Like our own bodies, cells have their own skeletons called 'cytoskeletons' and are made of proteins instead of bones.
These network-like structures maintain the cell's shape, provide mechanical support, and are involved in critical processes of the cell's lifecycle. The cytoskeleton is an object of intense scientific and medical research, which often requires being able to observe it directly in cells.
Ideally, this would involve highly-fluorescent molecules that can bind cytoskeletal proteins with high specificity without being toxic to the cell. Publishing in Nature Methods, EPFL scientists have exploited the properties of a new fluorescent molecule, also developed at EPFL, to generate two powerful probes for the imaging of the cytoskeleton with unprecedented resolution. These probes pave the way for the easier and higher quality imaging of cells, offering many scientific and medical advantages.
The cytoskeleton is a large structure inside cells that provides them with mechanical support, keeps their three-dimensional shape and internal organization, and enables them to move and divide. It consists of three major sub-structures inside the cell, which are made up of long, filamentous proteins: tubulin and actin.
Current techniques for observing the cytoskeleton can be difficult to get into living cells, can be toxic, and are usually limited in resolution and duration, since the signal wears off over time. A common technique is fluorescence microscopy, where fluorescent molecules ('probes') are attached to cell structures and then 'lit up' against a dark background.
The team of Kai Johnsson at EPFL has developed novel fluorescent probes that can easily enter live cells, are non-toxic, have long-lasting signals, and most importantly, offer unprecedented image resolution. In 2013, the researchers developed a fluorescent molecule called silicon-rhodamine (SiR), which switches 'on' only when it binds to the charged surface of a protein like the ones found on the cytoskeleton. When SiR switches 'on', it emits light at far-red wavelengths.
The challenge was getting SiR to bind specifically to the cytoskeleton's proteins, actin and tubulin. To achieve this, the scientists fused SiR molecules with compounds that bind tubulin or actin. The resulting hybrid molecules consist of a SiR molecule, which provides the fluorescent signal, and a molecule of a natural compound that can bind the target protein. One such compound was docetaxel, an anticancer drug that binds tubulin, and the other jasplakinolide, which specifically binds the cytoskeletal form of actin. Both compounds, which are used here in very low, non-toxic concentrations, can easily pass through the cell's membrane and into the cell itself.
The probes, named SiR-tubulin and SiR-actin, were used to visualize the dynamics of the cytoskeleton in human skin cells. Because the light signal of the probes is emitted in the far-red spectrum, it is easy to isolate from background noise, which generates images of unprecedented resolution when used with a technique called super-resolution microscopy.
An additional advantage is the practicality of the probes. "You just add them directly into your cell culture, and they are taken up by the cells", says Kai Johnsson. The probes also do not require any washing or preparation of the cells before administration or any subsequent washing steps, which greatly helps in maintaining the stability of their environment and their natural biological functions.
The scientists believe that they can extend their work into other types of proteins and tissues. "Cytoskeletal structures are imaged by biologists all the time", says Johnsson. "Up to now, no probes were available that would allow you to get high quality images of microtubules and microfilaments in living cells without some kind of genetic modification. With this work, we provide the biological community with two high-performing, high-contrast fluorogenic probes that emit in the non-phototoxic part of the light spectrum, and can be even used in tissues like whole-blood samples."
This work represents a collaboration between EPFL's Institute of Chemical Sciences and Engineering (ISIC), Institute of Bioengineering (IBI), and the Bioimaging and Optics Platform (BIOP), with the National Centre of Competence in Research (NCCR) in Chemical Biology; the Max-Planck Institutes for Biophysical Chemistry (Göttingen) and of Molecular Physiology (Dortmund); the Friedrich-Schiller-University's Institute of Organic Chemistry (Jena); and the Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA) (Vienna).
Lukinavičius G, Reymond L, D'Este E, Masharina A, Göttfert F, Ta H, Güther A, Fournier M, Rizzo S, Waldmann H, Blaukopf C, Sommer C, Gerlich DW, Arndt HD, Hell SW, Johnsson K. Fluorogenic probes for live-cell imaging of the cytoskeleton. Nature Methods 25 May 2014. DOI: 10.1038/nmeth.2972
Nik Papageorgiou | Eurek Alert!
A new potential biomarker for cancer imaging
05.02.2016 | Universiti Putra Malaysia (UPM)
NIH researchers identify striking genomic signature shared by 5 types of cancer
05.02.2016 | NIH/National Human Genome Research Institute
Automobiles increase the mobility of their users. However, their maneuverability is pushed to the limit by cramped inner city conditions. Those who need to...
Advance in biomedical imaging: The University of Würzburg's Biocenter has enhanced fluorescence microscopy to label and visualise up to nine different cell structures simultaneously.
Fluorescence microscopy allows researchers to visualise biomolecules in cells. They label the molecules using fluorescent probes, excite them with light and...
NASA's follow-on to the successful ICESat mission will employ a never-before-flown technique for determining the topography of ice sheets and the thickness of sea ice, but that won't be the only first for this mission.
Slated for launch in 2018, NASA's Ice, Cloud and land Elevation Satellite-2 (ICESat-2) also will carry a 3-D printed part made of polyetherketoneketone (PEKK),...
In the last decades, sea level has been rising continuously – about 3.3 mm per year. For reef islands such as the Maldives or the Marshall Islands a sinister picture is being painted evoking the demise of the island states and their cultures. Are the effects of sea-level rise already noticeable on reef islands? Scientists from the ZMT have now answered this question for the Takuu Atoll, a group of Pacific islands, located northeast of Papua New Guinea.
In the last decades, sea level has been rising continuously – about 3.3 mm per year. For reef islands such as the Maldives or the Marshall Islands a sinister...
The ‘Internet of Things’ is growing rapidly. Mobile phones, washing machines and the milk bottle in the fridge: the idea is that minicomputers connected to these will be able to process information, receive and send data. This requires electrical power. Transistors that are capable of switching information with a single electron use far less power than field effect transistors that are commonly used in computers. However, these innovative electronic switches do not yet work at room temperature. Scientists working on the new EU research project ‘Ions4Set’ intend to change this. The program will be launched on February 1. It is coordinated by the Helmholtz-Zentrum Dresden-Rossendorf (HZDR).
“Billions of tiny computers will in future communicate with each other via the Internet or locally. Yet power consumption currently remains a great obstacle”,...
02.02.2016 | Event News
26.01.2016 | Event News
26.01.2016 | Event News
05.02.2016 | Life Sciences
05.02.2016 | Materials Sciences
05.02.2016 | Physics and Astronomy