Biology subjects

Del Curto, D.

Publications and source records attributed to Del Curto, D..

2 recordsLinked to original sources

bioRxiv2026

Temperature-Switchable Genome Editors from Extremophile-Derived Integrases

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.

molecular biology↗

bioRxiv2026

Re-evaluating Reported Pseudolysogeny in Phage T3: T3 and T7 Show Similar Propagation Responses to Nutrient Limitation and Media Switching

Pseudolysogeny is a latent state in which phage development is delayed after infection and has been proposed to promote phage persistence under unfavorable conditions. Virulent phage T3 has been reported to establish pseudolysogeny after infecting starved E. coli, then resume lytic replication following transfer to nutrient-rich media, a phenotype linked to the T3 SAMase gene. Here, we revisited the findings of Krueger et al. (1975) to test pseudolysogeny in T3 and examine phage propagation under nutrient-limited conditions. Both T3 and T7 showed impaired propagation under nutrient limitation, with the most stringent conditions causing substantial losses in recoverable infective centers. T3 was modestly more resilient than T7 under these conditions, but we were unable to reproduce the reported phenotype in which T3 remained latent while T7 replicated normally. Supplementation of minimal medium with small amounts of LB supported propagation of both phages, and a repeat experiment designed to more closely match the historical protocol, including post-adsorption reduction of extracellular phage carryover, likewise failed to reveal a T3-specific pseudolysogenic state. Together, our results indicate that, in this experimental system, phage propagation dynamics are more consistently explained by nutrient conditions and media switching than by starvation prior to infection. These findings suggest that the previously reported T3 pseudolysogeny phenotype may depend on additional environmental or methodological factors and underscore the importance of revisiting historically reported phage behaviors using modern controls.

microbiology↗