Single cell variability of CRISPR-Cas interference and adaptation

Single cell variability of CRISPR-Cas interference and adaptation

McKenzie, R. (TU Delft BN/Stan Brouns Lab; Kavli institute of nanoscience Delft; AMOLF)
Keizer, Emma M. (Wageningen University & Research)
Vink, J.N.A. (TU Delft BN/Stan Brouns Lab; Kavli institute of nanoscience Delft)
van Lopik, Jasper (Kavli institute of nanoscience Delft; Student TU Delft)
Büke, F. (TU Delft BN/Greg Bokinsky Lab; Kavli institute of nanoscience Delft; AMOLF)
Kalkman, Vera (Kavli institute of nanoscience Delft; Student TU Delft)
Fleck, Christian (Albert-Ludwigs-Universität Freiburg)
Tans, S.J. (TU Delft BN/Sander Tans Lab; Kavli institute of nanoscience Delft; AMOLF)
Brouns, S.J.J. (TU Delft BN/Stan Brouns Lab; Kavli institute of nanoscience Delft)

2022

While CRISPR-Cas defence mechanisms have been studied on a population level, their temporal dynamics and variability in individual cells have remained unknown. Using a microfluidic device, time-lapse microscopy and mathematical modelling, we studied invader clearance in Escherichia coli across multiple generations. We observed that CRISPR interference is fast with a narrow distribution of clearance times. In contrast, for invaders with escaping PAM mutations we found large cell-to-cell variability, which originates from primed CRISPR adaptation. Faster growth and cell division and higher levels of Cascade increase the chance of clearance by interference, while slower growth is associated with increased chances of clearance by priming. Our findings suggest that Cascade binding to the mutated invader DNA, rather than spacer integration, is the main source of priming heterogeneity. The highly stochastic nature of primed CRISPR adaptation implies that only subpopulations of bacteria are able to respond quickly to invading threats. We conjecture that CRISPR-Cas dynamics and heterogeneity at the cellular level are crucial to understanding the strategy of bacteria in their competition with other species and phages.

agent-based simulations
single-cell analysis
spacer acquisition
time-lapse microscopy
type I CRISPR-Cas

http://resolver.tudelft.nl/uuid:55c3fe10-2ccc-475a-b19e-2ed24cfb529a

https://doi.org/10.15252/msb.202110680

1744-4292

Molecular Systems Biology, 18 (4)

Institutional Repository

journal article

© 2022 R. McKenzie, Emma M. Keizer, J.N.A. Vink, Jasper van Lopik, F. Büke, Vera Kalkman, Christian Fleck, S.J. Tans, S.J.J. Brouns