Optogenetics pp 389-403 | Cite as

Guidelines for Photoreceptor Engineering

  • Thea Ziegler
  • Charlotte Helene Schumacher
  • Andreas MöglichEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1408)


Sensory photoreceptors underpin optogenetics by mediating the noninvasive and reversible perturbation of living cells by light with unprecedented temporal and spatial resolution. Spurred by seminal optogenetic applications of natural photoreceptors, the engineering of photoreceptors has recently garnered wide interest and has led to the construction of a broad palette of novel light-regulated actuators. Photoreceptors are modularly built of photosensors that receive light signals, and of effectors that carry out specific cellular functions. These modules have to be precisely connected to allow efficient communication, such that light stimuli are relayed from photosensor to effector. The engineering of photoreceptors benefits from a thorough understanding of the underlying signaling mechanisms. This chapter gives a brief overview of key characteristics and signal-transduction mechanisms of sensory photoreceptors. Adaptation of these concepts in photoreceptor engineering has enabled the generation of novel optogenetic tools that greatly transcend the repertoire of natural photoreceptors.

Key words

Light Optogenetics Protein engineering Sensory photoreceptor Signal transduction 



Research in our laboratory is generously supported through a Sofja-Kovalevskaya Award by the Alexander-von-Humboldt Foundation (to A.M.) and by the Deutsche Forschungsgemeinschaft within the Cluster of Excellence ‘Unicat—Unifying Concepts in Catalysis’.


  1. 1.
    Möglich A, Moffat K (2010) Engineered photoreceptors as novel optogenetic tools. Photochem Photobiol Sci 9:1286–1300CrossRefPubMedGoogle Scholar
  2. 2.
    Schmidt D, Cho YK (2015) Natural photoreceptors and their application to synthetic biology. Trends Biotechnol 33:80–91CrossRefPubMedGoogle Scholar
  3. 3.
    Ziegler T, Möglich A (2015) Photoreceptor engineering. Front Mol Biosci 2:30CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Ernst OP, Lodowski DT, Elstner M, Hegemann P, Brown LS, Kandori H (2014) Microbial and animal rhodopsins: structures, functions, and molecular mechanisms. Chem Rev 114:126–163CrossRefPubMedGoogle Scholar
  5. 5.
    Fenno L, Yizhar O, Deisseroth K (2011) The development and application of optogenetics. Annu Rev Neurosci 34:389–412CrossRefPubMedGoogle Scholar
  6. 6.
    Schneider F, Grimm C, Hegemann P (2015) Biophysics of channelrhodopsin. Annu Rev Biophys 44:167–186CrossRefPubMedGoogle Scholar
  7. 7.
    Hegemann P (2008) Algal sensory photoreceptors. Annu Rev Plant Biol 59:167–189CrossRefPubMedGoogle Scholar
  8. 8.
    Möglich A, Yang X, Ayers RA, Moffat K (2010) Structure and function of plant photoreceptors. Annu Rev Plant Biol 61:21–47CrossRefPubMedGoogle Scholar
  9. 9.
    Rockwell NC, Su Y-S, Lagarias JC (2006) Phytochrome structure and signaling mechanisms. Annu Rev Plant Biol 57:837–858CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Brown BA, Cloix C, Jiang GH, Kaiserli E, Herzyk P, Kliebenstein DJ, Jenkins GI (2005) A UV-B-specific signaling component orchestrates plant UV protection. Proc Natl Acad Sci U S A 102:18225–18230CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Conrad KS, Manahan CC, Crane BR (2014) Photochemistry of flavoprotein light sensors. Nat Chem Biol 10:801–809CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Losi A, Mandalari C, Gärtner W (2015) The evolution and functional role of flavin-based prokaryotic photoreceptors. Photochem Photobiol 91:1021–1031. doi: 10.1111/php.12489 Google Scholar
  13. 13.
    Pudasaini A, El-Arab KK, Zoltowski BD (2015) LOV-based optogenetic devices: light-driven modules to impart photoregulated control of cellular signaling. Front Mol Biosci 2:18CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Rockwell NC, Duanmu D, Martin SS, Bachy C, Price DC, Bhattacharya D, Worden AZ, Lagarias JC (2014) Eukaryotic algal phytochromes span the visible spectrum. Proc Natl Acad Sci U S A 111:3871–3876CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Ikeuchi M, Ishizuka T (2008) Cyanobacteriochromes: a new superfamily of tetrapyrrole-binding photoreceptors in cyanobacteria. Photochem Photobiol Sci 7:1159–1167CrossRefPubMedGoogle Scholar
  16. 16.
    Rockwell NC, Lagarias JC (2010) A brief history of phytochromes. Chemphyschem 11:1172–1180CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Filonov GS, Piatkevich KD, Ting L-M, Zhang J, Kim K, Verkhusha VV (2011) Bright and stable near-infrared fluorescent protein for in vivo imaging. Nat Biotechnol 29:757–761CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Gasser C, Taiber S, Yeh C-M, Wittig CH, Hegemann P, Ryu S, Wunder F, Möglich A (2014) Engineering of a red-light-activated human cAMP/cGMP-specific phosphodiesterase. Proc Natl Acad Sci U S A 111:8803–8808CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Ryu M-H, Kang I-H, Nelson MD, Jensen TM, Lyuksyutova AI, Siltberg-Liberles J, Raizen DM, Gomelsky M (2014) Engineering adenylate cyclases regulated by near-infrared window light. Proc Natl Acad Sci U S A 111:10167–10172CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Shu X, Royant A, Lin MZ, Aguilera TA, Lev-Ram V, Steinbach PA, Tsien RY (2009) Mammalian expression of infrared fluorescent proteins engineered from a bacterial phytochrome. Science 324:804–807CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Alexandre MT, Arents JC, van Grondelle R, Hellingwerf KJ, Kennis JT (2007) A base-catalyzed mechanism for dark state recovery in the Avena sativa phototropin-1 LOV2 domain. Biochemistry 46:3129–3137CrossRefPubMedGoogle Scholar
  22. 22.
    Zhou XX, Chung HK, Lam AJ, Lin MZ (2012) Optical control of protein activity by fluorescent protein domains. Science 338:810–814CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Lee J, Natarajan M, Nashine VC, Socolich M, Vo T, Russ WP, Benkovic SJ, Ranganathan R (2008) Surface sites for engineering allosteric control in proteins. Science 322:438–442CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Ohlendorf R, Vidavski RR, Eldar A, Moffat K, Möglich A (2012) From dusk till dawn: one-plasmid systems for light-regulated gene expression. J Mol Biol 416:534–542CrossRefPubMedGoogle Scholar
  25. 25.
    Jansen V, Alvarez L, Balbach M, Strünker T, Hegemann P, Kaupp UB, Wachten D (2015) Controlling fertilization and cAMP signaling in sperm by optogenetics. eLife 4:e05161Google Scholar
  26. 26.
    Grusch M, Schelch K, Riedler R, Reichhart E, Differ C, Berger W, Inglés-Prieto Á, Janovjak H (2014) Spatio-temporally precise activation of engineered receptor tyrosine kinases by light. EMBO J 33:1713–1726CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Guntas G, Hallett RA, Zimmerman SP, Williams T, Yumerefendi H, Bear JE, Kuhlman B (2015) Engineering an improved light-induced dimer (iLID) for controlling the localization and activity of signaling proteins. Proc Natl Acad Sci U S A 112:112–117CrossRefPubMedGoogle Scholar
  28. 28.
    Cosentino C, Alberio L, Gazzarrini S et al (2015) Optogenetics. Engineering of a light-gated potassium channel. Science 348:707–710CrossRefPubMedGoogle Scholar
  29. 29.
    Goldsmith M, Tawfik DS (2012) Directed enzyme evolution: beyond the low-hanging fruit. Curr Opin Struct Biol 22:406–412CrossRefPubMedGoogle Scholar
  30. 30.
    Bugaj LJ, Choksi AT, Mesuda CK, Kane RS, Schaffer DV (2013) Optogenetic protein clustering and signaling activation in mammalian cells. Nat Methods 10:249–252CrossRefPubMedGoogle Scholar
  31. 31.
    Lamb JS, Zoltowski BD, Pabit SA, Crane BR, Pollack L (2008) Time-resolved dimerization of a PAS-LOV protein measured with photocoupled small angle X-ray scattering. J Am Chem Soc 130:12226–12227CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Takahashi F, Yamagata D, Ishikawa M, Fukamatsu Y, Ogura Y, Kasahara M, Kiyosue T, Kikuyama M, Wada M, Kataoka H (2007) AUREOCHROME, a photoreceptor required for photomorphogenesis in stramenopiles. Proc Natl Acad Sci U S A 104:19625–19630CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Nash AI, McNulty R, Shillito ME, Swartz TE, Bogomolni RA, Luecke H, Gardner KH (2011) Structural basis of photosensitivity in a bacterial light-oxygen-voltage/helix-turn-helix (LOV-HTH) DNA-binding protein. Proc Natl Acad Sci U S A 108:9449–9454CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Christie JM, Arvai AS, Baxter KJ et al (2012) Plant UVR8 photoreceptor senses UV-B by tryptophan-mediated disruption of cross-dimer salt bridges. Science 335:1492–1496CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Kennedy MJ, Hughes RM, Peteya LA, Schwartz JW, Ehlers MD, Tucker CL (2010) Rapid blue-light-mediated induction of protein interactions in living cells. Nat Methods 7:973–975CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Yazawa M, Sadaghiani AM, Hsueh B, Dolmetsch RE (2009) Induction of protein-protein interactions in live cells using light. Nat Biotechnol 27:941–945CrossRefPubMedGoogle Scholar
  37. 37.
    Levskaya A, Weiner OD, Lim WA, Voigt CA (2009) Spatiotemporal control of cell signalling using a light-switchable protein interaction. Nature 461:997–1001CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Shimizu-Sato S, Huq E, Tepperman JM, Quail PH (2002) A light-switchable gene promoter system. Nat Biotechnol 20:1041–1044CrossRefPubMedGoogle Scholar
  39. 39.
    Harper SM, Neil LC, Gardner KH (2003) Structural basis of a phototropin light switch. Science 301:1541–1544CrossRefPubMedGoogle Scholar
  40. 40.
    Christie JM, Blackwood L, Petersen J, Sullivan S (2015) Plant flavoprotein photoreceptors. Plant Cell Physiol 56:401–413CrossRefPubMedGoogle Scholar
  41. 41.
    Renicke C, Schuster D, Usherenko S, Essen L-O, Taxis C (2013) A LOV2 domain-based optogenetic tool to control protein degradation and cellular function. Chem Biol 20:619–626CrossRefPubMedGoogle Scholar
  42. 42.
    Herman E, Kottke T (2015) Allosterically regulated unfolding of the A′α helix exposes the dimerization site of the blue-light-sensing aureochrome-LOV domain. Biochemistry 54:1484–1492CrossRefPubMedGoogle Scholar
  43. 43.
    Conrad KS, Bilwes AM, Crane BR (2013) Light-induced subunit dissociation by a light-oxygen-voltage domain photoreceptor from Rhodobacter sphaeroides. Biochemistry 52:378–391CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Rubinstenn G, Vuister GW, Mulder FAA, Düx PE, Boelens R, Hellingwerf KJ, Kaptein R (1998) Structural and dynamic changes of photoactive yellow protein during its photocycle in solution. Nat Struct Mol Biol 5:568–570CrossRefGoogle Scholar
  45. 45.
    Rivera-Cancel G, Ko W, Tomchick DR, Correa F, Gardner KH (2014) Full-length structure of a monomeric histidine kinase reveals basis for sensory regulation. Proc Natl Acad Sci U S A 111:17839–17844CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Takala H, Björling A, Berntsson O et al (2014) Signal amplification and transduction in phytochrome photosensors. Nature 509:245–248CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Anders K, Gutt A, Gärtner W, Essen L-O (2014) Phototransformation of the red light sensor cyanobacterial phytochrome 2 from Synechocystis species depends on its tongue motifs. J Biol Chem 289:25590–25600CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Chang K-Y, Woo D, Jung H et al (2014) Light-inducible receptor tyrosine kinases that regulate neurotrophin signalling. Nat Commun 5:4057PubMedGoogle Scholar
  49. 49.
    Kim N, Kim JM, Lee M, Kim CY, Chang K-Y, Heo WD (2014) Spatiotemporal control of fibroblast growth factor receptor signals by blue light. Chem Biol 21:903–912CrossRefPubMedGoogle Scholar
  50. 50.
    Nihongaki Y, Suzuki H, Kawano F, Sato M (2014) Genetically engineered photoinducible homodimerization system with improved dimer-forming efficiency. ACS Chem Biol 9:617–621CrossRefPubMedGoogle Scholar
  51. 51.
    Wang X, Chen X, Yang Y (2012) Spatiotemporal control of gene expression by a light-switchable transgene system. Nat Methods 9:266–269CrossRefPubMedGoogle Scholar
  52. 52.
    Aoki K, Kumagai Y, Sakurai A, Komatsu N, Fujita Y, Shionyu C, Matsuda M (2013) Stochastic ERK activation induced by noise and cell-to-cell propagation regulates cell density-dependent proliferation. Mol Cell 52:529–540CrossRefPubMedGoogle Scholar
  53. 53.
    Wu YI, Frey D, Lungu OI, Jaehrig A, Schlichting I, Kuhlman B, Hahn KM (2009) A genetically encoded photoactivatable Rac controls the motility of living cells. Nature 461:104–108CrossRefPubMedPubMedCentralGoogle Scholar
  54. 54.
    Schmidt D, Tillberg PW, Chen F, Boyden ES (2014) A fully genetically encoded protein architecture for optical control of peptide ligand concentration. Nat Commun 5:3019PubMedPubMedCentralGoogle Scholar
  55. 55.
    Bonger KM, Rakhit R, Payumo AY, Chen JK, Wandless TJ (2014) General method for regulating protein stability with light. ACS Chem Biol 9:111–115CrossRefPubMedGoogle Scholar
  56. 56.
    Anantharaman V, Balaji S, Aravind L (2006) The signaling helix: a common functional theme in diverse signaling proteins. Biol Direct 1:25CrossRefPubMedPubMedCentralGoogle Scholar
  57. 57.
    Möglich A, Ayers RA, Moffat K (2009) Design and signaling mechanism of light-regulated histidine kinases. J Mol Biol 385:1433–1444CrossRefPubMedGoogle Scholar
  58. 58.
    Möglich A, Ayers RA, Moffat K (2010) Addition at the molecular level: signal integration in designed Per-ARNT-Sim receptor proteins. J Mol Biol 400:477–486CrossRefPubMedGoogle Scholar
  59. 59.
    Rockwell NC, Ohlendorf R, Möglich A (2013) Cyanobacteriochromes in full color and three dimensions. Proc Natl Acad Sci U S A 110:806–807CrossRefPubMedPubMedCentralGoogle Scholar
  60. 60.
    Levskaya A, Chevalier AA, Tabor JJ et al (2005) Synthetic biology: sngineering Escherichia coli to see light. Nature 438:441–442CrossRefPubMedGoogle Scholar
  61. 61.
    Losi A, Polverini E, Quest B, Gärtner W (2002) First evidence for phototropin-related blue-light receptors in prokaryotes. Biophys J 82:2627–2634CrossRefPubMedPubMedCentralGoogle Scholar
  62. 62.
    Avelar GM, Schumacher RI, Zaini PA, Leonard G, Richards TA, Gomes SL (2014) A rhodopsin-guanylyl cyclase gene fusion functions in visual perception in a fungus. Curr Biol 24:1234–1240CrossRefPubMedPubMedCentralGoogle Scholar
  63. 63.
    Yoshihara S, Suzuki F, Fujita H, Geng XX, Ikeuchi M (2000) Novel putative photoreceptor and regulatory genes required for the positive phototactic movement of the unicellular motile Cyanobacterium Synechocystis sp. PCC 6803. Plant Cell Physiol 41:1299–1304CrossRefPubMedGoogle Scholar
  64. 64.
    Davis SJ, Vener AV, Vierstra RD (1999) Bacteriophytochromes: phytochrome-like photoreceptors from nonphotosynthetic eubacteria. Science 286:2517–2520CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Thea Ziegler
    • 1
    • 2
  • Charlotte Helene Schumacher
    • 1
  • Andreas Möglich
    • 1
    • 3
    Email author
  1. 1.Institut für Biologie, Biophysikalische ChemieHumboldt-Universität zu BerlinBerlinGermany
  2. 2.Lehrstuhl für BiochemieUniversität BayreuthBayreuthGermany
  3. 3.Faculty of Biology, Chemistry and Earth Sciences, Lehrstuhl für BiochemieUniversität BayreuthBayreuthGermany

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