(6a) Molecular Machines As Tools for Engineering with Biological Systems

Before the establishment of engineering disciplines, the biological world had long designed a menagerie of molecular machines capable of performing catalysis, separations, sensing, and locomotion. A number of these machines either act on or incorporate nucleic acids; information-dense biomolecules that both contain all of the instructions for life and perform catalytic and regulatory activities. Control of these molecular machines has led to amazing technologies such as PCR, gene editing, cloning, etc. that have made the entire discipline of bioengineering possible and promise to generate new tools and technologies.

My interests lie in fundamentally understanding the molecular machines we have available for engineering new biological systems, discovering new systems, and enabling new biological technologies. To achieve these goals, I have studied how RNA folding leads to structures that affect functional changes in molecular systems [1-6] during my PhD and transitioned to discovering and understanding new mechanisms in CRISPR biology [7] during my postdoctoral research.

My future lab will focus on both discovery and application of new molecular machines to develop new technologies for disease detection, biodefense, and biocomputation with a particular focus on tools derived from CRISPR and RNA enzymes and regulators. The high specificity of CRISPR-Cas proteins and RNA regulators for nucleic acids sequences allows for facile engineering by simply changing nucleic acid sequences, while maintaining the high capacity for diversity afforded by a deep sequence space. Thus, a major focus of my lab will be engineering new nucleic acid-based technologies.

PhD Research:

My graduate research was performed at Cornell University with Prof. Julius Lucks in Chemical and Biomolecular Engineering. During my time at Cornell, my work focused on two elements of RNA synthetic biology: 1) designing synthetic systems for biological process control and 2) investigating the mechanisms that RNA regulators use to control gene expression. To understand the RNA structure-function relationship, I developed an improved version of SHAPE-Seq [1,2], a high-throughput technique for measuring RNA structural features, that was less restrictive than previous techniques. These early improvements led to further breakthroughs in measuring RNA structures inside of living cells [3] and within cell lysates [4]. My graduate work culminated in a new technique for measuring the RNA structures that form during the process of transcription [5,6] that give regulatory RNAs their unique properties for controlling gene expression.

Postdoctoral Research:

As a postdoctoral fellow, I sought to gain a more fundamental comprehension of basic biological processes and to join the CRISPR community, which has been the source of myriad biological tools. Therefore, I joined the lab of Prof. Jennifer Doudna, in the department of Molecular and Cell Biology, which had a rich history studying the RNA structure-function relationship and discovering new CRISPR biology and applications. My postdoctoral work has been focused on the discovery of anti-CRISPR proteins with high-throughput approaches, harnessing both bioinformatics and experimental screens. These efforts have led to the discovery of the first anti-CRISPR proteins that inhibit the Cas12 (Cpf1) nuclease, which is commonly used for gene editing applications as an alternative to Cas9, as well as other new anti-CRISPRs that inhibit Cas9. Other research is currently underway to create a high-throughput screening technique amenable to screening genomes more generally for new protein functions.

Successful Proposals:

NIH F32 Fellowship, Life Sciences Research Fellowship, DARPA SafeGenes

Education:

Ph.D., Chemical and Biomolecular Engineering, Cornell University, Ithaca, NY, 2016

B.S., Chemical Engineering, Rensselaer Polytechnic Institute, Troy, NY, 2011

Teaching Interests:

My goal in teaching is to combine classical Chemical Engineering topics with modern applications in bioengineering, tying these disciplines together to create exciting example-driven learning that draws from real data and experiments. Of the core curriculum, I am best suited to teach subjects most penitent to biological systems such as kinetics, thermodynamics, separations, or control theory. I am also excited about the prospect of developing and/or teaching cross-disciplinary courses that focus on bioengineering topics within synthetic biology, bioinformatics, and biochemistry with an engineer’s lens to motivate questions about downstream applications, increased throughput/scaleup, and alternative approaches. Throughout my courses, I plan on creating an atmosphere of shared learning and establishing a dialog between myself, the students, and each other. I believe that classroom discussions promote student-student learning and allow easier assessment of student knowledge and where I can improve as a teacher. Additionally, I am interested in developing hands-on projects to solidify the materials delivered during lectures.

As a mentor to graduate students and postdoctoral scholars, I plan on synthesizing the different approaches to mentorship that I experienced during my own training. I believe that a lab with as a high degree of diversity (both cultural and scientific) as possible is immensely beneficial to lab output, morale, and peer-directed learning. In general, I will focus on fostering a culture of lab cohesiveness through celebrated group accomplishments, intra-lab collaborations and teamwork, and out-of-lab events to strengthen bonds between lab members.

Invention Disclosures:

Blackburn-Marino, N., Bondy-Denomy, J.B., Watters, K.E., Doudna, J.A. Proteins that Inhibit Cas12a (Cpf1), a CRISPR-Cas DNA Nuclease (pending)

(Others pending)

Selected Publications (out of 14):

1. Watters, K. E., et al. Characterizing RNA structures in vitro and in vivo with selective 2'-hydroxyl acylation analyzed by primer extension sequencing (SHAPE-Seq). Methods 103, 34–48 (2016).
2. Loughrey, D.*, Watters, K. E.*, Settle, A. H. & Lucks, J. B. SHAPE-Seq 2.0: systematic optimization and extension of high-throughput chemical probing of RNA secondary structure with next generation sequencing. Nucleic Acids Research 42, (2014).
3. Watters, K. E., Abbott, T. R. & Lucks, J. B. Simultaneous characterization of cellular RNA structure and function with in-cell SHAPE-Seq. Nucleic Acids Research 44, e12–e12 (2016).
4. Watters, K. E. et al. Probing of RNA structures in a positive sense RNA virus reveals selection pressures for structural elements. Nucleic Acids Research (2017). doi:10.1093/nar/gkx1273
5. Watters, K. E., Strobel, E. J., Yu, A. M., Lis, J. T. & Lucks, J. B. Cotranscriptional folding of a riboswitch at nucleotide resolution. Nature Structural & Molecular Biology 23, 1124–1131 (2016).
6. Strobel, E. J., Watters, K. E., Nedialkov, Y., Artsimovitch, I. & Lucks, J. B. Distributed biotin-streptavidin transcription roadblocks for mapping cotranscriptional RNA folding. Nucleic Acids Research 45, e109 (2017).
7. Watters, K.E., et al. Systematic discovery of natural CRISPR-Cas12 inhibitors. under review at Science as of June 2018.