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| Title | Structural basis of TnsC oligomerization and transposase recruitment in type I-B CRISPR-associated transposons. |
|---|---|
| Journal, issue, pages | Nucleic Acids Res, Vol. 53, Issue 5, Year 2025 |
| Publish date | Feb 27, 2025 |
 Authors | Giada Finocchio / Irma Querques / Christelle Chanez / Katarzyna J Speichert / Martin Jinek /  |
| PubMed Abstract | CRISPR-associated transposon (CAST) systems employ CRISPR-Cas systems as RNA-directed targeting modules for site-specific transposon DNA insertion. Among them, type I CASTs rely on the coordinated ...CRISPR-associated transposon (CAST) systems employ CRISPR-Cas systems as RNA-directed targeting modules for site-specific transposon DNA insertion. Among them, type I CASTs rely on the coordinated action of the guide RNA-bound Cascade complex and the transposon proteins TniQ, TnsC, and TnsAB. The interaction between the transposase TnsAB and the ATPase TnsC is crucial for transposition activity, yet the underlying molecular details have remained elusive. Here, we investigate the type I-B CAST system from Peltigera membranacea cyanobiont. Cryo-electron microscopic structures of TnsC and its complex with the C-terminal region of TnsAB reveal that TnsC forms a heptameric ring that recruits TnsAB by interacting with its C-terminal tail. In vitro binding assays indicate that TnsAB exclusively interacts with the TnsC heptamer without inducing its disassembly, in contrast to type V-K CAST systems. Mutational analysis of key structural features corroborates the significance of TnsC multimerization and TnsB interaction for transposon activity in vivo. Altogether, these findings offer detailed structural and functional insights into the molecular mechanism of type I-B CAST, with the aim of facilitating their development as genome engineering tools. |
 External links | Nucleic Acids Res / PubMed:40103227 / PubMed Central |
| Methods | EM (single particle) |
| Resolution | 3.2 - 4.47 Å |
| Structure data | EMDB-50932, PDB-9g0f:  EMDB-50933: CryoEM volume of a heptameric stretched ring conformation of the PmcTnsC-dsDNA-AMPPNP complex  EMDB-50934: CryoEM volume of an octameric stretched ring conformation of the PmcTnsC-dsDNA-AMPPNP complex EMDB-51458, PDB-9gmz: |
| Chemicals |  ChemComp-ANP:  ChemComp-MG: |
| Source |
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 Keywords | DNA BINDING PROTEIN / AAA+ ATPase / CRISPR / Tn7-like transposon / CAST / transposase / Integrase |
peltigera membranacea (fungus)
escherichia coli (E. coli)