Science: Chen Lingling's group and Liu Jiaquan's group collaborated to discover the important regulatory mechanism of long non-coding RNA on cell nucleolar structure and RNA polymerase I transcription

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July 30, internationally renowned academic journal "Science" ( Science ) published the results of research cooperation Innovation Center Molecular Cell Science, Chinese Academy of Excellence (Institute of Biochemistry and Cell Biology) Chen Lingling Liu Jia Quan study groups and study groups in the form of Research Article " LncRNA SLERT controls phase separation of FC/DFCs to facilitate Pol I transcription", and was reviewed by Perspectives during the same period
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This study revealed for the first time that long non-coding RNA SLERT changes the conformation of the interacting nucleolar protein DDX21 through the mechanism of RNA molecular chaperones, thereby affecting the size and fluidity of the FC/DFC region, and maintaining the efficient transcription of RNA polymerase I to generate ribosomal RNA

The nucleolus is the ribosomal RNA processing plant in the nucleus.
From the inside to the outside, it can be divided into three layers: the fiber center area (FC), the high density fiber area (DFC) and the granular area (GC)
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The Chen Lingling group used the ultra-high resolution microscope system to analyze the three-dimensional fine structure of the nucleolus of human living cells in detail.

In the latest study, researchers found SLERT missing DDX21 tufted not only causes small spherical shell proteins and flowability decreases, and the reduction can also cause curing entire FC / DFC transcription unit
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Based on the characteristics of the nucleolus structure, the researchers used ultra-high resolution structured illumination microscope (SIM) imaging technology and found that DDX21 relies on the formation of the DFC area to be positioned outside the DFC area, and the DDX21 layer has a restrictive effect on the inner structure and has a restrictive effect on the nucleolus.

The nucleolus is the largest membraneless organelle in the cell, and its phase separation feature has an important effect on the structure, dynamic changes and physiological functions of the nucleolus
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In order to further study the mechanism of SLERT regulating nucleolar FC/DFC, the researchers used in vitro protein phase separation, fluorescence bleaching recovery, precipitation separation and other technical methods to find that DDX21 protein presents a fibrous (fiber) morphology in vitro.

DDX21 has intramolecular and intermolecular interactions with comparable strength and mutual influence.
Strong intermolecular interactions lead to increased protein multimerization.
The high aggregation of DDX21 molecules compresses the size of the FC/DFC region and restricts the flow of the FC/DFC region.
The combination of SLERT and DDX21 increases the intramolecular interaction of DDX21 and makes the DDX21 molecule present a closed conformation; the closed conformation of DDX21 has a lower intermolecular binding effect, weakens the inhibition of DDX21 on the size of FC/DFC, and forms a relatively loose space environment to maintain high efficiency of RNA polymerase I transcription
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In order to further study how SLERT- regulated DDX21 clusters affect rDNA transcription, Chen Lingling’s group and Liu Jiaquan’s group used single-molecule total internal reflection fluorescence microscopy (smTIRF) to find that DDX21 forms a protein aggregation of hundreds of molecules at 50 nM, and DDX21 molecular clusters can bound and crimped linear rDNA molecules, the wound can not be combined with DDX21 rDNA important RPA49 RNA polymerase I subunits
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SLERT promotes the closed conformation of DDX21 and increases the fluidity of DDX21 can prevent DDX21 from encapsulating rDNA, thereby ensuring the effective occupancy of RNA polymerase I complex on rDNA and transcription

lncRNA is generally expressed at a low level, and the mode of its regulation has always been an important issue in the field.
However, the one-to-one targeting theory of traditional biology is not completely applicable to the number of low-dose molecules of lncRNA and the organisms it controls.
Phenotype
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This study found that SLERT can be used as an RNA chaperone to regulate the polymer state of DDX21 at low doses.

Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences (Institute of Biochemistry and Cell Biology) Chen Lingling's PhD students Wu Man, Xu Guang, and Liu Jiaquan PhD students in the research group Han Chong are the co-first authors of the paper, and Researcher Chen Lingling and Researcher Liu Jiaquan He is the co-corresponding author of the paper
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This work found for the first time that SLERT regulates the conformation and polymer state of proteins by RNA molecular chaperone mechanism, and observed the functional characteristics and regulatory mechanisms of protein microscopic phase separation at the single-molecule level
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The polymer state of protein is directly related to its fluidity, intermolecular interaction and function.
RNA chaperones maintain the normal morphological function of nucleolus by regulating the phase separation characteristics of nucleolar protein across orders of magnitude.
In-depth research in the field of lncRNA and phase separation It is of great significance
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This work received financial support from the National Natural Science Foundation of China, the Chinese Academy of Sciences, the Ministry of Science and Technology and the Shanghai Municipal Science and Technology Commission
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　　Article link: https://science.
sciencemag.
org/content/373/6554/547

　　(A) Top left: In the in vitro phase separation experiment, the formation of fibrous DDX21 decreases with the decrease of protein concentration; bottom left: In different experimental systems, different concentrations of DDX21 protein present different molecular states, SLERT can increase the flow of DDX21 protein sex; Right: in vivo and in vitro experiments reconstruction, SLERT increase the size and flowability FC / DFC transcription unit by combining DDX21
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　　(B) DDX21 molecular clusters gather on the outside of the nucleolar DFC area, and limit the size and fluidity of the FC/DFC area
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SLERT regulated by the open conformation DDX21 into closed, DDX21 oligomeric molecules exhibit loose state, to provide a suitable environment for space efficient transcription of Pol I
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Pol I transcription occurs at the junction of FC and DFC, and DDX21 coil-wound rDNA prevents the Pol I transcription complex from occupying rDNA
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SLERT regulates the conformation and mobility of DDX21, inhibits the winding of DDX21 on rDNA and promotes Pol I transcription
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