More than a decade ago, scientists harnessed a bacterial molecular machine that identifies and cuts specific sections of DNA, revolutionizing the ability to edit genes and accelerating research into treatments for all manner of diseases with a genetic link. But the technology known as CRISPR-Cas9 works by cutting DNA, not moving it. At Purdue University, researchers are investigating a similar molecular machine that moves so-called \u201cjumping genes\u201d into new locations in bacterial DNA, laying the groundwork for a more powerful gene-editing tool.<\/p>\n
The team, led by\u00a0Leifu Chang<\/a>, a Purdue associate professor of biological sciences, has produced high-resolution structural snapshots of the Tn7-like transpososome, a complex of nucleic acids and proteins that can accurately cut and paste an entire gene from one location to another in the genome of a cell. This structural information is analogous to an exploded parts diagram for an engine, showing all the parts in atomic-level detail and how they work together. In a pair of papers published in the journal Cell in\u00a02023<\/a>\u00a0and\u00a02024<\/a>, the team captured the structure of all the components needed to understand how the molecular machine recognizes a jumping gene and the location in the genome where it will insert that gene.<\/p>\n \u201cWe\u2019ve captured all the components that are really essential in this structure. This is a quite complicated process, requiring recognition of specific DNA in the gene and in the target and, while previous research has shown many partial structures, it\u2019s important to see the whole picture,\u201d said Chang, a member of the\u00a0Purdue Institute for Cancer Research<\/a>. \u201cWe\u2019re providing a lot of information to understand how that process happens.\u201d<\/p>\n Chang\u2019s research is part of Purdue\u2019s presidential One Health initiative, which involves research at the intersection of human, animal and plant health and well-being.<\/p>\n There are many similarities between the Tn7-like transpososome and the CRISPR system. CRISPR, which evolved as part of a bacterial defense system against known viral invaders, identifies and stores snippets of DNA from invaders between DNA brackets in a pattern called \u201cclustered regularly interspersed short palindromic repeats,\u201d hence CRISPR. Using the stored DNA as a template, the system generates an RNA mirror image of the viral DNA and surveils the cell looking for a match. When a match is made and an invader identified, a protein cuts the DNA of the viruses, blocking replication. The beauty of the system, from the standpoint of researchers, is that it uses RNA \u2014 a molecule easily synthesized in a lab \u2014 to identify and target DNA. By synthesizing an RNA snippet that searches for its own targets, researchers can use the system to snip DNA with precision.<\/p>\n But while CRISPR is great at what it does in bacteria, Chang said, it\u2019s not a one-stop gene editing tool in human cells. For starters, cutting DNA in humans triggers a DNA repair pathway that could undo the cut or unintentionally introduce a mutation as part of the repair. Also, once the DNA is cut, it might not be repaired in which case the cell would die. Researchers have devised various workarounds, but they aren\u2019t yet efficient and only work in dividing cells, putting repairs to cells that don\u2019t divide, like neurons, out of reach.<\/p>\n By contrast, the transpososome is a complete package that includes the machinery for inserting genes. The transpososome facilitates the movement of transposons, or \u201cjumping genes,\u201d that can be copied and moved to different locations in the genome. Jumping genes make up about half of the genome in animal cells, including in humans, and are believed to increase genetic diversity.<\/p>\n \u201cIn general, breaking DNA to achieve genome editing is not ideal,\u201d Chang said. \u201cThe transposon system is a more efficient approach because the proteins insert DNA seamlessly, so that avoids the harmful consequence of breaking DNA.\u201d<\/p>\n Scientists who study the Tn7-like transpososome have discovered two pathways that it uses to find its target. The first uses a protein to directly recognize specific DNA sequences. The second is similar to CRISPR-Cas9, in that it uses a snippet of RNA as a guide to find the target. Chang,\u00a0who is an expert in CRISPR-Cas systems<\/a>, is interested in understanding how the CRISPR-type pathways work.<\/p>\n The two Cell papers taken together present complete cryo-electron microscopy structures of essential components in atom-by-atom detail. The structures show the point at which DNA attaches to two proteins, which triggers formation of the entire transpososome complex, and initiate transposition. As the 2024 paper states, \u201cThe findings provide mechanistic insights into targeted DNA insertion by Tn7-like transposons with implications for improving the precision and efficiency of their genome-editing applications.\u201d<\/p>\n Chang said researchers are already trying to use the Tn7-like transpososome to edit animal cells but, so far, the process isn\u2019t effective. While much work needs to be done to arrive at systems useful in nonbacterial cells, the full structural information he has provided will accelerate that work.<\/p>\n References:\u00a0<\/b><\/p>\n 1. Wang S, Gabel C, Siddique R, Klose T, Chang L. Molecular mechanism for Tn7-like transposon recruitment by a type I-B CRISPR effector. Cell. 2023;186(19):4204-4215.e19. doi:\u00a010.1016\/j.cell.2023.07.010<\/a> 2. Wang S, Siddique R, Hall MC, Rice PA, Chang L. Structure of TnsABCD transpososome reveals mechanisms of targeted DNA transposition. Cell. 2024;187(24):6865-6881.e16. doi:\u00a010.1016\/j.cell.2024.09.023<\/a> <\/p>\n This article has been republished from the following materials<\/a>. Note: material may have been edited for length and content. For further information, please contact the cited source. 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