206 Biology East
Department of Biology
Johns Hopkins University
3400 N. Charles Street
Baltimore, MD 21218-2685
Office: 410 516-4954
Departmental fax 410 516-5213
Postdoctoral ResearchNew York University
Why is stochastic cell fate specification important?Genes are expressed in either uniform, regionalized, or stochastic patterns in developing tissues. Though we know a great deal about the regulatory mechanisms determining uniform and regionalized expression, the mechanisms controlling the fundamental process of stochastic gene expression have remained mysterious. In contrast with other mechanisms that override underlying molecular noise to dictate reproducible outcomes, stochastic gene regulatory mechanisms exploit variation to induce On or Off expression states randomly in individual cells. Stochastic gene expression mechanisms are critical for stem cell differentiation, B cell specification, visual and olfactory receptor selection, and adhesion molecule diversification. Thus, studying stochastic gene expression phenomena will impact our understanding of human medical conditions such as immunodeficiencies, lymphoma, anosmia, and vision disorders.
Why study the fly eye?We are using the fly eye as a paradigm to elucidate the mechanisms controlling stochastic gene expression during development. Similar to the human color vision system, the photoreceptors of the fly eye randomly express several light-detecting Rhodopsin proteins. The fly eye is an ideal system to study this phenomenon because it provides a simple binary output for stochastic gene expression, the general mechanisms of cell-fate specification are well-understood, and a vast array of genetic and transgenic tools are available to manipulate cis-regulatory inputs and upstream trans-acting factors.
What do we know?The transcription factor Spineless is the critical regulator controlling the random mosaic pattern of photoreceptor subtypes in the fly eye. Each allele of spineless makes its own random expression choice independent of the other. Stochastic on/off expression of spineless is determined by global activation coupled with random repression requiring combinatorial inputs from cis-regulatory elements acting at long range. Through interchromosomal communication (InterCom), the two alleles coordinate their expression state. The next goal is to determine the molecular mechanisms controlling intrinsically stochastic expression decisions and InterCom.
What's the plan?Since stochastic Spineless expression requires inputs from an enhancer and two silencer elements, we propose three main mechanistic models controlling the intrinsically random expression decision:
- the spineless locus randomly assumes one of two DNA looping configurations (i.e. active and repressed) similar to LCR looping in vertebrates
- one silencer facilitates the nucleation of closed chromatin state spreading from the other silencer
- one silencer generally lowers expression in all R7s whereas the other specifically provides the stochastic input
- a temporally distinct two step mechanism involving both alleles making the expression decision followed by an activating and repressing tug of war
- a temporally distinct two step mechanism in which one allele makes the decision and then imposes the decision onto the other naïve allele
- a mechanism involving contemporaneous decisions that average the activating and repressing inputs from each allele
For additional information, please check our lab website.
25. Johnston, R.J., Jr., and Desplan, C. Interchromosomal communication coordinates an intrinsically stochastic expression decision between alleles. (submitted).
24. Hsiao, H.Y., Jukam, D., Johnston, R.J., Jr., and Desplan, C. The neuronal transcription factor Erect wing regulates specification and maintenance of Drosophila R8 photoreceptor subtypes. Dev Biol (in revision).
23. Johnston, R.J., Jr. Lessons about terminal differentiation from the specification of color-detecting photoreceptors in the Drosophila retina. Ann N Y Acad Sci (in press).
22. Thanawala, S., Rister, J., Goldberg, G.W., Zuskov, A., Olesnicky, E.C., Flowers, J.M., Jukam, D., Purugganan, M.D., Gavis, E.R., Desplan, C., and Johnston, R.J., Jr. (2013) Regional modulation of a stochastically expressed factor determines ommatidial subtypes in the Drosophila retina. Dev Cell, 25, 93-105.
21. Hsiao H.Y., Johnston, R.J., Jr., Jukam, D., Vasiliauskas, D., Desplan, C., and Rister, J. (2012) Dissection and Immunohistochemistry of Larval, Pupal and Adult Drosophila Retinas. JoVE, 69, e4347.
20. Sood, P., Johnston, R.J., Jr., and Kussell, E. (2011) Regulatory motif behavior depends on different network contexts in the fly Rhodopsin patterning system. PLoS Comput Biol, 8, e1002357.
19. Vasiliauskas, D., Mazzoni, E.O, Sprecher, S.G., Brodetskiy, K., Johnston R.J., Jr., Lidder, P., Vogt, N., Celik, A., and Desplan, C. (2011) Feedback from Rhodopsin protein controls rhodopsin exclusion in Drosophila R8 photoreceptors. Nature, 479, 108-112.
18. Johnston, R.J., Jr., Otake, Y., Sood, P., Vogt, N., Behnia, R., Vasiliauskas, D., McDonald, E., Xie, B., Koenig, S., Wolf, R., Cook, T., Gebelein, B., Kussell, E., Nakagoshi, H., and Desplan, C. (2011) Interlocked feedforward loops control cell-type-specific rhodopsin expression in the Drosophila eye. Cell, 145, 956-968.
17. Johnston, R.J., Jr., and Desplan, C. (2010) Stochastic mechanisms of cell fate specification that yield random or robust outcomes. Annu Rev Cell Dev Biol, 26, 689-719.
16. Johnston, R.J., Jr., and Desplan, C. (2010) A penetrating look at stochasticity in development. Cell, 140, 610-612.
15. Vasiliauskas, D., Johnston, R., and Desplan, C. (2009) Maintaining a stochastic neuronal cell fate decision. Genes Dev, 23, 385-390.
14. Johnston, R.J., Jr., and Desplan, C. (2008) Stochastic neuronal cell fate choices. Curr Opin Neurobiol, 18, 20-27.
13. Mazzoni, E.O., Celik, A., Wernet, M.F., Vasiliauskas, D., Johnston, R.J., Cook, T.A., Pichaud, F., and Desplan, C. (2008) Iroquois complex genes induce co-expression rhodopsins in Drosophila. PLoS Biol, 6, e97 825-835.
12. Sarin, S., O’Meara, M.M., Flowers, E.B., Antonio, C., Poole, R.J., Didiano, D., Johnston R.J., Jr., Chang, S, Narula, S., and Hobert, O. (2007) Genetic screens for Caenorhabditis elegans mutants defective in left/right asymmetric neuronal fate specification. Genetics, 176, 2109-2130.
11. Johnston, R.J., Jr., Copeland, J.W., Fasnacht, M., Etchberger, J.F., Liu, J., Honig, B., and Hobert, O. (2006) An unusual Zn-finger/FH2 domain protein controls a left/right asymmetric neuronal fate decision in C. elegans. Development, 133, 3317-3328.
10. Johnston, R.J., Jr., and Hobert, O. (2005) A novel C. elegans zinc finger transcription factor, lsy-2, required for the cell type-specific expression of the lsy-6 microRNA. Development, 132, 5451-5460.
9. Johnston, R.J., Jr., Chang, S., Etchberger, J.F., Ortiz, C.O., and Hobert, O. (2005) MicroRNAs acting in a double-negative feedback loop to control a neuronal cell fate decision. PNAS, 102, 12449-12454.
8. Chang, S., Johnston, R.J., Jr., Frokjaer-Jensen, C., Lockery, S., and Hobert, O. (2004) MicroRNAs act sequentially and asymmetrically to control chemosensory laterality in the nematode. Nature, 430, 785-789.
7. Johnston, R.J., and Hobert, O. (2003) A microRNA controlling left/right neuronal asymmetry in Caenorhabditis elegans. Nature, 426, 845-849.
6. Chang, S., Johnston, R.J., Jr., and Hobert, O (2003) A transcriptional regulatory cascade that controls left/right asymmetry in chemosensory neurons of C. elegans. Genes Dev, 17, 2123-2137.
5. Hobert, O., Johnston, R.J., Jr., and Chang, S. (2002) Left-right asymmetry in the nervous system: the Caenorhabditis elegans model. Nat Rev Neurosci, 3, 629-640.
4. Melia, T.J., Weber, T., Mcnew, J.A., Fisher, L.E., Johnston, R.J., Parlati, F., Mahal, L.K., Sollner, T.H., and Rothman, J.E. (2002) Regulation of membrane fusion by the membrane-proximal coil of the t-SNARE during zippering of SNAREpins. J Cell Biol, 158, 929-940.
3. McNew, J.A., Parlati, F., Fukuda, R., Johnston, R.J., Paz, K., Paumet, F., Sollner, T.H., and Rothman, J.E. (2000) Compartmental specificity of cellular membrane fusion encoded in SNARE proteins. Nature, 407, 153-159.
2. McNew, J.A., Weber, T., Parlati, F., Johnston, R.J., Melia, T.J., Sollner, T.H., and Rothman, J.E. (2000) Close is not enough: SNARE-dependent membrane fusion requires an active mechanism that transduces force to membrane anchors. J Cell Biol, 150, 105-117.
1. Weber, T., Parlati, F., McNew, J. A., Johnston, R.J., Westermann, B., Sollner, T.H., and Rothman, J.E. (2000) SNAREpins are functionally resistant to disruption by NSF and alphaSNAP. J Cell Biol, 149, 1063-1072.