As an indispensable process for all life, gene expression is divided into two steps: DNA is transcribed into RNA, and then RNA is translated into protein.
In a new study, researchers from Georgia State University, the University of California at Berkeley, and Northwestern University combined cryo-electron microscopy (Cryo-EM) with the latest computational modeling methods, unprecedentedly The molecular structure of the human transcriptional pre-initiation complex (PIC) at near-atomic resolution was analyzed in detail. Human PIC is a protein assembly that organizes RNA polymerase in place to ensure transcription is enabled.
These new structures help to gain insight into a range of conformational changes that occur in human PIC during the entire process of transcription initiation, including identifying the promoter region of DNA promoter initiation, opening this promoter region, and initiating transcription. . The relevant research results were published online in the Nature Journal on May 11, 2016, and the title of the paper is "Near-atomic resolution visualization of human transcription promoter opening".
Genes are made up of DNA that stores genetic information. In order to use the genetic information encoded in the gene, the RNA polymerase must copy the gene into messenger RNA (mRNA). This copying process is called transcription.
At the beginning of this tightly controlled transcription process, RNA polymerase and general transcription factors (general transcription)
Factor) assembles together at specific sites in the DNA to form a PIC. This PIC assembly is required to open the double-stranded DNA helix of the promoter to place the DNA on the active site of the RNA polymerase and initiate the transcription process. The resulting mRNA transcript is then used to make the protein.
Ivaylo Ivanov, author of the paper and associate professor of chemistry at Georgia State University, said, "This paper provides detailed structural information on the complexes involved in the early stages of the transcription process. We studied RNA polymerase in order to open the transcription bubble and initiate the transcription process. And every step taken by a general transcription factor. This is a very important system that could not be analyzed by crystal crystallization or any other structural method before. This is the first near-atom resolution Cryo of the human PIC complex. - EM structure reconstruction map."
Chemical cross-linking and crystal crystallization have provided structural information for partial RNA polymerase complexes from eukaryotes such as yeast, but these techniques are not capable of resolving the structure of the intact PIC complex. PIC causes events and processes of DNA unwinding, as well as transcriptional bubble formation - a molecular structure formed during the unwinding of a portion of a double-stranded DNA fragment during transcription, which is formed by RNA polymerase core enzymes, DNA template strands, and transcription The transcriptional complexes formed by the combination of the three new RNA strands are not well understood.
To obtain the detailed structure of the human PIC complex, the researchers combined Cryo-EM with integrated molecular computational modeling techniques that rely on supercomputing techniques to capture human PIC structures in three different functional states: 1) with promoters The closed state of the DNA double-helical contact of the region; 2) the open state when contacted with the transcription bubble, and 3) the initial transcription state ready to perform mRNA synthesis. They are also able to visualize a variety of previously unidentified components in the human PIC complex. These findings revealed a complete subunit assembly structure of the transcription factor TFIIH, in which TFIIH plays a crucial role in opening the promoter region. TFIIH is one of the most difficult to resolve components of the PIC complex.
By comparing the PIC's off-state, on-state, and initial transcriptional state, the researchers developed new insights into DNA contact, promoter unwinding, and transcriptional bubble stabilization.
Ivanov said, "Without the advances in electron microscopy, it would have been impossible without the recent advances in integrated computational modeling techniques. The electron microscopy of near-atomic resolution was only available in electron microscopy. This is only possible when combined with the powerful computational algorithms of new analytical images."
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