Temperature-Switchable Genome Editors from Extremophile-Derived Integrases
- 1Sandia National Labs
Abstract
Integrases are site-specific recombinases encoded by phages and other mobile genetic elements. They mediate DNA integration, excision, and inversion between cognate attachment (att) sites. Although integrases are powerful tools for genetic engineering and synthetic biology, most systems lack intrinsic mechanisms that limit activity after expression, creating potential for unintended recombination. We hypothesized that extremophiles could provide temperature-responsive integrases because their enzymes evolved under selective pressure to operate within the thermal ranges experienced by their hosts. To test this concept, we linked integrase-att pairs from 458,683 prokaryotic genome assemblies to curated host growth-temperature metadata. This analysis revealed temperature-associated structure among integrase clusters and established a candidate pool for testing temperature-responsive recombinases. GC content in tyrosine integrase-associated attB sites showed a modest increase in higher-temperature hosts. We developed an inversion assay using a single-copy reporter plasmid and sacB counterselection to quantify integrase activity across temperatures. Thermophile derived integrases from Thermus thermophilus and Geobacillus stearothermophilus displayed hot-ON/cold-OFF activity profiles, whereas an integrase derived from the psychrotroph Pseudomonas cerasi showed cold-ON/hot-OFF activity. Together, these results establish host thermal niche as a guide for discovering intrinsically temperature switchable integrases and provide a foundation for engineering thermally controlled genome editing systems.
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BibTeX
@misc{arnold2026temperatureswitchable,
title = {{Temperature-Switchable Genome Editors from Extremophile-Derived Integrases}},
author = {J. Arnold and I. G. Romano and D. Del Curto and E. K. Wilbourn and E. L. Torrance and V. A. Fierro and J. B. Ricken and K. P. Williams and J. S. Schoeniger and J. CAHILL},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.10.749218},
url = {https://www.biorxiv.org/content/10.64898/2026.09.10.749218v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Temperature-Switchable Genome Editors from Extremophile-Derived Integrases
AU - J. Arnold
AU - I. G. Romano
AU - D. Del Curto
AU - E. K. Wilbourn
AU - E. L. Torrance
AU - V. A. Fierro
AU - J. B. Ricken
AU - K. P. Williams
AU - J. S. Schoeniger
AU - J. CAHILL
PY - 2026
DA - 2026/09/10
PB - bioRxiv
DO - 10.64898/2026.09.10.749218
UR - https://www.biorxiv.org/content/10.64898/2026.09.10.749218v1
AB - Integrases are site-specific recombinases encoded by phages and other mobile genetic elements. They mediate DNA integration, excision, and inversion between cognate attachment (att) sites. Although integrases are powerful tools for genetic engineering and synthetic biology, most systems lack intrinsic mechanisms that limit activity after expression, creating potential for unintended recombination. We hypothesized that extremophiles could provide temperature-responsive integrases because their enzymes evolved under selective pressure to operate within the thermal ranges experienced by their hosts. To test this concept, we linked integrase-att pairs from 458,683 prokaryotic genome assemblies to curated host growth-temperature metadata. This analysis revealed temperature-associated structure among integrase clusters and established a candidate pool for testing temperature-responsive recombinases. GC content in tyrosine integrase-associated attB sites showed a modest increase in higher-temperature hosts. We developed an inversion assay using a single-copy reporter plasmid and sacB counterselection to quantify integrase activity across temperatures. Thermophile derived integrases from Thermus thermophilus and Geobacillus stearothermophilus displayed hot-ON/cold-OFF activity profiles, whereas an integrase derived from the psychrotroph Pseudomonas cerasi showed cold-ON/hot-OFF activity. Together, these results establish host thermal niche as a guide for discovering intrinsically temperature switchable integrases and provide a foundation for engineering thermally controlled genome editing systems.
ER -