N. Gautam, Ph.D.
Molecular Cell Biology Program
5548 Clinical Sciences Research Building
Subcellular optogenetics, light, imaging, signaling, GPCRs, migration, differentiation, contractility, secretion
Optogenetic Control of Signaling and Cell Behavior
One of our goals is to develop tools for subcellular optogenetics using light sensitive proteins. We define subcellular optogenetics as the ability to optically regulate molecular activity in selected regions within a cell. Coordinated variations in subcellular signaling play a critical role in governing cell behavior. For instance, asymmetric signaling activity across a cell governs processes such as cell migration and neuron differentiation. Signaling is also confined often to subcellular sites such as the plasma membrane, Golgi or endosomes. Localized optical control can help decipher the role such signaling plays in regulating cell function. We have developed tools based on visual color opsins and a plant cryptochrome that provide spatial control of signaling within a single cell. They evoke specific cellular responses such as migration and neurite growth with a light signal. Over the years it has been possible to visualize subcellular variation in signaling activity by generating a set of fluorescent protein tools. Similarly, developing a library of optogenetic tools to control this subcellular signaling activity can provide transformative experimental control over cell behaviors.
Optical control over multiple proteins in a cell
We are also interested in developing methods to exert optical control over more than one signaling protein in the same cell. For example, spectrally distinct opsins can be used to activate GPCRs coupled to different G proteins and increase or decrease cardiomyocyte contractility or insulin secretion with different wavelengths of light. Spectrally selective opsins can also be used with light sensitive domains such as CRY2 or LOV to activate an entire pathway and simultaneously target an individual protein in a cell. We plan to engineer such optical triggers incorporating properties such as a wider range of spectral tuning, spectral selectivity and the ability to sustain prolonged activity. The vast family of opsins in metazoans with diverse spectral tuning and second messenger activity provide a natural resource to develop these tools.
The optical triggers will be used to control critical cell behaviors in culture and in whole animals using light.
Karunarathne WK, O`Neill PR & Gautam N (2015) Subcellular optogenetics - controlling signaling and single-cell behavior. J Cell Sci 128:15-25. PMC4282045
O’Neill, P.R., & Gautam, N. (2014) Subcellular optogenetic inhibition of G proteins generates signaling gradients and cell migration. Mol Biol Cell 25:2305-14. PMC 24920824
Giri, L., Patel, A.K., Karunarathne, W.K.A., Kalyanaraman, V., Venkatesh, K.V., & Gautam, N. (2014) A G Protein Subunit Translocation Embedded Network Motif Underlies GPCR Regulation of Calcium Oscillations. Biophys J 107:242-54. PMC4119271
O`Neill, P.R., Giri, L., Karunarathne, W.K., Patel, A.K., Venkatesh, K.V., & Gautam, N. (2014) The structure of dynamic GPCR signaling networks. Wiley Interdiscip Rev Syst Biol Med. 6:115-23. PMC4007319
Karunarathne, W.K., Giri, L., Patel, A.K., Venkatesh, K.V., & Gautam, N. (2013) Optical control demonstrates switch-like PIP3 dynamics underlying the initiation of immune cell migration. Proc Natl Acad Sci USA. 110(17):E1575-83. PMC3637758
Karunarathne, W.K., Giri, L., Kalyanaraman, V., & Gautam, N. (2013) Optically triggering spatiotemporally confined GPCR activity in a cell and programming neurite initiation and extension. Proc Natl Acad Sci USA 110(17):E1565-74. PMC3637763
O`Neill, P.R., Karunarathne, W.K., Kalyanaraman, V., Silvius, J.R., & Gautam, N. (2012) G-protein signaling leverages subunit-dependent membrane affinity to differentially control βγ translocation to intracellular membranes. Proc Natl Acad Sci USA 109(51):E3568-77. PMC3529095
Saini, D.K., Karunarathne, W.K., Angaswamy, N., Saini, D., Cho, J.H., Kalyanaraman, V., & Gautam, N. (2010) Regulation of Golgi structure and secretion by receptor-induced G protein betagamma complex translocation. Proc Natl Acad Sci USA 107: 11417-11422. PMC2895111
Saini, D.K., Chisari, M., & Gautam, N. Shuttling and translocation of heterotrimeric G proteins and Ras. (2009) Trends Pharmacol Sci 30: 278-286. PMC19427041
Last Updated: 4/11/2017 4:18:02 PM