From bench to barn
January 20, 2011 § Leave a comment
Has synthetic biology ever gone further than this? If so, I’m not aware of it. A recent paper (Kemmer et al. 2010. A designer network coordinating bovine artificial insemination by ovulation-triggered release of implanted sperms. J. Controlled Release PMID: 21108977) starts by engineering a simple hormone-response circuit and ends with pregnant cows.
First, a little motivation. Huge industries (milk, cheese, beef) depend, basically, on getting cows pregnant. But successfully inseminating a cow can be quite tricky, because there’s no easy way to know exactly when a cow is ovulating. Since the oocyte isn’t available for fertilization for very long, and sperms are also fairly fragile, it’s easy to miss the perfect moment. There have been attempts to encapsulate sperm in protective coats to extend their lifespan, but — although this does seem to preserve the viability of the sperm — the challenge of releasing them at the right time remains.
The most reliable signal that a cow (or most female vertebrates) is about to ovulate is a surge of luteinizing hormone, which triggers oocyte release (in cows) 22 to 31 hours later. Kemmer et al reasoned that if they could key the release of encapsulated sperm to the surge of luteinizing hormone, they might be able to coordinate the delivery of the two active gametes, so that sperm and egg would arrive in the same place at the same time. So they set out to build a circuit that would respond to luteinizing hormone by producing an enzyme (in the final form of the circuit, cellulase) that would chew up the capsule protecting the sperm. The capsules they chose were cellulose-sulfate implants, and the luteinizing-hormone-responsive circuit was expressed in human embryonic kidney HEK-293 cells.
The design of the circuit is shown in the figure. It depends on an exogenous luteinizing hormone receptor (LHR) being able to connect appropriately with the adenyl cyclase in HEK-293 cells, producing cyclic AMP (cAMP). cAMP then activates the cAMP-responsive-element binding protein (CREB), which in turn triggers the transcription of a cellulase gene driven by a cAMP-responsive element (CRE)-containing promoter. Once the cellulase protein is produced, it’s secreted, and it can therefore chew up the cellulose encapsulating the sperm.
Kemmer et al. showed first that their circuit gave nice dose-responsive behavior when the cells are cultured normally. Next, they inserted the engineered cells into the matrix of the sperm-containing cellulose-sulfate implants, and showed that they’re still able to produce the cellulase in response to the luteinizing hormone signal. The final step needed to show that the whole approach is feasible is to show luteinizing-hormone-dependent destruction of the cellulase capsules; this works too, and they provide movies to prove that the encapsulated sperm are still alive and kicking after release.
But can all these steps work together? Yes, they can. As a grand finale, Kemmer et al. tested the whole cascade: they implanted their combination sperm-protection/triggered release device (encapsulated sperm plus the engineered cells) in twelve Swiss dairy cows, either at the time of ovulation or 24 hours earlier, and allowed the luteinizing hormone produced by the ovulating cow to trigger the production of the cellulase, which then released the encapsulated sperm. The final figure of the paper is a micrograph of successfully-fertilized actual cow embryos, obtained from the uterus of a cow treated with their sperm capsule/cell-based release device.
What the paper doesn’t show is whether this fertilization strategy indeed has benefits relative to the old method of introducing the sperm when the farmer thinks — based on behavioral cues that presumably evolved to give a bull a decent chance of knowing when a cow is fertile — that it’s the right time. I assume that this is because getting a statistically sensible answer to this question would involve testing the implants on hundreds of cows, not just the 12 reported here; I can understand the authors not wanting to make quite such a large investment in the first report. Still, this paper takes an impressively large step towards actual commercial realism. (Thanks to Roland Eils for pointing this one out!)
Kemmer C, Fluri DA, Witschi U, Passeraub A, Gutzwiller A, & Fussenegger M (2010). A designer network coordinating bovine artificial insemination by ovulation-triggered release of implanted sperms. Journal of controlled release : official journal of the Controlled Release Society PMID: 21108977