, Volume 241, Issue 2, pp 387–402 | Cite as

Distinct subfunctionalization and neofunctionalization of the B-class MADS-box genes in Physalis floridana

  • Shaohua Zhang
  • Ji-Si Zhang
  • Jing Zhao
  • Chaoying HeEmail author
Original Article


Main conclusion

This work suggested that in Physalis PFGLO1–PFDEF primarily determined corolla and androecium identity, and acquired a novel role in gynoecia functionality, while PFGLO2–PFTM6 functioned in pollen maturation only.


The B-class MADS-box genes play a crucial role in determining the organ identity of the corolla and androecium. Two GLOBOSA-like (GLO-like) PFGLO1 and PFGLO2 and two DEFICIENS-like (DEF-like) PFDEF and PFTM6 genes were present in Physalis floridana. However, the double-layered-lantern1 (doll1) mutant is the result of a single recessive mutation in PFGLO1, hinting a distinct divergent pattern of B-class genes. In this work, we utilized the tobacco rattle virus (TRV)-mediated gene silencing approach to further verify this assumption in P. floridana. Silencing of PFGLO1 or/and PFDEF demonstrated their primary role in determining corolla and androecium identity. However, specific PFGLO2 or/and PFTM6 silencing did not affect any organ identity but showed a reduction in mature pollen. These results suggested that both PFGLO2 and PFTM6 had lost their role in organ identity determination but functioned in pollen maturation. Evaluation of fruit setting in reciprocal crosses suggested that both PFGLO1 and PFDEF might have acquired an essential and novel role in the functionality of gynoecia. Such a divergence of the duplicated GLO–DEF heterodimer genes in floral development is different from the existing observations within Solanaceae. Therefore, our research sheds new light on the functional evolution of the duplicated B-class MADS-box genes in angiosperms.


MADS-box gene Functional evolution Fertility Organ identity Physalis 



Bimolecular fluorescence complementation


Complementary DNA


Green fluorescence protein


Open reading frame


Tobacco rattle virus


Virus-induced gene silencing


Yellow fluorescence protein



This work was supported by the grants (31070203 and 91331103) from the National Natural Science Foundation of China.

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

425_2014_2190_MOESM1_ESM.pdf (775 kb)
Supplementary material 1 (PDF 774 kb)


  1. Coen ES, Meyerowitz EM (1991) The war of the whorls: genetic interactions controlling flower development. Nature 353:31–37PubMedCrossRefGoogle Scholar
  2. Davies B, Egea-Cortines M, de Andrade Silva E, Saedler H, Sommer H (1996) Multiple interactions amongst floral homeotic MADS box proteins. EMBO J 15:4330–4343PubMedCentralPubMedGoogle Scholar
  3. Davies B, Motte P, Keck E, Saedler H, Sommer H, Schwarz-Sommer Z (1999) PLENA and FARINELLI: redundancy and regulatory interactions between two Antirrhinum MADS-box factors controlling flower development. EMBO J 18:4023–4034PubMedCentralPubMedCrossRefGoogle Scholar
  4. de Martino G, Pan I, Emmanuel E, Levy A, Irish VF (2006) Functional analyses of two tomato APETALA3 genes demonstrate diversification in their roles in regulating floral development. Plant Cell 18:1833–1845PubMedCentralPubMedCrossRefGoogle Scholar
  5. Egea-Cortines M, Saedler H, Sommer H (1999) Ternary complex formation between the MADS-box proteins SQUAMOSA, DEFICIENS and GLOBOSA is involved in the control of floral architecture in Antirrhinum majus. EMBO J 18:5370–5379PubMedCentralPubMedCrossRefGoogle Scholar
  6. Geuten K, Irish V (2010) Hidden variability of floral homeotic B genes in Solanaceae provides a molecular basis for the evolution of novel functions. Plant Cell 22:2562–2578PubMedCentralPubMedCrossRefGoogle Scholar
  7. Geuten K, Viaene T, Irish VF (2011) Robustness and evolvability in the B-system of flower development. Ann Bot 107:1545–1556PubMedCentralPubMedCrossRefGoogle Scholar
  8. Goto K, Meyerowitz EM (1994) Function and regulation of the Arabidopsis floral homeotic gene PISTILLATA. Genes Dev 8:1548–1560PubMedCrossRefGoogle Scholar
  9. He CY, Saedler H (2005) Heterotopic expression of MPF2 is the key to the evolution of the Chinese lantern of Physalis, a morphological novelty in Solanaceae. Proc Natl Acad Sci USA 102:5779–5784PubMedCentralPubMedCrossRefGoogle Scholar
  10. He CY, Saedler H (2007) Hormonal control of the inflated calyx syndrome, a morphological novelty, in Physalis. Plant J 49:935–946PubMedCrossRefGoogle Scholar
  11. He CY, Münster T, Saedler H (2004) On the origin of morphological floral novelties. FEBS Lett 567:147–151PubMedCrossRefGoogle Scholar
  12. He CY, Sommer H, Grosardt B, Huijser P, Saedler H (2007) PFMAGO, a MAGO NASHI-like factor, interacts with the MADS-box protein MPF2 from Physalis floridana. Mol Biol Evol 24:1229–1241PubMedCrossRefGoogle Scholar
  13. Hernandez-Hernandez T, Martinez-Castilla LP, Alvarez-Buylla ER (2007) Functional diversification of B MADS-box homeotic regulators of flower development: adaptive evolution in protein–protein interaction domains after major gene duplication events. Mol Biol Evol 24:465–481PubMedCrossRefGoogle Scholar
  14. Hofer KA, Ruonala R, Albert VA (2012) The double-corolla phenotype in the Hawaiian lobelioid genus Clermontia involves ectopic expression of PISTILLATA B-function MADS box gene homologs. EvoDevo 3:26PubMedCentralPubMedCrossRefGoogle Scholar
  15. Immink RGH, Kaufmann K, Angenent GC (2010) The ‘ABC’ of MADS domain protein behavior and interactions. Semin Cell Dev Biol 21:87–93PubMedCrossRefGoogle Scholar
  16. Innan H, Kondrashov F (2010) The evolution of gene duplications: classifying and distinguishing between models. Nat Rev Genet 11:97–108PubMedCrossRefGoogle Scholar
  17. Irish VF, Litt A (2005) Flower development and evolution: gene duplication, diversification and redeployment. Curr Opin Genet Dev 15:454–460PubMedCrossRefGoogle Scholar
  18. Jack T, Brockman LL, Meyerowitz EM (1992) The homeotic gene APETALA3 of Arabidopsis thaliana encodes a MADS box and is expressed in petals and stamens. Cell 68:683–697PubMedCrossRefGoogle Scholar
  19. Kanno A, Saeki H, Kameya T, Saedler H, Theißen G (2003) Heterotopic expression of class B floral homeotic genes supports a modified ABC model for tulip (Tulipa gesneriana). Plant Mol Biol 52:831–841PubMedCrossRefGoogle Scholar
  20. Kramer EM, Holappa L, Gould B, Jaramillo MA, Setnikov D, Santiago P (2007) Elaboration of B gene function to include the identity of novel floral organs in the lower eudicot Aquilegia (Ranunculaceae). Plant Cell 19:750–766PubMedCentralPubMedCrossRefGoogle Scholar
  21. Lamb RS, Irish VF (2003) Functional divergence within the APETALA3/PISTILLATA floral homeotic gene lineages. Proc Natl Acad Sci USA 100:6558–6563PubMedCentralPubMedCrossRefGoogle Scholar
  22. Lange M, Orashakova S, Lange S, Melzer R, Theißen G, Smyth DR, Becker A (2013) The seirena B class floral homeotic mutant of California poppy (Eschscholzia californica) reveals a function of the enigmatic PI motif in the formation of specific multimeric MADS domain protein complexes. Plant Cell 25:438–453PubMedCentralPubMedCrossRefGoogle Scholar
  23. Lee HL, Irish VF (2011) Gene duplication and loss in a MADS box gene transcription factor circuit. Mol Biol Evol 28:3367–3380PubMedCrossRefGoogle Scholar
  24. Leseberg CH, Eissler CL, Wang X, Johns MA, Duvall MR, Mao L (2008) Interaction study of MADS-domain proteins in tomato. J Exp Bot 59:2253–2265PubMedCrossRefGoogle Scholar
  25. Liu Y, Nakayama N, Schiff M, Litt A, Irish VF, Dinesh-Kumar SP (2004) Virus induced gene silencing of a DEFICIENS ortholog in Nicotiana benthamiana. Plant Mol Biol 54:701–711PubMedCrossRefGoogle Scholar
  26. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCt method. Methods 25:402–408PubMedCrossRefGoogle Scholar
  27. McGonigle B, Bouhidel K, Irish VF (1996) Nuclear localization of the Arabidopsis APETALA3 and PISTILLATA homeotic gene products depends on their simultaneous expression. Genes Dev 10:1812–1821PubMedCrossRefGoogle Scholar
  28. Mondragón-Palomino M, Theissen G (2011) Conserved differential expression of paralogous DEFICIENS- and GLOBOSA-like MADS-box genes in the flowers of Orchidaceae: refining the ‘orchid code’. Plant J 66:1008–1019PubMedCrossRefGoogle Scholar
  29. Moore RC, Purugganan MD (2005) The evolutionary dynamics of plant duplicate genes. Curr Opin Plant Biol 8:122–128PubMedCrossRefGoogle Scholar
  30. Oliver SN, Van Dongen JT, Alfred SC, Mamun EA, Zhao XC, Saini HS, Fernandes SF, Blanchard CL, Sutton BG, Geigenberger P, Dennis ES, Dolferus R (2005) Cold-induced repression of the rice anther-specific cell wall invertase gene OSINV4 is correlated with sucrose accumulation and pollen sterility. Plant, Cell Environ 28:1534–1551CrossRefGoogle Scholar
  31. Poupin MJ, Federici F, Medina C, Matus JT, Timmermann T, Arce-Johnson P (2007) Isolation of the three grape sub-lineages of B-class MADS-box TM6, PISTILLATA and APETALA3 genes which are differentially expressed during flower and fruit development. Gene 404:10–24PubMedCrossRefGoogle Scholar
  32. Rijpkema AS, Royaert S, Zethof J, van der Weerden G, Gerats T, Vandenbussche M (2006) Analysis of the Petunia TM6 MADS box gene reveals functional divergence within the DEF/AP3 lineage. Plant Cell 18:1819–1832PubMedCentralPubMedCrossRefGoogle Scholar
  33. Schwarz-Sommer Z, Hui I, Huijser P, Flor PJ, Hansen R, Tetens F, Lönnig WE, Saedler H, Sommer H (1992) Characterization of the Antirrhinum floral homeotic MADS-box gene deficiens: evidence for DNA binding and autoregulation of its persistent expression throughout flower development. EMBO J 11:251–263PubMedCentralPubMedGoogle Scholar
  34. Sharma B, Kramer E (2013) Sub- and neo-functionalization of APETALA3 paralogs have contributed to the evolution of novel floral organ identity in Aquilegia (columbine, Ranunculaceae). New Phytol 197:949–957PubMedCrossRefGoogle Scholar
  35. Sommer H, Beltran JP, Huijser P, Pape H, Lönnig WE, Saedler H, Schwarz-Sommer Z (1990) DEFICIENS, a homeotic gene involved in the control of flower morphogenesis in Antirrhinum majus: the protein shows homology to transcription factors. EMBO J 9:605–613PubMedCentralPubMedGoogle Scholar
  36. Theißen G, Saedler H (2001) Plant biology: floral quartets. Nature 409:469–471PubMedCrossRefGoogle Scholar
  37. Tröbner W, Ramirez L, Motte P, Hue I, Huijser P, Lönnig WE, Saedler H, Sommer H, Schwarz-Sommer Z (1992) GLOBOSA: a homeotic gene which interacts with DEFICIENS in the control of Antirrhinum floral organogenesis. EMBO J 11:4693–4704PubMedCentralPubMedGoogle Scholar
  38. Vandenbussche M, Zethof J, Royaert S, Weterings K, Gerats T (2004) The duplicated B-class heterodimer model: whorl-specific effects and complex genetic interactions in Petunia hybrid flower development. Plant Cell 16:741–754PubMedCentralPubMedCrossRefGoogle Scholar
  39. VanderSluis B, Bellay J, Musso G, Costanzo M, Papp B, Vizeacoumar FJ, Baryshnikova A, Andrews B, Boone C, Myers CL (2010) Genetic interactions reveal the evolutionary trajectories of duplicate genes. Mol Syst Biol 6:429PubMedCentralPubMedCrossRefGoogle Scholar
  40. Viaene T, Vekemans D, Irish VF, Geeraerts A, Huysmans S, Janssens S, Smets E, Geuten K (2009) Pistillata—duplications as a mode for floral diversification in (basal) asterids. Mol Biol Evol 26:2627–2645PubMedCrossRefGoogle Scholar
  41. Walter M, Chaban C, Schütze K, Batistic O, Weckermann K, Näke C, Blazevic D, Grefen C, Schumacher K, Oecking C, Harter K, Kudla J (2004) Visualization of protein interactions in living plant cells using bimolecular fluorescence complementation. Plant J 40:428–438PubMedCrossRefGoogle Scholar
  42. Wang L, Li ZC, He CY (2012) Transcriptome-wide mining of the differentially expressed transcripts for natural variation of floral organ size in Physalis philadelphica. J Exp Bot 63:6457–6465PubMedCentralPubMedCrossRefGoogle Scholar
  43. Winter KU, Weiser C, Kaufmann K, Bohne A, Kirchner C, Kanno A, Saedler H, Theißen G (2002) Evolution of class B floral homeotic proteins: obligate heterodimerization originated from homodimerization. Mol Biol Evol 19:587–596PubMedCrossRefGoogle Scholar
  44. Wuest SE, O’Maoileidigh DS, Rae L, Kwasniewska K, Raganelli A, Hanczaryk K, Lohan AJ, Loftus B, Graciet E, Wellmer F (2012) Molecular basis for the specification of floral organs by APETALA3 and PISTILLATA. Proc Natl Acad Sci USA 109:13452–13457PubMedCentralPubMedCrossRefGoogle Scholar
  45. Yao J, Dong Y, Morris BA (2001) Parthenocarpic apple fruit production conferred by transposon insertion mutations in a MADS-box transcription factor. Proc Natl Acad Sci USA 98:1306–1311PubMedCentralPubMedCrossRefGoogle Scholar
  46. Zahn LM, Leebens-Mack J, DePamphilis CW, Ma H, Theißen G (2005) To B or not to B a flower: the role of DEFICIENS and GLOBOSA orthologs in the evolution of the angiosperms. J Hered 96:225–240PubMedCrossRefGoogle Scholar
  47. Zhang JS, Li ZC, Zhao J, Zhang SH, Quan H, Zhao M, He CY (2014a) Deciphering the Physalis floridana double-layered-lantern1 mutant provides insights into functional divergence of the GLOBOSA duplicates within the Solanaceae. Plant Physiol 164:748–764PubMedCrossRefGoogle Scholar
  48. Zhang JS, Zhao J, Zhang SH, He CY (2014b) Efficient gene silencing mediated by tobacco rattle virus in an emerging model plant Physalis. PLoS One 9:e85534PubMedCentralPubMedCrossRefGoogle Scholar
  49. Zhao J, Tian Y, Zhang JS, Zhao M, Gong PC, Riss S, Saedler R, He CY (2013) The euAP1 protein MPF3 represses MPF2 to specify floral calyx identity and displays crucial roles in Chinese lantern development in Physalis. Plant Cell 25:2002–2021PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Shaohua Zhang
    • 1
    • 2
  • Ji-Si Zhang
    • 1
    • 2
    • 3
  • Jing Zhao
    • 1
    • 2
  • Chaoying He
    • 1
    Email author
  1. 1.State Key Laboratory of Systematic and Evolutionary BotanyInstitute of Botany, Chinese Academy of SciencesBeijingChina
  2. 2.University of Chinese Academy of SciencesBeijingChina
  3. 3.Anshan Normal UniversityAnshanChina

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