IGEM:Caltech/2008/Ideas

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iGEM 2008

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Big Ideas Bacteriofood After reviewing UC Berkeley's BactoBlood in the 2007 iGEM Jamboree, Bacteriofood could be used to produce and carry vital nutrients that can maintain a healthy human. Most useful in third world countries where food is hard to come around. Bacteria being easy to grow and maintain, could be a simple way to feed the underfed. Just trying to throw out any idea that comes across. Some thoughts - could we make bacteria more nutritious? I.e. you'd probably need an E. coli culture plus some list of vitamins/minerals/etc. Could we come up with that list of necessary extras and then engineer E. coli to remove a couple items from the list? Cholesterol Degradation Could we make a strain of E. coli that circulates in the blood stream and feeds on cholesterol plaques as its food source? I know part of the 2007 UC Berkeley project, the bactoblood one, was to make E. coli not trigger sepsis when in the bloodstream. This chassis would be very useful for a project like this. Similar idea, but use it for diabetes also and have E. coli feed off sugar. Maybe both? Taking advantage of noise From an engineering perspective, noise is usually a problem we need to overcome. But in biology, a uniform response simply means that your entire population gets wiped out simultaneously (think agricultural monocultures). Biology often hedges its bets by producing a diversity of phenotypes, some fraction of which are favored in any given environment [1, 2]. Can we do the same - continually producing a range of phenotypes and allowing the environment to select the ones best suited at that moment? I've always liked the idea of directed evolution. Usually mutation occur at random throughout the genome, but a lot of the time it seems researchers only want a few genes to change. Instead of using the time consuming site directed or random mutagenesis, could we engineer a bacteria mutation a specif portion of its genome at an accelerated rate? I'm thinking something akin to the cre-lox in which the researcher could flank the stretch of DNA to be mutated by two DNA sequences, and then an enzyme or set of enzymes would catalyze the accelerated mutation of only that portion. The ter sites and Tus proteins are normally used to terminate E. coli genome replication by slowing down and kicking off the polymerase. Maybe a toned down ter-Tus system would be enough to screw up the polymerase as it passes through, but not totally derail it. There are mutator strains that increase the genome-wide mutation rate. Ignoring implementation, I think it would be tricky (not impossible, but tricky) to find a niche for targeted in vivo mutation. Mutator strains are generally used when you don't know what your target is. If you know what your target is, the question is what the advantage is over error prone PCR. Best I can come up with is that you might want to hit multiple genomic locations simultaneously, such that error prone PCR would be really time consuming. But those situations seem relatively rare. Endosymbiosis Attempt to recreate endosymbiosis by introducing a bacteria (E. coli or B. subtilis or other) into yeast. This would essentially be creating an artificial organelle. I know yeast can grow without mitocondria, being called petite yeast. What new things could they do with an entirely new organelle? I'd imagine this could be done with some sort of double selection. We could take an auxotrophic yeast strain, say for uracil, and then "inject" a bacteria that secretes the missing enzyme in the uricil pathway. The missing enzyme would be secreted directly into the yeast cytoplasm by the bacteria. The bacteria in turn could gain shelter from some antibiotic in the medium. Only by living inside the yeast cell could both the yeast and bacterium survive. Could we hijack an existing organelle rather than introducing a new one? Hacking mitochondria, say? Sounds interesting? What do you mean by "hacking?" What would the new organelle be hacked to do? Would it be like transplanting an organelle from one species to another? (ie - giving chloroplasts to yeast?) Well, there's always two questions - first, what can we do in a summer? And second, how can you spin the summer project as a model system for something more exciting? For the summer - depends on what people have done previously (and I'm not really sure what the current state of the field is). Might be as simple as showing that we can reprogram a specific organelle (worst case, just adding a marker). Longer term, you can make the same argument as UCSF last year - segregate your engineered system off into a separate compartment will less interaction with the host. CO2 Bacteria It seems many researchers are looking to find or develop a biofuel that can sustain our cars, planes, machines, etc. As time passes by, we are still polluting our atmosphere with greenhouse gases, so why not develop a bacteria that will *chew up* or turn CO2 into a less harmful substance. (Is there any bacteria that consumes CO2?) Maybe we could create filters of bacteria to be put along our car exhausts, etc. I don't have much knowledge about greenhouse gases, but just an idea to throw out. You're talking about photosynthesis here - fixing CO2. It's an important problem, but one that industry is already working on (see this for instance). Cool outputs Pimp my E. coli We can trick out our E. coli similar to our car -- spinners, rims, spoilers, etc. One of many modifications is have the cells express reflectins on their surface. Reflectins are highly reflective proteins only found in squid reflective tissue [3]. Random thought - we can probably localize proteins to the poles of the cell. So make the tips a different color from the body If I remember correctly from last summer, UCSF used Pleckstrin Homology (PH) Domains for localizing different fluorescent proteins to different parts of the cell, such as the cell membrane. Could we do polka dots? Make protein clusters in the membrane? Changing shape Can we make E. coli become helical using crescentin[4] from Caulobacter? Motility As an output, we could turn motility on and off[5]. Stickiness Make the cells express an adhesive protein[6] or turn on biofilm EPS expression (absent any other biofilm phenotype). Random thoughts A more analog device Digital and analog responses, a common feature of electrical circuits, are also displayed by biological networks. While recent research has focused on engineering a more digital response using cooperativity or transcriptional cascades, we go the other way and engineer a more analog device. One way to do this is to express mutiple tetR variants that have different affinities for aTc, the primary inducer used in bacteria. I have found one paper that reports tetR variants with different affinities [7], although I'm sure more can be tracked down. Others Using network motifs? Uri Alon has done some interesting stuff with it. Should we need to use time-delayed releases, might be handy. http://www.nature.com/nrg/journal/v8/n6/pdf/nrg2102.pdf Using ribozymes? No idea where this is going or if its valid, but who said that we needed to follow how the cell does it.... perhaps RNAi or one of those unique forms could be used as a branching statement in what seems to be a rather linear DNA programming methodology (or at least from first glance). Kind of like this[8, 9]?: Random Number Generator FimE inverts a specific stretch of DNA, defined by a pair of sequence elements (IRR and IRL), forming a DNA loop between the two elements[10]. If we add multiple copies of one of these elements (one IRR, two IRL), would FimE randomly choose one of the sites (one IRL out of the pair) to invert between? Either choose one of several promoters to attach to a given gene, or one of several genes to attach to a given promoter. Then, can we tune the probability (from, say, 60:40 to 80:20 to 20:80)? Ideally do this dynamically (based on some small molecule) - use proteins that bend DNA to affect the probability of loop formation. Population Variability Slipped-strand mispairing (SSM)[11] can produce a heritable variation in the expression from a promoter. Roughly one in 1000 divisions produces a change in expression. Couple this expression to a selectable/counterselectable marker. Under any given condition (selection, say), the population thrives, but with a small group of the opposite phenotype (non-expressing). Switch conditions (to counterselecting), and the population can use these revertants to recover. The switching is stochastic by nature and can be directly compared to both natural [1] and synthetic [2] systems that utilize stochastic switching to adapt to variable and fluctuating environments. Under constantly varying conditions, most circuits would die. These cells, though, can adapt and pass that adaptation on to their descendants. References Süel GM, Garcia-Ojalvo J, Liberman LM, and Elowitz MB. An excitable gene regulatory circuit induces transient cellular differentiation. Nature. 2006 Mar 23;440(7083):545-50. DOI:10.1038/nature04588 | Acar M, Mettetal JT, and van Oudenaarden A. Stochastic switching as a survival strategy in fluctuating environments. Nat Genet. 2008 Apr;40(4):471-5. DOI:10.1038/ng.110 | Crookes WJ, Ding LL, Huang QL, Kimbell JR, Horwitz J, and McFall-Ngai MJ. Reflectins: the unusual proteins of squid reflective tissues. Science. 2004 Jan 9;303(5655):235-8. DOI:10.1126/science.1091288 | Margolin W. Bacterial shape: concave coiled coils curve caulobacter. Curr Biol. 2004 Mar 23;14(6):R242-4. DOI:10.1016/j.cub.2004.02.057 | Topp S and Gallivan JP. Guiding bacteria with small molecules and RNA. J Am Chem Soc. 2007 May 30;129(21):6807-11. DOI:10.1021/ja0692480 | Hwang DS, Yoo HJ, Jun JH, Moon WK, and Cha HJ. Expression of functional recombinant mussel adhesive protein Mgfp-5 in Escherichia coli. Appl Environ Microbiol. 2004 Jun;70(6):3352-9. DOI:10.1128/AEM.70.6.3352-3359.2004 | Kintrup M, Schubert P, Kunz M, Chabbert M, Alberti P, Bombarda E, Schneider S, and Hillen W. Trp scanning analysis of Tet repressor reveals conformational changes associated with operator and anhydrotetracycline binding. Eur J Biochem. 2000 Feb;267(3):821-9. DOI:10.1046/j.1432-1327.2000.01063.x | Win MN and Smolke CD. A modular and extensible RNA-based gene-regulatory platform for engineering cellular function. Proc Natl Acad Sci U S A. 2007 Sep 4;104(36):14283-8. DOI:10.1073/pnas.0703961104 | An CI, Trinh VB, and Yokobayashi Y. Artificial control of gene expression in mammalian cells by modulating RNA interference through aptamer-small molecule interaction. RNA. 2006 May;12(5):710-6. DOI:10.1261/rna.2299306 | Ham TS, Lee SK, Keasling JD, and Arkin AP. A tightly regulated inducible expression system utilizing the fim inversion recombination switch. Biotechnol Bioeng. 2006 May 5;94(1):1-4. DOI:10.1002/bit.20916 | Torres-Cruz J and van der Woude MW. Slipped-strand mispairing can function as a phase variation mechanism in Escherichia coli. J Bacteriol. 2003 Dec;185(23):6990-4. DOI:10.1128/JB.185.23.6990-6994.2003 |