OVERVIEW – Quantitative Biology of Gene Expression
When you order meatballs at an Ikea restaurant, you always get exactly 15 meatballs per order (fewer if you are outside the US). That is because your server can count. But how are protein serving sizes specified in a cell? And what determines the ideal abundance of each protein? Biological phenomena can be remarkably precise and quantitative despite being made of floppy molecules that live on thermal fluctuations. We use bacteria as a model to decipher the physical and chemical principles behind the precise control that cells have mastered through evolution. At the systems level, we are interested in how a bacterial proteome may be optimally configured for specific functions. Our approach combines technical development of quantitative measurements, genomics analysis, biophysical modeling, and synthetic biology. Our ultimate goal is to gain both high-level and mechanistic understanding of a cell in relation to its genetic blueprint.
Many major advances in modern biology have been driven by breakthrough biophysical techniques that enabled better quantitation of cellular constituents. In our lab, we are continuing to develop precision measurements for both bacterial transcriptome and proteome. Our Rend-seq approach (RNA end-enriched sequencing, Lalanne et al. Cell 2018) provides a high-resolution view of the diverse mRNA isoforms in vivo, which are shaped by alternative promoters, partial terminators, RNA processing, and decay. We have also developed methods to quantify the absolute rates of protein synthesis across the proteome (Li et al. Cell 2014 and Taggart et al. Cell Systems 2018). These and other unique measurements power our studies on the optimization of proteome and mRNA processing.
A fundamental challenge of systems biology is to combine individual, well-characterized, molecular players into a functional cell. Even a mundane-looking problem of mixing a few proteins to form a pathway can quickly become intractable due to the large combinatorial space of possible protein abundances. Our work over the past few years has revealed remarkable precision in cells’ ability to synthesize their proteomes. The genomes of both prokaryotes and eukaryotes encode quantitative instructions to produce stoichiometric amounts of components for protein complexes (Taggart et al. Cell Systems 2018). Taking advantage of this evolved precision, we found that most multi-enzyme pathways also have exquisitely preferred stoichiometry of proteins (Lalanne et al. Cell 2018). However, even for proteins with completely characterized molecular properties, we still lack frameworks to predict what constitutes appropriate abundances and how susceptible cells are to expression perturbations. We are tackling this problem using closely integrated experimental and theoretical approaches.
How do bacteria cells set protein levels so precisely? We study the molecular basis for differentially tuning expression among genes in the same operon. These mechanisms occur at both the transcriptional level, such as fractional termination due to different U-tract lengths (Lalanne et al. Cell 2018), and the translational level, such as variable translation efficiency due to long-range RNA secondary structure (Burkhardt, Rouskin, Yan et al. eLife 2017 and Mol Cell 2018). A less appreciated control point is operon mRNA maturation, which is initiated by endonuclease cleavage that leads to differential RNA stability among neighboring genes (DeLoughery et al. PNAS 2018). Compared to the wealth of information for transcriptional and translational control, our understanding of the code for mRNA maturation lags far behind. Using the endonuclease RNase Y in B. subtilis as a model, we leverage high-throughput technologies to investigate what instructs the initial cleavage that determines the stoichiometry of its products.