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Aquaporins (AQPs) constitute a highly diverse family of channel proteins that transport water and neutral solutes. AQPs play crucial roles in plant development and stress responses. However, the characterization and biological functions of RsAQPs in radish (Raphanus sativus L.) remain elusive. In this study, 61 non-redundant members of AQP-encoding genes were identified from the radish genome database and located on nine chromosomes. Radish AQPs (RsAQPs) were divided into four subfamilies, including 21 plasma membrane intrinsic proteins (PIPs), 19 tonoplast intrinsic proteins (TIPs), 16 NOD-like intrinsic proteins (NIPs), and 5 small basic intrinsic proteins (SIPs), through phylogenetic analysis. All RsAQPs contained highly conserved motifs (motifs 1 and 4) and transmembrane regions, indicating the potential transmembrane transport function of RsAQPs. Tissue- and stage-specific expression patterns of AQP gene analysis based on RNA-seq data revealed that the expression levels of PIPs were generally higher than TIPs, NIPs, and SIPs in radish. In addition, quantitative real-time polymerase chain reaction (qRT-PCR) revealed that seven selected RsPIPs, according to our previous transcriptome data (e.g., RsPIP1-3, 1-6, 2-1, 2-6, 2-10, 2-13, and 2-14), exhibited significant upregulation in roots of salt-tolerant radish genotype. In particular, the transcriptional levels of RsPIP2-6 dramatically increased after 6 h of 150 mM NaCl treatment during the taproot thickening stage. Additionally, overexpression of RsPIP2-6 could enhance salt tolerance by Agrobacterium rhizogenes-mediated transgenic radish hairy roots, which exhibited the mitigatory effects of plant growth reduction, leaf relative water content (RWC) reduction and alleviation of O2– in cells, as shown by nitro blue tetrazolium (NBT) staining, under salt stress. These findings are helpful for deeply dissecting the biological function of RsAQPs on the salt stress response, facilitating practical application and genetic improvement of abiotic stress resistance in radish.

Impulsive stimulated thermal scattering (ISTS) allows one to access the structural relaxation dynamics in supercooled molecular liquids on a time scale ranging from nanoseconds to milliseconds. Till now, a heuristic semi-empirical model has been commonly adopted to account for the ISTS signals. This model implicitly assumes that the relaxation of specific heat, C, and thermal expansion coefficient, γ, occur on the same time scale and accounts for them via a single stretched exponential. This work proposes two models that assume disentangled relaxations, respectively, based on the Debye and Havriliak-Negami assumptions for the relaxation spectrum and explicitly accounting for the relaxation of C and γ separately in the ISTS response. A theoretical analysis was conducted to test and compare the disentangled relaxation models against the stretched exponential. The former models were applied to rationalize the experimental ISTS signals acquired on supercooled glycerol. This allows us to simultaneously retrieve the frequency-dependent specific heat and thermal expansion up to the sub-100 MHz frequency range and further to compare the fragility and time scale probed by thermal, mechanical, and dielectric susceptibilities. ispartof: JOURNAL OF CHEMICAL PHYSICS vol:155 issue:16 ispartof: location:United States status: published

A generalized physical model is introduced to describe the impulsive stimulated scattering (ISS) response of relaxing systems to photothermal excitation in a periodical grating geometry. The proposed approach starts from Debye and Havriliak-Negami expressions for both the frequency-dependent heat capacity, C({\omega}), and thermal expansion coefficient, {\gamma}({\omega}). Simulations are carried out on glycerol to test and compare the developed models with the existing semi-empirical model [1]. Debye behavior of the specific heat capacity is shown to be compatible with a two-temperature scenario, in which, in addition to the classical, experimentally observable temperature that characterizes the distribution of the system over vibrational energy states, a second temperature characterizes the state of the configurational energy landscape. The models here developed have been applied for the interpretation of the experimental ISS signals of supercooled glycerol, illustrating simultaneous and separate assessment of C({\omega}) and {\gamma}({\omega}) up to sub-100 MHz from thermoelastic transients., Comment: 18 pages, 10 figures

Specific heat and linear thermal expansivity are fundamental thermal dynamics and have been proven as interesting relaxing quantities to investigate in glass transition and glassy state. However, their possibility has much less been exploited compared to mechanical and dielectric susceptibilities due to the limited spectroscopy bandwidth. This work reports on simultaneous spectroscopy of the two by making use of ultrafast time-resolved thermal lens (TL) spectroscopy. Detailed modeling of the thermoelastic transients of a relaxing system subjected to ultrashort laser heating is presented to describe the TL response. The model has been applied to analyze a set of experimentally recorded TL waveforms, allowing the determination of relaxation strength and relaxation frequency from sub-kilohertz to sub-100 MHz and in a wide temperature range from 200-280 K., Comment: 27 pages, 9 figures

International audience; Picosecond acoustics enable the mechanical characterization of transparent media by means of Time Domain Brillouin Scattering (TDBS) detection in the GHz frequency range and at the micrometer scale1. This technique is based on the interferences of a probe light with a propagating acoustic wave. We have demonstrated recently that silica nanoparticules, whose size matches the wavelength of phonons at the Brillouin frequency, can be used to discriminate the Brillouin oscillations in a soft transparent medium immersed in water2. Indeed, the multiple scattering of waves cancels the coherent acoustic propagation in water and consequently attenuates the Brillouin oscillations in the fluid whereas the Brillouin signature of the transparent medium remains unaffected. For ten years3, picosecond ultrasonics has been used to image the mechanics of biological cells. However, until now for most experimental achievements, the cells are fixed and dried on the opto-acoustic transducers. Few experiments have been done with immersed cells4-5 whereas it is an essential step toward the achievement of the characterization of living cells. In this work, we present experimental results of the use of a sediment of nanoparticules to improve the imaging of an immersed cell in water. [1] V. Gusev and P. Ruello, Appl. Phys. Rev. 5(3), 031101 (2018). [2 ] M.-F. Ponge, L. Liu, C. Aristégui, and B. Audoin, Appl. Phys. Lett. 115, 133101 (2019). [3] C. Rossignol, N. Chigarev, M. Ducousso, B. Audoin, G. Forget, F. Guillemot, and M. C. Durrieu, Appl. Phys. Lett. 93, 123901 (2008). [4] S. Danworaphong, M. Tomoda, Y. Matsumoto, O. Matsuda, T. Ohashi, H. Watanabe, M. Nagayama, K. Gohara, P. H. Otsuka, and O. B. Wright, Appl. Phys. Lett. 106, 163701 (2015). [5] F. Perez-Cota, R. J. Smith, E. Moradi, L. Marques, K. F. Webb, and M. Clark, Sci. Rep. 6, 39326 (2016).

International audience; Picosecond optoacoustic microscopy is a technique that allows imaging single cells with an in-plane resolution limited by optics and with the mechanical properties as the contrast mechanism.[1,2] In this talk, the basic principles involved in single cell picosecond ultrasonics will be briefly reminded, and the large variety of physical parameters that can be extracted from a single signal will be illustrated. Using a pulsed opto-acoustic microscope we produce notably high-resolution images of the stiffness and adhesion of single migrating cells. The technique is also capable to image cell 3D morphology with an out-of-plane accuracy down to a tenth of the optical wavelength.[3] This capability will be illustrated with the measurement of the 3D morphology of an entire osteosarcoma cell. The mapping complies with the image obtained by standard atomic force microscopy for the same cell, and both reveal very close roughness mean values. These results open perspectives for the morphological phenotyping of cancer cells. This opto-acoustic modality was inserted on a regular upright microscope, making it handy to cooperate with the built-in bright field and with fluorescence microscopies.

International audience; Picosecond opto-acoustic micro-spectroscopy is an optical technique based on ultrafast pump-probe generation and detection of hypersound on time durations of picoseconds and length scales of nanometers. This work reports on a dual-probe architecture to carry out real-time parallel detection of the hypersound propagation inside a transparent sample and, simultaneously, of the hypersound reflection at a metal/sample interface. This dual architecture offers unprecedented possibilities to image, at a micron scale, the mechanical properties of a non-homogeneous transparent material. We will show that, using this dual- optoacoustic modality, several biomechanical properties of a cell can be measured in a non-contact and label-free manner. The technique capabilities are notably illustrated with the imaging of multiple biomechanical properties of a mitotic macrophage-like cell, in a single run experiment. [1] Liu L., Plawinski L., Durrieu M-C, and Audoin B., "Label-free multi-parametric imaging of single cells: dual picosecond optoacoustic microscopy", Journal of Biophotonics 12(8), e201900045, 2019 doi:10.1002/jbio.201970028

Additional file 1 Figure S1. The sequence length distribution of small RNAs during radish taproot thickening. Figure S2. The heatmap of DEGs and DEMs in radish taproot thickening. a. Heatmap of DEMs in ‘NAU-DY’; b, c represents heatmap of DEGs in ‘NAU-DY’ and ‘NAU-YB’, respectively. Figure S3. Comparative analysis of gene expression profiles from RNA-seq and RT-qPCR. 20 randomly selected genes by RNA-seq and RT-qPCR showing different expression patterns in three comparative groups (DS2 vs DS1, DS3 vs DS1, and DS3 vs DS2) during taproot thickening. Each data point represents the log2 normalized expression level obtained from RNA-seq (x axis) and RT-qPCR (y axis) analyses. Figure S4. RT-qPCR validation of 14 DEMs during radish taproot thickening. The relative expression of DEMs between DS2 and DS1 libraries (a), DS3 and DS1 libraries (b) and DS3 and DS2 libraries (c) were analyzed by the 2−ΔΔCT method. Table S1. Primer sequences for RT-qPCR assay. Table S2. Summary of small RNA sequencing data. Table S3. Summary of mRNA sequencing data. Table S4. Detailed information of DEMs during radish taproot thickening in ‘NAU-DY’. Table S6. DEMs and their corresponding targets extracted from RNA-seq.

Cell morphological analysis has long been used in cell biology and physiology for abnormality identification, early cancer detection, and dynamic change analysis under specific environmental stresses. This work reports on the remote mapping of cell 3D morphology with an in-plane resolution limited by optics and an out-of-plane accuracy down to a tenth of the optical wavelength. For this, GHz coherent acoustic phonons and their resonance harmonics were tracked by means of an ultrafast opto-acoustic technique. After illustrating the measurement accuracy with cell-mimetic polymer films we map the 3D morphology of an entire osteosarcoma cell. The resulting image complies with the image obtained by standard atomic force microscopy, and both reveal very close roughness mean values. In addition, while scanning macrophages and monocytes, we demonstrate an enhanced contrast of thickness mapping by taking advantage of the detection of high-frequency resonance harmonics. Illustrations are given with the remote quantitative imaging of the nucleus thickness gradient of migrating monocyte cells. ispartof: SCIENTIFIC REPORTS vol:9 issue:1 ispartof: location:England status: published

In this study, 51,625 unique expressed sequence tag (ESTs) from a total of 289,621 radish ESTs in the National Center for Biotechnology Information (NCBI) database were used to search for simple sequence repeat (SSRs) by SSRLocate, and 2,917 SSRs in 2,891 ESTs were identified. The SSR marker motifs contained di-, tri-, tetra-, penta-, and hexa- nucleotide repeats, and the number was 945 (32.40%), 1,300 (44.57%), 179 (6.14%), 262 (8.98%) and 231 (7.92%), respectively. The motifs AG/CT (16.15%) and GA/TC (13.58%) were the most abundant type. Among 20 amino acids encoded by trinucleotide, Ser (16.17%) was the most common transcript, followed by Leu (13.00%) and Glu (9.90%). A total of 1,082 EST-SSR primers in Raphanus sativus were designed and synthesized, and 864 (79.85%) EST-SSRs were successfully amplified. Polymorphism of these loci was evaluated in a panel of 48 genotypes in Raphanus. The polymorphism information content (PIC) values of those primers varied from 0.33 to 0.84, with a mean value of 0.58. Moreover, it was found that 25 out of 53 EST-SSR primers could be successfully amplified in Brassica species. For its polymorphism, reproducibility and functionality, these novel EST-SSR markers could be used as a powerful tool for marker-assisted selection (MAS) and genetic mapping in radish.Keywords: Raphanus sativus L., expressed sequence tag (EST), simple sequence repeat (SSR), genetic diversity, polymorphism information contentAfrican Journal of Biotechnology Vol. 12(9), pp. 921-935

This PhD-project deals with the development and experimental validation of an all-optical thermometry method that enables to detect remotely and very high temporal resolution (10 nanoseconds, 100MHz bandwidth). The technique was used to tackle the challenge to investigate the relaxation behavior of the heat capacity of the glassformer glycerol, and thus supply information that can be compared with the relaxation features of other, classically investigated physical properties of glassformes such as the dielectric constant and the mechanical properties. The developed method is exploiting the temperature dependence and rapid response time of the fluorescence characteristics of a probe material. In the investigated application, a transient temperature jump was induced by irradiating the sample with a nanosecond laser pulse, whose energy was optically absorbed and converted into heat. Also a configuration in which the sample was periodically heated by optical excitation was implemented. Contents Acknowledgements Abstract Nederlandstalig abstract Abbreviations/Symbols Chapter 1 Introduction 1.1 Background and state of the art 1.2 Motivation and research objectives 1.3 Outline of the dissertation References Chapter 2 Temperature dependent fluorescence 2.1 Calibration setup for investigation of temperature dependent fluorescence 2.2 Experimental results and discussion 2.3 Conclusion References Chapter 3 Temperature retrieval by neural network (NN)recognition 3.1 Introduction 3.2 NN implementation of fluorescence-based thermometry 30 3.3 Performance of temperature reconstruction by NN 3.4 Conclusion References Chapter 4 Development of ultrafast fluorescence based thermometry 4.1 Introduction 4.2 CW probe-pulsed pump implementation 4.3 Pulsed probe-pulsed pump implementation 4.4 Conclusion References Chapter 5 Spatially resolved photothermal fluorescence spectroscopy in frequency domain 5.1 Experimental setup 5.2 Results and discussion 5.3 Conclusion References Chapter 6 Lock-in photothermal photoluminescence spectroscopy in frequency domain 6.1 Introduction 6.2 Temperature dependence of the QD photoluminescence 6.3 Lock-in detection of photothermal photoluminescence signals 6.4 Experimental results and discussion 6.5 Conclusion References Chapter 7 Temperature dependent time-resolved photothermal spectroscopy of glycerol 7.1 Introduction • Relaxation dynamics in supercooling systems • Relaxation of specific heat capacity 7.2 Time-resolved thermal lens spectroscopy of supercooled glycerol • Experimental setup • Results and discussion 7.3 Photothermal fluorescence spectroscopy of supercooled glycerol • Simulation of C effect on the impulsive temperature response • Experiment results and discussion 7.4 Conclusion References Chapter 8 General conclusion and perspective 8.1 Conclusion 8.2 Perspective Curriculum Vitae List of publications nrpages: 149 status: published

Muscarinic agonist specificity is limited, making it difficult to match receptor subtypes with signal transduction cascades that mediate ion channel modulation. We have characterized the inhibitory effects of two muscarinic agonists, oxotremorine-M (Oxo-M) and bethanechol chloride (BeCh), on Ca2+ currents in neonatal rat superior cervical ganglion neurons.Oxo-M-mediated (10 μM) inhibition occurred via two signaling pathways. The first pathway inhibited whole cell peak currents, consisting primarily of N-type current, but not FPL 64176-induced, long-lasting tail currents, comprised entirely of L-type current. Inhibited currents displayed slowed activation kinetics and voltage dependence, characteristics of membrane-delimited inhibition. Current inhibition was blocked by the selective M2 receptor antagonist, methoctramine (METH; 100 nM), or following pertussis toxin (PTX) pretreatment.Activation of the second pathway inhibited both peak and long-lasting tail currents. This pathway was voltage-independent, PTX-insensitive, but sensitive to internal Ca2+ chelator concentration. Muscarinic toxin 7 (MT-7, 100 nM), an irreversible M1 receptor antagonist, eliminated this inhibition. Oxo-M (100 μM) decreased L- and N-type channel activities in cell-attached patches, indicating that a diffusible second messenger is involved.BeCh (100 μM) also inhibited whole cell currents via the membrane-delimited pathway. Blocking M4 receptors with 100 nM pirenzepine (in the presence of MT-7) had no effect, while antagonizing M2 receptors with METH abolished inhibition. Concentrations of BeCh as high as 3 mM failed to inhibit either peak or long-lasting tail currents following PTX pretreatment.These results indicate that BeCh may be an effective tool for selectively activating M2 receptor stimulation of the membrane-delimited pathway.