Contactless control of gene expression
Biologists have learned to control genes unnoticed by the genome
Alexander Ershov, N+1
Biochemists from the Massachusetts Institute of Technology have developed a tracking system that allows you to see where, when and how active the RNA of the desired gene is. Making small changes to the tracking system turns it from passive to active – it becomes possible to control the work of genes without affecting the genome. The work was published in the journal Proceedings of the National Academy of Sciences (Adamala et al., Programmable RNA-binding protein composed of repeats of a single modular unit), briefly about it can be read in the press release MIT Controlling RNA in living cells: Modular, programmable proteins can be used to track or manipulate gene expression.
The system works as follows. A genetic construct is introduced into the cell, which ensures the synthesis of two proteins. Proteins are two halves of an artificial marker. Each of the proteins has a part responsible for RNA recognition, and a part responsible for the actual glow.
The second, fluorescent, part is an ordinary green fluorescent protein (GFP), divided into two halves. These halves do not glow separately from each other, but if the two parts are brought close to each other, they connect (non-covalently) into a single fluorescent complex and become visible in ultraviolet.
Now, if you "teach" the parts of the marker that are responsible for RNA recognition to find two neighboring sequences in the desired RNA, this will ensure the convergence of the fluorescent halves, and such RNA can be easily seen in ultraviolet. If half of the marker binds to non-targeted, random RNAs, then the probability of assembling a fluorescent complex of two molecules will be several orders of magnitude lower. This strategy of "splitting" the marker makes it possible to achieve high specificity of binding even when a single RNA-binding part of the protein is not too legible.
Scientists have shown that it is possible to create a system for controlling the work of genes on the same element base. To do this, it is enough to replace the fluorescent part of the marker with one of the translation initiator proteins (that is, protein synthesis in ribosomes). The recognizing part should be directed to the start point of the broadcast. Then the former marker will bind to the desired matrix RNA, but not glow, but attract ribosomes to it. As a result, the activity of the gene (that is, the number of copies of the protein that are produced from it per unit of time) will increase significantly, but not a single "letter" in the genome will be changed.
The scheme of the method. The coding sequence is responsible for synthesis
control fluorescent protein – red mRuby
(picture from an article in PNAS).
It should be noted that different parts of the new system (for example, the method of dividing GFP into two halves) were created by other researchers. The most important innovation of the authors of the article is the creation of a method for selecting the amino acid sequence of the part of the protein that recognizes the necessary RNA. To do this, the scientists used a modular approach – they found in the proteins of the Pumilio family short amino acid fragments that have a high specific affinity for a particular base in RNA. By combining these fragments with each other, biologists have learned to create completely artificial proteins specific to any desired RNA sequence.
Ideologically, this approach is close to the method of designing artificial nucleases based on transcription factor proteins (TALEN) and zinc finger proteins. But these proteins bind to DNA, and the proteins of the Pumilio family bind to RNA.
Pumilio Protein Complex with RNA (1M8W)
Alexander Ershov / Wang, X., McLachlan, J., Zamore, P.D., Hall, T.M.T.
The structure of single-stranded RNA is very different from the usual double-stranded DNA, so the transfer of the approach from TALEN to Pumilio is very far from trivial.
Portal "Eternal youth" http://vechnayamolodost.ru
27.04.2016