A mammalian cell phone

Sachin Singh Rawat

Bacchus W, Lang M, El-Baba MD, Weber W, Stelling J & FusseneggerSynthetic two-way communication between mammalian cellsNature Biotechnology, Published online 16/9/12

Introduction

Synthetic Biology of Animal Cells

Switches Logic gates Timers, time-

delay circuits Counters Oscillators Biocomputers

Discovery of anti-TB compounds

Design of bacteria that kill cancer cells

Prosthetic network for treating arthritis

Light-controlled regulation of glucose homeostasis

Developments Recent successes

Cell-cell interactions

enable cell populations to… Communicate during development Adapt to the environment Form and/or recognize patterns

engineered cell interactions include… Quorum-sensing dependent gene

expression Programmed pattern formation in yeast

Making the call

The two modules

Sender cells communicate through the essential amino acid L-tryptophan

Sender module: HEK-293 cells with pWB32, so they constitutively express ß-subunit of E. coli tryptophan synthase

Indole, present in media, was converted into the signalling molecule – tryptophan

Receiver module: HEK-293 cells with pWB24 to express a target-gene by L-tryptophan-inducible sensor circuit

The modules at work

Characterizing the communication

Sender (HEK-TrpB) and receiver (HEK-SEAP) cells co-cultivated in varying ratios

Communication established only if ratio of sender to total cell number is higher than tryptophan uptake rate per cell divided by tryptophan production rate of sender cell

Communication strength increase linearly with receiver cell concentration

The response was indole-dependent

Validating the interactions

Heat maps showing ideal conditions formaximum communication strength

Getting a call back

Cellular crosstalk

Developed combined sender/receiver as well as processor cells, creating a two-way, closed loop feedback circuit

The cells (HEK-TrpB/SEAP) were engineered for constitutive tryptophan synthase expression and acetaldehyde-inducible SEAP expression

The processor cells, HEK-ADH, were transgenic for L-tryptophan-inducible expression of alcohol dehydrogenase to convert ethanol in medium into acetaldehyde

The processing

The L-tryptophan signal triggers expression of reporter SAMY as well as production of volatile acetaldehyde

The signal is broadcasted back to sender/receiver cells which signal receipt of message by acetaldehyde-triggered SEAP expression

The system’s performance is quantized by dynamically monitoring the reporter

Implementing two-way communication

A therapeutic call

Producing blood vessels

VEGF and Ang1 act in a sequential and complementary manner to produce blood vessels

The communication device was modified: VEGF instead of SAMY and Ang1 instead of SEAP

Processor cells were engineered for constitutive tryptophan synthase expression and acetaldehyde-inducible Ang1 expression

The device programmed transient permeability increase after 24h, correlating with VEGF expression, and reversed after 48h by Ang1 expression

“ Such bidirectional and multistep communication

devices, inspired by naturally occurring fundamental processes, are key to the design and construction of next-generation synthetic mammalian networks that will likely comprise populations of specialized cells interconnected by orthogonal communication devices to fulfill complex computational operations

’’

Thank you