Building a cell-cycle counter

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Contents

Overview

The goal of my project is to engineer a cell cycle counter. Such a counter, when implemented in living cells, would allow them to count, store and display the number of time they divided. A cell cycle counter would thus enable sorting or tracking cells according to the number of time they divided. This would have profound impact for aging and cancer research or developmental biology.

  • Digital counters

Digital counters are built from multiple bits, each bit being able to take two values, zero or one. The cell cycle counter will be engineered as a biological analog of the simpler digital counter,the ripple (or asynchronous) counter. The basic building block for digital counters are flip-flops. Flip- flops can resides in two stable states and are therefore able to serve as one bit of memory. The JK flip-flop toggles between the two stable states in response to a same periodic input (the clock), then hold the state till the next imput. A digital ripple counter is built by chaining multiple JK flip-flops. Only the first flip-flop of the chain is connected to the external input. This first flip-flop toggles in response to the external input with a frequency equals to the input frequency. The output of a each flip-flop serves as the clock for the following flip-flop, which toggles with half the frequency. By chaining various JK flip flops in this way a counter can be engineered, with a counting capacity of 2n where n is the number of flip-flops. Therefore we need to build a biological analog of the JK flip flop.

  • Memory storage

Memory will be stored within DNA by flipping a DNA sequence two different orientation (states) in a controlled fashion, by using site specific recombinases. Importantly, once written, the state does not require energy to be maintained. Therefore, instead of using bistable switches, I propose to use transient activation of a specific recombinase to flip the target DNA sequence. Find more at .

  • Building a biological analog to JK flip flop

A device will be engineered to produce an input once per cell cycle. This input will activate a device containing two pulse generators, each of them controlling switching toward one state. Depending on the current state, the input will activate the device which triggers switching to the opposite state. For each state, a specific effector will be produced which will act cooperatively with the input to trigger activation of the device controlling switching to the opposite state. The effector or the input alone cannot activate switching, which will thus be restricted to a precise time window, when both input and effector molecules are present.

  • Chaining

One of the two pulse generating devices will serve as the clock for the following bit, by producing a molecule analogous to the first input, which will associate with an effector produced by the following bit, thus triggering state switching in the second flip-flop similarly than in the first one, except that it will be with half the frequency.

Notes

What signals can we use to activate state writing?

  • Cell cycle dependent degradation (cyclin D-box, scf/skp2 of Beta Trcp degrons)
  • Cell cycle dependent transcription
  • Telomeres mimics
  • Sensors of cell cycle kinases or phosphatases activity.
  • Centrosomes duplication
  • etc...

references

counter architecture

more soon

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