scholarly journals Linear elements are stable structures along the chromosome axis in fission yeast meiosis

Chromosoma ◽  
2021 ◽  
Author(s):  
Da-Qiao Ding ◽  
Atsushi Matsuda ◽  
Kasumi Okamasa ◽  
Yasushi Hiraoka
Keyword(s):  
Fission Yeast ◽  
Strand Break ◽  
Wide Field ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Meiotic Chromosomes ◽  
Linear Elements ◽  
Yeast Meiosis ◽  
Chromosome Axis ◽  
Stable Structures

AbstractThe structure of chromosomes dramatically changes upon entering meiosis to ensure the successful progression of meiosis-specific events. During this process, a multilayer proteinaceous structure called a synaptonemal complex (SC) is formed in many eukaryotes. However, in the fission yeast Schizosaccharomyces pombe, linear elements (LinEs), which are structures related to axial elements of the SC, form on the meiotic cohesin-based chromosome axis. The structure of LinEs has been observed using silver-stained electron micrographs or in immunofluorescence-stained spread nuclei. However, the fine structure of LinEs and their dynamics in intact living cells remain to be elucidated. In this study, we performed live cell imaging with wide-field fluorescence microscopy as well as 3D structured illumination microscopy (3D-SIM) of the core components of LinEs (Rec10, Rec25, Rec27, Mug20) and a linE-binding protein Hop1. We found that LinEs form along the chromosome axis and elongate during meiotic prophase. 3D-SIM microscopy revealed that Rec10 localized to meiotic chromosomes in the absence of other LinE proteins, but shaped into LinEs only in the presence of all three other components, the Rec25, Rec27, and Mug20. Elongation of LinEs was impaired in double-strand break-defective rec12− cells. The structure of LinEs persisted after treatment with 1,6-hexanediol and showed slow fluorescence recovery from photobleaching. These results indicate that LinEs are stable structures resembling axial elements of the SC.

scholarly journals The Linear Element is a Stable Structure Along the Chromosome Axis in Fission Yeast

2020 ◽  
Author(s):  
Da-Qiao Ding ◽  
Atsushi Matsuda ◽  
Kasumi Okamasa ◽  
Yasushi Hiraoka
Keyword(s):  
Fission Yeast ◽  
Wide Field ◽  
Structured Illumination ◽  
Dna Double Strand Breaks ◽  
Structured Illumination Microscopy ◽  
Time Treatment ◽  
Structure Changes ◽  
Structure Line ◽  
Short Time ◽  
Chromosome Axis

AbstractChromosomes structure changes dramatically upon entering meiosis to ensure the successful progression of meiosis-specific events. During this process, a multilayer proteinaceous structure called synaptonemal complex (SC) is formed in many eukaryotes. Instead, in the fission yeast Schizosaccharomyces pombe, linear elements (LinEs), which are structures related to an axial element of SC, form on the meiotic cohesin-based chromosome axis and are required for the formation of DNA double-strand breaks. In contrast to the well-organized SC structure, LinE structure had been observed only by silver-stained electron micrographs or in immuno-fluorescence stained spread nuclei. Thus, their fine structure and dynamics in intact living cells remain to be elucidated. In this study, we performed live cell imaging with wide-field fluorescence microscopy as well as 3D structured illumination microscopy (3D-SIM) for the four components of LinE, the Rec10, Rec25, Rec27 and Mug20. We found that LinEs consist of threads formed along the chromosome axes during the meiotic prophase. Rec10 binds to the chromosome itself and shapes into LinEs only in the presence of all the other LinE components. Rec25, Rec27, and Mug20 attach to the chromosome in the presence of Rec10. LinEs are stable in a short-time treatment with 1,6-hexanediol; and fluorescence recovery after photobleaching (FRAP) experiment reveals slow recovery from photobleaching, indicating a stable property of LinEs.

scholarly journals A technique for resolution assessment in blind-SIM experiments

2021 ◽  
Author(s):  
Imen Boujmil ◽  
Giancarlo Ruocco ◽  
Marco Leonetti
Keyword(s):  
High Resolution ◽  
Biological Sample ◽  
A Priori ◽  
Resolution Enhancement ◽  
Super Resolution ◽  
Experimental Setup ◽  
Wide Field ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Resolution Measurement

Super resolution techniques are an excellent alternative to wide field microscopy, providing high resolution also in (typically fragile) biological sample. Among the various super resolution techniques, Structured Illumination Microscopy (SIM) improve resolution by employing multiple illumination patterns to be deconvolved with a dedicated software. In the case of blind SIM techniques, unknown patterns, such as speckles, are used, thus providing super resolved images, nearly unaffected by aberrations with a simplified experimental setup. Scattering Assisted Imaging, a special blind SIM technique, exploits an illumination PSF (speckle grains size), smaller than the collection PSF (defined by the collection objectives), to surpass the typical SIM resolution enhancement. However, if SAI is used, it is very difficult to extract the resolution enhancement form a priori considerations. In this paper we propose a protocol and experimental setup for the resolution measurement, demonstrating the resolution enhancement for different collection PSF values.

scholarly journals Mug20, a novel protein associated with linear elements in fission yeast meiosis

2012 ◽  
Vol 58 (2) ◽  
pp. 119-127 ◽  
Author(s):  
Anna Estreicher ◽  
Alexander Lorenz ◽  
Josef Loidl
Keyword(s):  
Fission Yeast ◽  
Linear Elements ◽  
Yeast Meiosis ◽  
Novel Protein

Generation and Control of Wide-Field Three-Dimensional Structured Illumination for Advanced Microscopic Imaging

2012 ◽  
Vol 516 ◽  
pp. 640-644
Author(s):  
Shin Usuki ◽  
Hiroyoshi Kanaka ◽  
Kenjiro Takai Miura
Keyword(s):  
Three Dimensional ◽  
Fluorescent Imaging ◽  
Wide Field ◽  
Structured Illumination ◽  
Axial Resolution ◽  
Structured Illumination Microscopy ◽  
Spatial Control ◽  
Microscopic Imaging ◽  
Point Spread ◽  
Resolution Improvement

In a variety of practical microscopic imaging applications, many industries require not only lateral resolution improvement but also axial resolution improvement. The resolution in optical microscopy is limited by diffraction and determined by the wavelength of the incident light and the numerical aperture (NA) of the objective lens. The diffraction limit is mathematically described by a point spread function in the imaging system, and three-dimensional (3D) point spread functions describe both the lateral and axial resolutions. Thus, it is useful to focus on exceeding this limit and improving the resolution of optical imaging by the spatial control of structured illumination. Structured illumination microscopy is a familiar technique to improve resolution in fluorescent imaging, and it is expected to be applied to industrial applications. Microscopic imaging is convenient, non-destructive, and has a high-throughput performance and compatibility with a number of applications. However, the spatial resolution of conventional light microscopy is limited to wavelength scale and the depth of field is shallow; hence, it is difficult to obtain detailed 3D spatial data of the object to be measured. Here, we propose a new technique for generating and controlling wide-field 3D structured illumination. The technique, based on the 3D interference of multiple laser beams, provides lateral and axial resolution improvement, and a wide 3D field of view. The spatial configuration of the beams was theoretically examined and the optimal incident angle of the multiple beams was confirmed. Numerical simulations using the finite difference time domain (FDTD) method were carried out and confirmed the generation of 3D structured illumination and spatial control of the illumination by using the phase shift of incident beams.

Localized plasmon assisted structured illumination microscopy for wide-field high-speed dispersion-independent super resolution imaging

Nanoscale ◽  
2014 ◽  
Vol 6 (11) ◽  
pp. 5807-5812 ◽  
Author(s):  
Joseph Louis Ponsetto ◽  
Feifei Wei ◽  
Zhaowei Liu
Keyword(s):  
Antenna Array ◽  
High Speed ◽  
Super Resolution ◽  
Fluorescent Imaging ◽  
Wide Field ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Resolution Imaging ◽  
Localized Plasmon ◽  
Imaging Resolution

Fluorescent imaging resolution down to 51 nm is shown by generating tunable localized plasmon excitations on a nano-antenna array.

scholarly journals Laser-free super-resolution microscopy

2021 ◽  
Vol 379 (2199) ◽  
Author(s):  
Kirti Prakash
Keyword(s):  
Single Molecule ◽  
Super Resolution ◽  
Stimulated Emission ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Correlative Imaging ◽  
Deep Blue ◽  
Meiotic Chromosomes ◽  
Set Up ◽  
Super Resolution Microscopy

We report that high-density single-molecule super-resolution microscopy can be achieved with a conventional epifluorescence microscope set-up and a mercury arc lamp. The configuration termed as laser-free super-resolution microscopy (LFSM) is an extension of single-molecule localization microscopy (SMLM) techniques and allows single molecules to be switched on and off (a phenomenon termed as ‘blinking’), detected and localized. The use of a short burst of deep blue excitation (350–380 nm) can be further used to reactivate the blinking, once the blinking process has slowed or stopped. A resolution of 90 nm is achieved on test specimens (mouse and amphibian meiotic chromosomes). Finally, we demonstrate that stimulated emission depletion and LFSM can be performed on the same biological sample using a simple commercial mounting medium. It is hoped that this type of correlative imaging will provide a basis for a further enhanced resolution. This article is part of the Theo Murphy meeting issue ‘Super-resolution structured illumination microscopy (part 1)’.

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