Nucleation and growth of cadherin adhesions

Cell-cell contact formation relies on the recruitment of cadherin molecules and their anchoring to actin. However, the precise chronology of events from initial cadherin trans-interactions to adhesion strengthening is unclear, in part due to the lack of access to the distribution of cadherins within adhesion zones. Using N-cadherin expressing cells interacting with N-cadherin coated surfaces, we characterized the formation of cadherin adhesions at the ventral cell surface. TIRF and RIC microscopies revealed streak-like accumulations of cadherin along actin fibers. FRAP analysis indicated that engaged cadherins display a slow turnover at equilibrium, compatible with a continuous addition and removal of cadherin molecules within the adhesive contact. Association of cadherin cytoplasmic tail to actin as well as actin cables and myosin II activity are required for the formation and maintenance of cadherin adhesions. Using time lapse microscopy we deciphered how cadherin adhesions form and grow. As lamellipodia protrude, cadherin foci stochastically formed a few microns away from the cell margin. Neo-formed foci coalesced aligned and coalesced with preformed foci either by rearward sliding or gap filling to form cadherin adhesions. Foci experienced collapse at the rear of cadherin adhesions. Based on these results, we present a model for the nucleation, directional growth and shrinkage of cadherin adhesions.     

Bi-directional transport of GLUT4 vesicles near the plasma membrane of primary rat adipocytes

Insulin stimulates glucose uptake into adipocytes by mobilizing intracellular membrane vesicles containing GLUT4 proteins to the plasma membrane. Here we applied time-lapse total internal reflection fluorescence microscopy to study moving parameters and characters of exogenously expressed GLUT4 vesicles in basal, insulin and nocodazole treated primary rat adipocytes. Our results showed that microtubules were essential for long-range transport of GLUT4 vesicles but not obligatory for GLUT4 distribution in rat adipocytes. Insulin reduced the mobility of the vesicles, made them tethered/docked to the PM and finally had constitutive exocytosis. Moreover, long-range bi-directional movements of GLUT4 vesicles were visualized for the first time by TIRFM. It is likely that there are interactions between insulin signaling and microtubules, to regulating GLUT4 translocation in rat adipocytes.     

A micelle nucleation model for the interaction of dodecyl sulphate with Lys49-phospholipases A(2)

Bothropstoxin-I (BthTx-I) is a Lys49-PLA(2) from the venom of Bothrops jararacussu that lacks detectable catalytic activity, yet causes rapid Ca(2+)-independent membrane damage. With the aim of understanding the interaction between BthTx-I and amphiphilic molecules, we have studied the interaction of sodium dodecyl sulphate (SDS) with the protein. Circular dichroism and attenuated total reflection Fourier-transform infrared spectra of BthTx-I reveal changes in the alpha-helical organization of the protein at an SDS/BthTx-I molar ratio of 20-25. At SDS/BthTx-I ratios of 40-45 the alpha-helices return to a native-like conformation, although fluorescence emission anisotropy measurements of 2-amino-N-hexadecyl-benzamide (AHBA) demonstrate that the total SDS is below the critical micelle concentration when this transition occurs. These results may be interpreted as the result of SDS accumulation by the BthTx-I homodimer and the formation of a pre-micelle SDS/BthTx-I complex, which may subsequently be released from the protein surface as a free micelle. Similar changes in the alpha-helical organization of BthTx-I were observed in the presence of dipalmitoylphosphatidylcholine liposomes, suggesting that protein structure transitions coupled to organization changes of bound amphiphiles may play a role in the Ca(2+)-independent membrane damage by Lys49-PLA(2)s.     

Real-time Imaging of Plant Cell Surface Dynamics with Variable-angle Epifluorescence Microscopy

A plant’s cell surface is its interface for perceiving environmental cues; it responds with cell biological changes such as membrane trafficking and cytoskeletal rearrangement. Real-time and high-resolution image analysis of such intracellular events will increase the understanding of plant cell biology at the molecular level. Variable angle epifluorescence microscopy (VAEM) is an emerging technique that provides high-quality, time-lapse images of fluorescently-labeled proteins on the plant cell surface. In this article, practical procedures are described for VAEM specimen preparation, adjustment of the VAEM optical system, movie capturing and image analysis. As an example of VAEM observation, representative results are presented on the dynamics of PATROL1. This is a protein essential for stomatal movement, thought to be involved in proton pump delivery to plasma membranes in the stomatal complex of Arabidopsis thaliana. VAEM real-time observation of guard cells and subsidiary cells in A. thaliana cotyledons showed that fluorescently-tagged PATROL1 appeared as dot-like structures on plasma membranes for several seconds and then disappeared. Kymograph analysis of VAEM movie data determined the time distribution of the presence (termed ‘residence time’) of the dot-like structures. The use of VAEM is discussed in the context of this example.     

Optics of the ultraviolet reflecting scales of a jumping spider

The jumping spider Cosmophasis umbratica from Singapore is strongly sexually dimorphic. The males, but not the females, reflect ultraviolet as well as green-orange light. The scales responsible for this are composed of a chitin-air-chitin sandwich in which the chitin layers are three-quarters of a wavelength thick and the air gap a quarter wavelength (where lambda=600 nm, the peak wavelength of the principal reflection maximum). It is shown that this configuration produces a second reflectance peak at approximately 385 nm, accounting for the observed reflection in the ultraviolet. Other scales have a similar thickness of chitin but lack the air gap and thus produce a dull purple reflection. This novel mechanism provides the spiders with two colour signals, both of which are important in mating displays.     

德融课程盐溶于汤：微生物学课程思政的思考与实践

课程思政是以“课程”作为“思政”的载体,探索知识传授、能力培养与价值塑造三位融合的有效途径,当前高等教育已步入全面建设课程思政的新时期。微生物学专业面与应用范围均非常广泛,而且与人类社会发展、生产实践、日常生活都息息相关,蕴含着极为丰富的思政元素,是开展课程思政的优秀载体。我们微生物学教学团队一直以来秉承教书育人的教育理念,致力于微生物学创新教学改革,近年来又以此为基础进行了课程思政思考与实践。本文总结了“微生物学”开展课程思政的必要性、融入路径、教学评价、反思与持续改进等,以期为高等院校课程思政教学改革提供参考。     

Single-molecule imaging and manipulation of biomolecular machines and systems

Background

Biological molecular machines support various activities and behaviors of cells, such as energy production, signal transduction, growth, differentiation, and migration.

盐胁迫对大豆根系木质部压力和Na+吸收的影响

取栽培大豆的水培幼苗为材料,用木质部压力探针和原子吸收分光光度计测定了盐胁迫条件下其根木质部压力和伤流液中Na~+含量的变化,以分析大豆抗盐吸水的机制.结果表明:在25～150 mmol/L NaCl的浓度范围内,随着盐胁迫强度的增加,大豆根木质部负压力的绝对值逐渐增大,但相对负压力和根的径向反射系数则逐渐减小;木质部伤流液中Na~+含量逐渐增加,但Na~+的相对含量则逐渐降低.同时,虽然根系吸水所需的木质部负压力(压力势)及根木质部伤流液的渗透势随着盐胁迫强度的增加都有所下降,但两者共同作用使木质部水势下降的幅度远远小于根外溶液水势(渗透势)下降的幅度,即随着根外溶液盐浓度的升高,根木质部溶液的总水势逐渐高出根外溶液的水势.上述结果说明,在盐胁迫下大豆可以利用相对小的木质部负压力逆水势梯度吸水,且通过避免对Na~+的过量吸收来适应盐胁迫环境.     

海盐对绿竹叶片反射光谱及叶绿素荧光参数的影响

沿海防护林是海岸生态系统中最基本的生物资源,由于沿海地区地下水位高,土壤含盐量高,肥力低,生态环境恶劣等特点,所以优良植物种选择是加快沿海优良防护林体系建设的关键。为了探讨绿竹耐盐性以及为沿海地区防护林选择优良植物种,以2年生绿竹(Bambusa oldhamii)为材料,采用水培法进行不同浓度的海盐处理,利用Unispec-SC型单通道光纤光谱仪和非调制式叶绿素荧光仪对绿竹叶片反射光谱和叶绿素荧光参数等进行测定。结果表明:当海盐浓度小于1.2%时,绿竹叶片中叶绿素a、叶绿素b和类胡萝卜素含量,以及"三边"参数与对照无显著差异,当海盐浓度升高到1.6%时,色素与对照相比分别降低了63.2%、62.8%和47.2%(P0.01),红边位置(λr ed)和红边面积(Sred)相比对照显著减小(P0.05)。1.2%浓度的海盐处理,红边归一化指数(rNDVI)、绿度归一化指数(gNDVI)、类胡萝卜素反射指数2(CRI700)和光化学反射指数(PRI)显著降低(P0.05),与对照相比分别降低了27.3%、23.3%、19.5%和43.9%;当海盐浓度增加到1.6%时,rNDVI、改良红边比值指数(mND)、gNDVI、改良归一化差值指数(mSR700)、类胡萝卜素反射指数1(CRI550)、CRI700和PRI等参数与对照相比分别下降了42.4%、43.9%、32.6%、21.5%、47.2%、49.9%和58.5%。绿竹叶片PSⅡ最大量子产率(Fv/Fm)、电子传递的量子产额(ΦEo)、单位反应中心捕获的用于电子传递的能量(ETo/RC)、单位面积反应中心数目(RC/CS)和叶片性能指数(PIABS)等参数,1.6%海盐处理与对照相比分别降低了50.8%、28.6%、21.7%、52.1%和92.3%,单位反应中心复合体吸收的能量(ABS/RC)比对照提高了96.9%。说明绿竹具有一定的耐盐性。