The Technological Progress of CRISPR-Cas9

A common misconception about technological advancement is that they are ahistorical revolutions (Cook 1995). On this narrative, technological innovations emerge suddenly, without competition from other extant technologies, and are solely responsible for ushering in rapid, widespread social change. This misconception not only fails to account for the crucial social, political, and moral values that often drive technological change (a topic which I will not address here), but it also mischaracterizes the nature of technological progress. Contrary to this misconception, innovative progress is often achieved by overcoming problems hindering extant technologies (Laudan 1984). The same kind of misconception has characterized popular reception of gene-modification technologies since the 20^th century. Today, the misconception continues with the new gene-editing technology, CRISPR-Cas9. In this post, I attempt to correct the misconception that CRISPR-Cas9 alone is bringing about radical change.

In the first half of the 20^th century when researchers began using x-rays to induce random mutations in the genomes of living systems, expectations were high. Many anticipated (prematurely) that radiation would enable us to direct evolution to our exact specifications – “New Life Made to Order,” as one headline read (Curry 2016; Early 1931). The misconception of x-ray breeding at the time painted a picture of social, economic, and even dietary revolution. Today, the misconceptions of CRISPR-Cas9 are often subtler. Contemporary discussions often emphasize CRISPR-Cas9’s superior precision, ease of use, and affordability without fully examining how these features compare to other competing technologies (i.e., zinc fingers, TALENs, RNAi) (“60 Minutes;” Smolenski 2015). These discussions often imply that CRISPR-Cas9 will singularly bring about widespread social change, when the reality is much more modest.

CRISPR (short for Clustered Regularly Interspersed Palindromic Repeats) – Cas is an adaptive immunity mechanism that evolved in microbial lineages that was recently (2013-15) modified by biologists to function as a gene-editing tool. As an immune defense, the CRISPR array is a sequence in DNA consisting of several hundred palindromic repeats separated by unique spacers, which function as a genetic memory of past viral infections. Flanking the CRISRP array is a cluster of genes that encode different Cas proteins, some of which (i.e., Cas9) cleave DNA. The CRISPR array identifies foreign DNA by complimentary binding to strands that match the spacer sequence and recruits the Cas9 protein where it makes a precise break in the invading genetic material. The CRISPR array’s precise targeting abilities have been adapted by researchers to work as a customizable single guide RNA (sgRNA) that can identify (almost) any gene. Cas9 has also been altered to cleave DNA in multicellular organisms. Together these components can be used to modify the genomes of many, varied organisms.

Gene-editing, however, requires more than precise cleavage, it also requires repair and “rewriting” of the broken strands. Researchers rely on DNA repair mechanisms that are inherent to an organism to accomplish this. DNA repair mechanisms rejoin the strands by inserting or removing nucleic acid bases. Gene-editing technologies enable researchers to “knockout” a gene – preventing its expression – or “knock-in” a gene – introducing a new gene that an organism would not otherwise have.

CRISPR-Cas9 is an advancement insofar as it has overcome some functional shortcomings of other molecular tools. A technology has functional shortcomings when it can’t meet all of our demands. For instance, a bridge has a functional shortcoming when it cannot bear the load of a semi-truck (Laudan 1984). In genetics, many of the molecular tools biologists use have functional shortcomings regarding precision. CRISPR-Cas9, zinc fingers and TALENs can be used to “knock out” or “knock in” genes; whereas, RNAi is commonly used to “knock down” a gene – by significantly reducing how much it gets expressed. These tools are relatively precise, but all have some degree of error. Early in CRISPR-Cas9’s history as a technology, a number of researchers anticipated that it would make fewer mistakes than zinc fingers and TALENs (Doudna et al. 2014). But as more research has unfolded, it now appears that CRISPR-Cas9’s precision is comparable to these tools (Schaefer et al. 2017). When compared to RNAi, however, CRISPR-Cas9 is more accurate (Smith et al. 2017). Yet, RNAi and CRISPR-Cas9 are used for slightly different purposes making them complimentary rather than competing tools (Barrangou et al. 2015).

Where CRISPR-Cas9 has overcome the shortcomings of zinc fingers and TALENs concerns economics. A technology has an economic shortcoming when it is too costly (in terms of money and time) to make and implement on a large scale. The protein-based guide components of zinc fingers and TALENs are extremely expensive and challenging to customize (Ledford 2015). By contrast, the time and expense required to synthesize the guide components for RNAi and CRISPR-Cas9 is much less. It is largely due to ease and affordability with which customized sgRNAs are produced that makes CRISPR-Cas9 preferable to zinc fingers and TALENs.

Some have claimed that CRISPR-Cas9’s ease-of-use is what will affect widespread change, but I remain skeptical. How CRISPR-Cas9 scores on this category will depend on the relative ease-of-use of other tools. Gene “knockout,” “knockdown,” and “knock-in” applications all present different experimental challenges, and there’s always tradeoffs between using one technology and using any other. Determining which tool is preferable for any given task is not always obvious. Simply because a tool is easier for a scientist to use (for some purposes) doesn’t mean it is world-changing. CRISPR-Cas9 is not so easy to use that just anyone can successfully implement it, nor has it overcome all the experimental hurdles that complex biological systems present.

CRISPR-Cas9’s functional and economic improvements only make sense in comparison to other molecular tools. Its low cost (compared to zinc fingers and TALENs) has brought about change in biology by making “knockouts” studies a more common methodology. Its improved precision (relative to RNAi) has also made “knockout” studies a viable alternative to gene “knockdowns.” However, this tool is comparable to these technologies on many other fronts, including applications that have serious ethical consequences (like making heritable modifications to mammalian genomes). If CRISPR-Cas9 is “revolutionary,” then this revolution is proceeding by small improvements to existing practices. That’s my take, what do you all think?

 

 

 

References

Barrangou R., A. Birmingham, S. Wiemann, R. Beijersbergen, V. Hornung, A. van Brabant Smith. 2015. “Advances in CRISPR-Cas9 Genome Engineering: Lessons Learned from RNA Interference.” Nucleic Acid Research, vol. 43, no. 7.

Cook, S. 1995. “The Structure of Technological Revolutions and the Gutenberg Myth.” In New Directions in the Philosophy of Technology. MA: Kluwer Academic Publishers.

Curry H. 2016. Evolution Made to Order. IL: The University of Chicago Press.

Doudna J., E. Charpentier. 2014. “The New Frontier of Genome Engineering with CRISPR-Cas9.” Science, vol. 346, issue 6213.

Early E. 1931. “X0Rays to Produce Giant Flowers, Fruits and Animals.” Public Ledger Sunday Magazine.

Laudan, R. 1984. “Cognitive Change in Technology and Science.” In The Nature of Technological Knowledge: Are Models of Scientific Change Relevant? MA: D. Reidel Publishing Co.

Ledford H. 2015. “CRISPR, The Disruptor.” Nature, vol. 522.

Schaefer K., W. Wu, D. Colgan, S. Tsang, A. Bassuk, V. Mahajan. “Unexpected Mutations after CRISPR-Cas9 Editing in vivo.” Nature Methods, vol. 14, no. 6.

Smith I., P. Greenside, T. Natoli, D. Lahr, D. Wadden, I Tirosh, R. Narayan, D. Root, R. Golub, A. Subramanian, J. Doench. 2017. “Evaluation of RNAi and CRISPR Technologies by Large-Scale Gene Expression Profiling in the Connectivity Map.” PLOS Biology, 15(11).

Smolenski J. 2015. “CRISPR/Cas9 and Germline Modification: New Difficulties in Obtaining Informed Consent.” The American Journal of Bioethics, 15(12).

“CRISPR: The Gene-Editing Tool Revolutionizing Biomedical Research.” 60 Minutes, produced by Nichole Marks. Associated producers, Kate Morris, Jamie Woods. April 29, 2018. https://www.cbsnews.com/news/crispr-the-gene-editing-tool-revolutionizing-biomedical-research/