope experiment of (13)CH4 addition revealed that DAMO microbes coupled to S. oneidensis MR-1 in a ferric iron reduction process with (13)CH4 consumption and (13)CO2 production. Compared with ferrihydrite, the reduced products showed increased crystallinity (from amorphous state to crystallinity 77.1%) and magnetism (from paramagnetic to ferromagnetic). The produced ferrous iron was formed into minerals primarily composed of siderite with a small amount vivianite and magnetite. A portion of products covered the cell surface and hindered further reactions. The results presented herein widen the current understanding of iron metabolism and mineralization in the ocean, and show that the coculture systems of DAMO microbes and Shewanella have the potential to be globally important to iron reduction and methane oxidation.