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Mannans and endo-β-mannanase transcripts are located in different seed compartments during Brassicaceae germination

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Abstract

Main conclusion

Mannans but not endo-β-mannanases are mainly found in the mucilage layer of two Brassicaceae seeds. Nonetheless, mannanase mobilization from inner to outer seed layers cannot be ruled out.

The contribution of endo-β-mannanase (MAN) genes to the germination of the wild-type Sisymbrium officinale and cultivated Brassica rapa (Brassicaceae) species has been explored. In both species, mannans have been localized to the imbibed external seed coat layer (mucilage) by fluorescence immunolocalization and MAN enzymatic activity increases in seeds as imbibition progresses, reaching a peak before 100% germination is achieved. The MAN gene families have been annotated and the expression of their members analyzed in vegetative and reproductive organs. In S. officinale and B. rapa, MAN2, MAN5, MAN6, and MAN7 transcripts accumulate upon seed imbibition. SoMAN7 is the most expressed MAN gene in S. officinale germinating seeds, as occurs with its ortholog in Arabidopsis thaliana, but in B. rapa, the most abundant transcripts are BrMAN2 and BrMAN5. These genes (MAN2, MAN5, MAN6, and MAN7) are localized, by mRNA in situ hybridization, to the micropylar at the endosperm layer and to the radicle in S. officinale, but in B. rapa, these mRNAs are faintly found to the micropylar living seed coat layer and are mainly present at the radicle tip and the vascular bundles. If the domestication process undergone by B. rapa is responsible for these different MAN expression patterns, upon germination remains to be elucidated. Since mannans and MAN genes are not spatially distributed in the same seed tissues, a movement of MAN enzymes that are synthesized with typical signal peptides from the embryo tissues to the mucilage layer (via apoplastic space) is necessary for the mannans to be hydrolyzed.

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Abbreviations

CWME:

Cell wall modifying enzymes

LSCL:

Living seed coat layer

MAN:

Endo-β-mannanase

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Acknowledgements

We thank Prof. P. Carbonero (Universidad Politécnica de Madrid, Centro de Biotecnología y Genómica de Plantas, Madrid, Spain) for critical reading of the manuscript. Financial support was granted by Ministerio de Ciencia e Innovación (CGL2009-11425; MICINN, Spain).

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Authors and Affiliations

  1. Departamento de Fisiología Vegetal, Facultad de Farmacia, Universidad de Santiago de Compostela, 15780, Santiago de Compostela, Spain

    Néstor Carrillo-Barral, Angel J. Matilla & María del Carmen Rodríguez-Gacio

  2. Centro de Biotecnología y Genómica de Plantas (UPM-INIA), Universidad Politécnica de Madrid, Campus de Montegancedo, Pozuelo de Alarcón, 28223, Madrid, Spain

    Raquel Iglesias-Fernández

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  1. Néstor Carrillo-Barral
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Correspondence to Raquel Iglesias-Fernández.

Electronic supplementary material

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425_2017_2815_MOESM1_ESM.tif

Fig. S1 Transcription levels (Ct values) of the housekeeping SoACT8 and BrACT8 during seed germination (a and d), silique development (b and e) and in different organs (c and f) of S. officinale and B. rapa (TIFF 34544 kb)

425_2017_2815_MOESM2_ESM.tif

Fig. S2 a Expression analysis by RT-qPCR of SoMAN2, SoMAN5, SoMAN6, and SoMAN7 in several plant organs of S. officinale. b Transcript abundance of BrMAN2, BrMAN5, BrMAN6, and BrMAN7 in different plant organs of B. rapa. Data are means ± SE of two technical replicates of three biological samples (TIFF 32187 kb)

Table S1 Sequences of primers used (TIFF 7137 kb)

425_2017_2815_MOESM4_ESM.tif

Table S2 Major characteristics of the endo-β-mannanase proteins analyzed in S. officinale and B. rapa. Those that are important in the germination of Arabidopsis thaliana seeds are also included (Iglesias-Fernández et al. 2011a) (TIFF 9586 kb)

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Carrillo-Barral, N., Matilla, A.J., Rodríguez-Gacio, M.d.C. et al. Mannans and endo-β-mannanase transcripts are located in different seed compartments during Brassicaceae germination. Planta 247, 649–661 (2018). https://doi.org/10.1007/s00425-017-2815-4

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