Cellular processes are controlled by gene regulatory circuits that are comprised of interactions among genes and proteins. One such process common to organisms ranging from bacteria to mammalian stem cells is pluripotent differentiation, where cells can differentiate into one of several possible fates. We propose to investigate how interactions at the molecular level within and across genetic circuits determine pluripotent differentiation at the cellular level. We will study pluripotent differentiation in Bacillus subtilis, a simple, well characterized and experimentally accessible system as a model for genetic control of cell fate choice. Environmental stress induces B. subtilis cells to undergo sporulation or differentiate into the competence state and take up extracellular DNA and incorporate it into their chromosome. We have recently identified the core competence circuit and showed that it exhibits excitable dynamics triggered by noise. Many of the molecular components that regulate competence and sporulation in B. subtilis are known. How interactions within and across competence and sporulation circuits regulate the choice and execution of appropriate differentiation programs is, however, poorly understood. We will study this problem using quantitative fluorescence time- lapse microscopy at the single-cell level to establish how the dynamics of molecular interactions regulate this process. Exploiting genetic manipulation techniques available for B. subtilis, we will measure how systematic re-engineering of circuit interactions control differentiation. Utilizing established connectivity maps of competence and sporulation circuits, we will also construct mathematical frameworks to generate predictions and analyze results. Specifically, we will apply these methods to: (1) Determine the functional importance of competence circuit architecture by comparing it to engineered alternative topologies in silico and in vivo. (2) Determine the functional importance of cross-regulation in cell fate choice. (3) Determine how the transient activity of the competence circuit alters the progression and execution of sporulation. This integrative research is necessary to determine how molecular interactions within and across genetic circuits control pluripotent differentiation at the cellular level. Identification of the mechanics that dictate the choice and execution of cell fate in this model organism are likely to be relevant to pluripotent differentiation in diverse organisms including mammalian systems.

Public Health Relevance

This proposal will establish a comprehensive description of how bacteria undergo differentiation. The resulting data will be relevant to: 1) Controlling differentiation in bacteria to prevent bacterial spore formation in foods. 2) Preventing the ability of bacteria to naturally become resistant to antibiotics. 3) Developing new techniques to control how cells differentiate, which can be applied in the future to control differentiation of mammalian stem cell to substitute for any missing or diseased cell types.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Modeling and Analysis of Biological Systems Study Section (MABS)
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Brazhnik, Paul
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University of Texas Sw Medical Center Dallas
Schools of Medicine
United States
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Mugler, Andrew; Kittisopikul, Mark; Hayden, Luke et al. (2016) Noise Expands the Response Range of the Bacillus subtilis Competence Circuit. PLoS Comput Biol 12:e1004793
Narula, Jatin; Kuchina, Anna; Zhang, Fang et al. (2016) Slowdown of growth controls cellular differentiation. Mol Syst Biol 12:871
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Espinar, Lorena; Dies, Marta; Cagatay, Tolga et al. (2013) Circuit-level input integration in bacterial gene regulation. Proc Natl Acad Sci U S A 110:7091-6
Asally, Munehiro; Kittisopikul, Mark; Rue, Pau et al. (2012) Localized cell death focuses mechanical forces during 3D patterning in a biofilm. Proc Natl Acad Sci U S A 109:18891-6
Orchard, Robert C; Kittisopikul, Mark; Altschuler, Steven J et al. (2012) Identification of F-actin as the dynamic hub in a microbial-induced GTPase polarity circuit. Cell 148:803-15
Kuchina, Anna; Espinar, Lorena; Cagatay, Tolga et al. (2011) Temporal competition between differentiation programs determines cell fate choice. Mol Syst Biol 7:557

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