Monday, March 26, 2018

BacterioFiles 333 - Transposons Take Targeting Tool

Transposon structure
By Jacek FH, CC BY-SA 3.0
This episode: Certain transposons, genetic elements that move around the genome on their own, have co-opted the bacterial immune system, CRISPR, to use for jumping to new hosts!

Thanks to Dr. Joseph Peters for his contribution!
Download Episode (10.7 MB, 11.75 minutes)

Show notes:
Microbe of the episode: Streptomyces yokosukanensis

Journal Paper:
Peters JE, Makarova KS, Shmakov S, Koonin EV. 2017. Recruitment of CRISPR-Cas systems by Tn7-like transposons. Proc Natl Acad Sci 114:E7358–E7366.

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    Episode outline:
    • Background: CRISPR/Cas, microbial adaptive immune system
      • When viruses etc invade, captures bit of genome and stores
      • Uses to target genome in future, Cas protein chops it up
      • Many different kinds/versions
    • Cas etc used in a lot of gene editing applications recently, very powerful
    • Nature complicated; ep 120 virus uses system against bacterial antivirus defense
    • What’s new: Now, Joseph Peters, Kira Makarova, Sergey Shmakov, and Eugene Koonin, publishing in PNAS, have discovered that some mobile genetic elements have adapted parts of the CRISPR/Cas system to help them spread to new hosts!
    • Transposons – pieces of genetic material that can move around and copy selves to new places in genome, informally known as “jumping genes”
      • All organisms
      • More or less complex, number of types
      • Here, Tn7; very targeted site selection
    • Methods: Here’s the first author, Joseph Peters, describing how the initial finding came about: statement 1
      • So these transposons have a version of CRISPR-like system
      • No cutting protein, Cas, used to chop up invaders
      • Particular variant not found outside transposons, must not be useful except
    • So what is it doing?
      • Looking at CRISPR sequences, different transposons have different
      • Even in closely related bacteria
      • Must be replacing some with new as needed
      • Many didn’t match any known sequence
      • But of others, matched plasmids and phage genomes
    • Thought maybe helps target transposition to new places
      • Found in same place as a gene that helps other versions do this
      • Statement 2
      • Pretty clever, first known RNA guiding of transposons
    • Applications and implications: Here’s Dr. Peters again: Statement applications
      • So could be useful tool for large-scale genome editing applications
    • What do I think: Good strategy for spreading
      • Jump on genetic elements that are good at spreading already
    • Could knock out target phage by inserting
      • Sorta helpful to bacterial hosts, selective pressure to keep
    • Wonder: if spacers acquired, is target already recognized by cells?
    • System could have other effects too, like modifying host behavior to favor transposons
    • Any genes for proteins that are good at something have potential to be used by other things
      • Microbes, viruses, now transposons
      • And also us.

    Author Transcript:
    The work described in the paper stemmed from this fortuitous conversation I had at a Cold Spring Harbor meeting on transposons in November 2016. After hearing my seminar, Kira, who is an investigator in Eugene's lab, mentioned she was finding transposon genes associated with one of the CRISPR/Cas systems they had discovered bioinformatically. So we got to talking from there, and because the Koonin group already had a bioinformatic pipeline for working with these datasets and looking at associations with other genes, we were able to work quickly to determine if any one class of transposon had really picked up the new CRISPR/Cas variant. As luck would have it, the new CRISPR variants were associated with the exact system, the Tn7 system that I've been working on for a long time now. Within a few months we were able to work out the association using phylogeny, basically you can make a family tree of sorts that spans millions of years, to see if it matches between these two systems, the CRISPR/Cas system and this sort of unique transposon system. And we found that there was a match between these systems, and that really suggested that the CRISPR/Cas systems were captured by the transposon.

    we were also able to analyze the guide RNAs that were contained in these systems, and this really provided a smoking gun that these elements weren't using this as a defense, but really as a roadmap to travel to new bacteria.

    We're excited that this could really let us learn more about CRISPR/Cas systems and how they've been adapted in the environment, but we're really excited that, one potential advantage of this system is we could use it as a sort of targeted gene insertion system, where if we could have a synthetic guide RNA, we could direct a transposon to a specific site, but in an application-based setting, we wouldn't have the normal transposon information, but we would just use the ends of the transposon to flank any kind of series of genetic information we wanted to place in some specific place in a genome, because these transposons could be very big, we think even large blocks of sequence, so whole operons or very big genes could be done, you know, even if it was 100kb or more that wanted to be put in one place.


    1. Has there been other transposons using the CRISPR/Cas system other than TN7?

      1. This was the first report of such an occurrence, so though it's quite possible that other transposons may have the system, they haven't yet been discovered.

    2. What effect, if any, do you think this would have on mutation control/monitoring?

      1. You mean like, monitoring where transposons are moving around? I suppose awareness of the CRISPR system's existence could provide clues as to where the transposons might go or have already gone in moving between hosts, for example.