Pollen-expressed RLCKs control pollen tube burst.

In angiosperms, the pollen tube enters the receptive synergid cell, where it ruptures to release its cytoplasm along with two sperm cells. This interaction is complex, and the exact signal transducers that trigger the bursting of pollen tubes are not well understood. In this study, we identify three homologous receptor-like cytoplasmic kinases (RLCKs) expressed in pollen tubes of Arabidopsis, Delayed Burst 1/2/3 (DEB1/2/3), which play a crucial role in this process. These genes produce proteins localized on the plasma membrane, and their knockout causes delayed pollen tube burst and entrance of additional pollen tubes into the embryo sac due to fertilization recovery. We show that DEBs interact with the Ca2+ pump ACA9, influencing the dynamics of cytoplasmic Ca2+ in pollen tubes through phosphorylation. These results highlight the importance of DEBs as key signal transducers and the critical function of the DEB-ACA9 axis in timely pollen tube burst in synergids.

MAP3Kε1/2 Interact with MOB1A/1B and Play Important Roles in Control of Pollen Germination through Crosstalk with JA Signaling in Arabidopsis.

Restriction of pollen germination before the pollen grain is pollinated to stigma is essential for successful fertilization in angiosperms. However, the mechanisms underlying the process remain poorly understood. Here, we report functional characterization of the MAPKKK kinases, MAP3Kε1 and MAP3Kε2, involve in control of pollen germination in Arabidopsis. The two genes were expressed in different tissues with higher expression levels in the tricellular pollen grains. The map3kε1 map3kε2 double mutation caused abnormal callose accumulation, increasing level of JA and precocious pollen germination, resulting in significantly reduced seed set. Furthermore, the map3kε1 map3kε2 double mutations obviously upregulated the expression levels of genes in JA biosynthesis and signaling. The MAP3Kε1/2 interacted with MOB1A/1B which shared homology with the core components of Hippo singling pathway in yeast. The Arabidopsis mob1a mob1b mutant also exhibited a similar phenotype of precocious pollen germination to that in map3kε1 map3kε2 mutants. Taken together, these results suggested that the MAP3Kεs interacted with MOB1s and played important role in restriction of the precocious pollen germination, possibly through crosstalk with JA signaling and influencing callose accumulation in Arabidopsis.

The Arabidopsis AGC kinases NDR2/4/5 interact with MOB1A/1B and play important roles in pollen development and germination.

Pollen formation and pollen tube growth are essential for the delivery of male gametes into the female embryo sac for double fertilization. Little is known about the mechanisms that regulate the late developmental process of pollen formation and pollen germination. In this study, we characterized a group of Arabidopsis AGC kinase proteins, NDR2/4/5, involved in pollen development and pollen germination. The NDR2/4/5 genes are mainly expressed in pollen grains at the late developmental stages and in pollen tubes. They function redundantly in pollen formation and pollen germination. At the tricellular stages, the ndr2 ndr4 ndr5 mutant pollen grains exhibit an abnormal accumulation of callose, precocious germination and burst in anthers, leading to a drastic reduction in fertilization and a reduced seed set. NDR2/4/5 proteins can interact with another group of proteins (MOB1A/1B) homologous to the MOB proteins from the Hippo signaling pathway in yeast and animals. The Arabidopsis mob1a mob1b mutant pollen grains also have a phenotype similar to that of ndr2 ndr4 ndr5 pollen grains. These results provide new evidence demonstrating that the Hippo signaling components are conserved in plants and play important roles in sexual plant reproduction.

Protective effect of ulinastatin on hepatic ischemia reperfusion injury through autophagy activation in Chang liver cells.

This study aimed to investigate the protective effect of ulinastatin in hepatic ischemia-reperfusion progress, involving its association with the role of autophagy during hypoxia-induced hypoxia-reoxygenation injury in vitro. The model of hepatic hypoxia/reoxygenation (H/R) injury in Chang liver cells was established. After treatment with ulinastatin at the doses of 10, 100, and 1000 U/mL in H/R liver cells, the cell proliferation was significantly increased, morphological damage was reduced, and the cell apoptosis rate was decreased. The protein levels of antiapoptotic myeloid cell leukemia-1 (Mcl-1) and caspase-3 were upregulated, and C-PARP protein was downregulated. Meanwhile, ulinastatin led to an increase in the messenger RNA and protein levels of autophagy maker Unc-like kinase 1 (ULK1), Beclin-1, and microtubule-associated protein 1 light chain 3 (LC-3) and a decrease in p62. Then, 3-methyladenine (3-MA), an inhibitor of autophagy, made morphological damage and cell apoptosis worsen in ulinastatin-treated H/R liver cells. And the expression levels of caspase-3, C-PARP, p62, Beclin-1, and LC-3, proteins were also reversed by 3-MA. Taken together, our results demonstrate that ulinastatin inhibited the hepatic H/R injury in Chang liver cells, which was, to some extent, related to the autophagy activation.

The predictive value of serial changes in diaphragm function during the spontaneous breathing trial for weaning outcome: a study protocol.

INTRODUCTION: There is a variety of tools being used in clinical practice for the prediction of weaning success from mechanical ventilation. However, their diagnostic performances are less than satisfactory. The purpose of this study is to investigate the value of serial changes in diaphragm function measured by ultrasound during the spontaneous breathing trial (SBT) as a weaning predictor. METHODS AND ANALYSIS: This is a prospective observational study conducted in a 10-bed medical emergency intensive care unit (EICU) in a university-affiliated hospital. The study will be performed from November 2016 to December 2017. All patients in the EICU who are expected to have mechanical ventilation for more than 48 hours through endotracheal tube are potentially eligible for this study. Patients will be included if they fulfil the criteria for SBT. All enrolled patients will be ventilated with an Evita-4 by using volume assist control mode prior to SBT. Positive end-expiratory pressure (PEEP) will be set to 5 cmH2O and fractional inspired oxygen (FiO2) will be set to a value below 0.5 that guarantees oxygen saturation by pulse oximetry (SpO2) greater than 90%. Enrolled patients will undergo SBT for 2 hours in semirecumbent position. During the SBT, the patients will breathe through the ventilator circuit by using flow triggering (2 L/min) with automatic tube compensation of 100% and 5 cmH2O PEEP. The FiO2 will be set to the same value as used before SBT. If the patients fail to tolerate the SBT, the trial will be discontinued immediately and the ventilation mode will be switched to that used before the trial. Patients who pass the 2-hour SBT will be extubated. Right diaphragm excursion and bilateral diaphragm thickening fraction will be measured by ultrasonography during spontaneous breathing. Images will be obtained immediately prior to the SBT, and at 5, 30, 60, 90 and 120 min after the initiation of SBT. Rapid shallow breathing index will be simultaneously calculated at the bedside by a respiratory nurse. ETHICS AND DISSEMINATION: The study protocol is approved by the ethics committee of Sir Run Run Shaw Hospital, an affiliate of Zhejiang University, Medical College. The results will be published in a peer-reviewed journal and shared with the worldwide medical community. TRIAL REGISTRATION NUMBER: ISRCTN42917473; Pre-results.

The THERMOSENSITIVE MALE STERILE 1 Interacts with the BiPs via DnaJ Domain and Stimulates Their ATPase Enzyme Activities in Arabidopsis.

The Arabidopsis TMS1 encodes a heat shock protein identical to the Hsp40 protein AtERdj3A and plays important roles in the thermotolerance of pollen tubes and other plant tissues. Despite its importance to plant growth and reproduction, little has been known about its mechanisms underlying thermotolerance of plants. In this study, the relationship between TMS1 and the Hsp70 proteins, Binding Immunoglobulin Proteins (BiPs) was explored to understand the molecular mechanisms of TMS1 in thermotolerance of plants. The expression of TMS1 was induced not only by heat shock, but also by dithiothreitol (DTT) and L-azetidine-2-carboxylic acid (AZC), similarly to the three BiP genes, indicating that TMS1 may be involved in unfolded protein response (UPR). The firefly luciferase complementary imaging (LCI), GST pull-down and ATPase enzyme activity assays demonstrated that the DnaJ domain of TMS1 could interact with BiP1 and BiP3, and could stimulate their ATPase enzyme activities. In addition, the expression level of TMS1 was reduced in the bzip28 bzip60 double mutant. These results suggest that TMS1 may function at the downstream of bZIP28 and bZIP60 and be involved in termotolerance of plants, possibly by participating in refolding or degradation of unfolded and misfolded proteins through interaction with the BiPs.