Team:Chiba/Project

E.coli Time Manager Since 2008

Introduction

決められた時間の長さを生物が認識する機能作りは、生物工学において切望されてきた。例えば物質合成や物質感知などの優れた生物機能を時間制御することができれば、ドラッグデリバリー(定めた時間に薬となる有効成分を私たちの体内でださせる)や健康診断(定めた時間に体内の有害物質濃度を感知できれようになる)活かすことができるだろう。 しかし細胞毎の時間制御ができたとしても、集団で発揮させることができなければ、時間制御の効果は薄くなるのではないか。そこで、私たちは、「生物が集団で発揮するタイマーシステム」を作ろうと考えた。

Project Design

シンクロシステムのいらない、同一系内で使える数種類のタイマーを作るために、私たちは以下の様なデザインを考えている。

複数細胞の同時コントロール

時間を扱った2008年のMelbourneや、2007・2008年のParisチーム、2008年のNYMUチームのプロジェクトでは、細胞集団の振る舞いを同時にするためにシンクロシステムが必要だった。そこで千葉はシンクロシステムを使わずに複数の細胞を同一のタイミングでコントロールできるシステムを作ろうと考えた。ここで使うのが細胞間のSignaling Systemだ。このSignaling Systemによって系内の何万モノ細胞に、「感じている時間」を共有させることができるようにしている。

Signaling System

In this project, we use acylated homoserine lactones (AHLs), signaling molecules used for quorum sensing in gram negative bacteria. Senders express LuxI or similar enzymes, which catalyze the production of AHLs, under the control of a constitutive (Tet) promoter. Each cell thus generates AHL more or less at a constant rate. AHL can freely permeate cell membranes and are detected by neighboring cells. Receivers constitutively express LuxR proteins (or a similar ortholog), the protein that detects AHL concentrations. When AHLs bind LuxR proteins, the AHL-LuxR complex activates the Lux promoter. The threshold [AHL] at which switching occurs is determined by the affinity of AHL for the particulr LuxR ortholog. (more about quorum sensing)

Constructing A Delay Switch, Multiple Ways (Since 2008)

We are constructing delay switches to control/preset the timing of target gene expression. Our project uses two classes of bacteria: senders and receivers. Senders produce signaling molecules, and receivers are activated only after a particular concentration of this molecule is reached. The combinatorial use of senders/receivers allows us to make a‘switching consortium’which activates different genes at the preset times.

In principle, there are three ways to delay the activation of chemical communications;

1. Silencing the Speakers: Rate of signal accumulation down-regulated, for instance, by slowing down the signal generators.
2. Desensitize Receivers: Switching threshold elevated, for instance, by using insensitive receiver/ reporter systems.
3. Partial Blocking: Decreasing the by chewing the signal up.

Inter-species communications!

We decided to go for the strategy inspired by Japanese-classic experience; Whenever we speak to somebody in English, we often experience a certain delay in activating the communication. We though this is exactly what we pursued in. See Exp #4. Same applies to the reverse, too. When somebody speaks to us, we definitely need some time (sometime infinite) to get activated. This is all in spite that he/ she was loud and clear enough. The less affinity (perception) we have to English, the longer we need to activate them.See exp #5.

-> Get more information about a Delay Switch!

Constructing A Flow of Activation

Conclusions