In a new study, researchers from Colorado State University (CSU) has acquired an unprecedented achievement: made in living cells in vivo observation of ribosomal RNA translation --- basic cellular processes protein. Research results on May 5, 2016, published online in the journal Science, the paper titled "Real-time quantification of single RNA translation dynamics in living cells".
Francis - Crick first described in such centers after 60 years of rule, the researchers clarify this final step in gene expression in single living cells. Their tools are some smart protein modification and customization of a microscope, this microscope can be observed where a single RNA translation, with nanoscale precision.
This groundbreaking study was conducted by Assistant Professor, Department of Biology, Faculty of Science in Biochemistry and Molecular Colorado State University Tim Stasevich leadership. First author is a researcher Tatsuya Morisaki, wherein Morisaki created this microscope and conduct these experiments.
Stasevich said, "Before this, no one protein is expressed for imaging. This is the start-up and one of the key steps to turn genes, and genes at the translational level of regulation and many diseases are associated. This is very important, the reason is that such incidents can not be imaged, we can not recognize what went wrong. people in vitro to do this, but can not do this in living cells."
Proteins perform most cellular functions, and is the reason we are alive. Protein is translated from the RNA. But why it is difficult to observe it happen? This is because it takes time to mature and protein folding, and protein can be irradiated latest technology too slow and not able to catch the earliest stages of life of the protein. Even the use of green fluorescent protein (GFP) of the RNA is labeled, it takes too long. Stasevich explained, "When the light is turned on, the translation has been completed a good long time."
To solve this problem, the researchers encoded a protein receptor sites, sites where these receptors like a key in a lock. They let their experimental living cells express a simple fluorescent antibody fragments (ie, marked with GFP antibody fragment), wherein the antibody fragment is like the key to the lock. Once the translation occurs in cells that need to join the key in the lock, the results of this protein emit bright green fluorescence. Nascent protein remains attached to its messenger RNA (mRNA) on, and the researchers were able to observe and record everything that happens. By using different biochemical markers, they can perform a variety of posttranslational RNA imaging, wherein each protein could be identified by a different fluorescent colors.
This tagging process is one thing, but how to capture it is another matter. The researchers used their custom microscope capture. The microscope has no moving parts, but comprises two highly sensitive cameras, two colors can be simultaneously imaged on the RNA and protein.
Through these experiments, other researchers shared knowledge, including the proteins in living cells 10 extending in the rate of occurrence of the second amino acid. They also confirmed that the polysomes (polysome) consists of a string consisting of the ribosome is spherical, rather than elongated. Finally, they found that more ribosomal sometimes interact with each other, even when they are completely different encoded protein.
Translation of the RNA can be imaged may be able to more fully understand the mechanisms of gene expression. For example, the virus is too small to not have their own translation system, which was described as a battlefield translation virus and host cells. Stasevich said, "how viruses hijack our translation system for imaging will be very interesting."
Furthermore, such diseases such as cancer almost always involve multiple genes, and these genes often "talk" to each other. The researchers hope that they can study more deeply into the network of genes in order to understand how they function together and finally understand how to prevent their failure.
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