FM4-64,a kind of membrane-selective fluorescent dye,was utilized to stain the membrane structure of Botrytis cinerea and Phytophthora capsici.The effect of FM4-64 on hypha was observed under confocal microscope.The re...FM4-64,a kind of membrane-selective fluorescent dye,was utilized to stain the membrane structure of Botrytis cinerea and Phytophthora capsici.The effect of FM4-64 on hypha was observed under confocal microscope.The results indicated that the plasma membrane,speta and vesicles of the hyphae could be stained with FM4-64 at the concentrations of 1 to 20 μmol/L.The optimal dye concentration was 6 μmol/L for the both pathogens.At the same concentration of FM4-64,the staining of organelles was in a time-dependent manner and the different organelles were stained in diferrent times.The staining of plasma membrane and vesicles should be completed in 5 minutes,as other organelles can be stained after 10 minutes.展开更多
Multiphoton microscopy(MPM)is a powerful imaging technology for brain research.The imaging depth in MPM is partly determined by emission wavelength of fluorescent labels.It has been demonstrated that a longer emission...Multiphoton microscopy(MPM)is a powerful imaging technology for brain research.The imaging depth in MPM is partly determined by emission wavelength of fluorescent labels.It has been demonstrated that a longer emission wavelength is favorable for signal detection as imaging depth increases.However,there has been no comparison with near-infrared(NIR)emission.In order to quantitatively analyze the effect of emission wavelength on 3-photon imaging of mouse brains in vivo,we utilize the same excitation wavelength to excite a single fluorescent dye and simultaneously collect NIR and orange-red emission fluorescence at 828 nm and 620 nm,respectively.Both experimental and simulation results show that as the imaging depth increases,NIR emission decays less than orange-red fluorescent emission.These results show that it is preferable to shift the emission wavelength to NIR to enable more e±cient signal collection deep in the brain.展开更多
Tip growth is a common strategy for the rapid elongation of cells to forage the environment and/or to target to long-distance destinations. In the model tip growth system of Arabidopsis pollen tubes, several small- mo...Tip growth is a common strategy for the rapid elongation of cells to forage the environment and/or to target to long-distance destinations. In the model tip growth system of Arabidopsis pollen tubes, several small- molecule hormones regulate their elongation, but how these rapidly diffusing molecules control extremely localized growth remains mysterious. Here we show that the interconvertible salicylic acid （SA） and meth- ylated SA （MESA）, well characterized for their roles in plant defense, oppositely regulate Arabidopsis pollen tip growth with SA being inhibitory and MeSA stimulatory. The effect of SA and MeSA was independent of known NPR3/NPR4 SA receptor-mediated signaling pathways. SA inhibited clathrin-mediated endocytosis in pollen tubes associated with an increased accumulation of less stretchable demethylated pectin in the apical wall, whereas MeSA did the opposite. Furthermore, SA and MeSA alter the apical activation of ROP1 GTPase, a key regulator of tip growth in pollen tubes, in an opposite manner. Interestingly, both MeSA methylesterase and SA methyltransferase, which catalyze the interconversion between SA and MESA, are localized at the apical region of pollen tubes, indicating of the tip-localized production of SA and MeSA and consistent with their effects on the apical cellular activities. These findings suggest that local generation of a highly diffusible signal can regulate polarized cell growth, providing a novel mechanism of cell polarity control apart from the one involving protein and mRNA polarization.展开更多
The phragmoplast is a special apparatus that functions in establishing a cell plate in dividing plant cells. It is known that microfilaments (MFs) are involved in constituting phragmoplast structure, but the dynamic d...The phragmoplast is a special apparatus that functions in establishing a cell plate in dividing plant cells. It is known that microfilaments (MFs) are involved in constituting phragmoplast structure, but the dynamic distribution and role of phragmoplast MFs are far from being understood. In this study, the precise structure and dynamics of MFs during the initiation and the late lateral expansion of the phragmoplast were observed by using a tobacco BY-2 cell line stably expressing the microfilament reporter construct GFP-fABD2. Three-dimensional imaging showed that the phragmoplast MFs were initiated by two populations of MFs emerging between the reconstituting daughter nuclei at anaphase, which migrated to the mid-zone and gave rise to two layers of microfilament arrays. FM4-64 stained vesicles accumulated and fused with the cell plate between the two populations of MFs. The two layers of microfilament arrays of phragmoplast with ends overlapped always surrounded the centrifugally expanding cell plate. Partial disruption of MFs at metaphase by low concentration of latrunculin B resulted in the inhibition of the cell plate consolidation and the blockage of cell plate lateral expansion, whereas high concentration of latrunculin B restrained the progression of the cell cycle. Treating the cell after the initiation of phragmoplast led to the cease of the expansion of the cell plate. Our observations provide new insights into the precise structure and dynamics of phragmoplast MFs during cytokinesis and suggest that dynamic phragmoplast MFs are important in cell plate formation.展开更多
文摘FM4-64,a kind of membrane-selective fluorescent dye,was utilized to stain the membrane structure of Botrytis cinerea and Phytophthora capsici.The effect of FM4-64 on hypha was observed under confocal microscope.The results indicated that the plasma membrane,speta and vesicles of the hyphae could be stained with FM4-64 at the concentrations of 1 to 20 μmol/L.The optimal dye concentration was 6 μmol/L for the both pathogens.At the same concentration of FM4-64,the staining of organelles was in a time-dependent manner and the different organelles were stained in diferrent times.The staining of plasma membrane and vesicles should be completed in 5 minutes,as other organelles can be stained after 10 minutes.
基金work is funded by the National Natural Sci-ence Foundation of China(Grant/Award Numbers 62075135 and 61975126)Shenzhen Science and Technology Planning Project(ZDSYS2021-0623092006020)+2 种基金Key R&D Program of Shandong Province(Grant Number 2021CXGC010202)the Science and Technology Innovation Commission of Shenzhen(Grant/Award Numbers JCYJ201908-08174819083 and JCYJ20190808175201640)and Natural Science Foundation of Shandong Province(Grant Number ZR2022MA046)Major Innovation Projects for Integrating Science,Education&Industry of Qilu University of Technology(Shan-dong Academy of Sciences,Grant Number 2022JBZ01-04).
文摘Multiphoton microscopy(MPM)is a powerful imaging technology for brain research.The imaging depth in MPM is partly determined by emission wavelength of fluorescent labels.It has been demonstrated that a longer emission wavelength is favorable for signal detection as imaging depth increases.However,there has been no comparison with near-infrared(NIR)emission.In order to quantitatively analyze the effect of emission wavelength on 3-photon imaging of mouse brains in vivo,we utilize the same excitation wavelength to excite a single fluorescent dye and simultaneously collect NIR and orange-red emission fluorescence at 828 nm and 620 nm,respectively.Both experimental and simulation results show that as the imaging depth increases,NIR emission decays less than orange-red fluorescent emission.These results show that it is preferable to shift the emission wavelength to NIR to enable more e±cient signal collection deep in the brain.
文摘Tip growth is a common strategy for the rapid elongation of cells to forage the environment and/or to target to long-distance destinations. In the model tip growth system of Arabidopsis pollen tubes, several small- molecule hormones regulate their elongation, but how these rapidly diffusing molecules control extremely localized growth remains mysterious. Here we show that the interconvertible salicylic acid （SA） and meth- ylated SA （MESA）, well characterized for their roles in plant defense, oppositely regulate Arabidopsis pollen tip growth with SA being inhibitory and MeSA stimulatory. The effect of SA and MeSA was independent of known NPR3/NPR4 SA receptor-mediated signaling pathways. SA inhibited clathrin-mediated endocytosis in pollen tubes associated with an increased accumulation of less stretchable demethylated pectin in the apical wall, whereas MeSA did the opposite. Furthermore, SA and MeSA alter the apical activation of ROP1 GTPase, a key regulator of tip growth in pollen tubes, in an opposite manner. Interestingly, both MeSA methylesterase and SA methyltransferase, which catalyze the interconversion between SA and MESA, are localized at the apical region of pollen tubes, indicating of the tip-localized production of SA and MeSA and consistent with their effects on the apical cellular activities. These findings suggest that local generation of a highly diffusible signal can regulate polarized cell growth, providing a novel mechanism of cell polarity control apart from the one involving protein and mRNA polarization.
基金Supported by National Natural Science Foundation of China(Grant Nos.30630005,30470176)National Key Basic Research and Development Program of China(Grant Nos.2006CB100100,2007CB108700)
文摘The phragmoplast is a special apparatus that functions in establishing a cell plate in dividing plant cells. It is known that microfilaments (MFs) are involved in constituting phragmoplast structure, but the dynamic distribution and role of phragmoplast MFs are far from being understood. In this study, the precise structure and dynamics of MFs during the initiation and the late lateral expansion of the phragmoplast were observed by using a tobacco BY-2 cell line stably expressing the microfilament reporter construct GFP-fABD2. Three-dimensional imaging showed that the phragmoplast MFs were initiated by two populations of MFs emerging between the reconstituting daughter nuclei at anaphase, which migrated to the mid-zone and gave rise to two layers of microfilament arrays. FM4-64 stained vesicles accumulated and fused with the cell plate between the two populations of MFs. The two layers of microfilament arrays of phragmoplast with ends overlapped always surrounded the centrifugally expanding cell plate. Partial disruption of MFs at metaphase by low concentration of latrunculin B resulted in the inhibition of the cell plate consolidation and the blockage of cell plate lateral expansion, whereas high concentration of latrunculin B restrained the progression of the cell cycle. Treating the cell after the initiation of phragmoplast led to the cease of the expansion of the cell plate. Our observations provide new insights into the precise structure and dynamics of phragmoplast MFs during cytokinesis and suggest that dynamic phragmoplast MFs are important in cell plate formation.
