2020

Sara Tedesco, Ana Pina, Pedro Fevereiro, Friedrich Kragler

Agronomy, 2020, doi: 10.3390/agronomy10050706

Grafting is the most used propagation method in viticulture and is the unique control strategy against Phylloxera. Nevertheless, its practice remains limited mainly due to inconsistent graft success and difficulties in predicting graft compatibility responses of proposed scion–rootstock combinations, slowing down the selection of elite rootstocks. Aiming to identify optimal phenotypic parameters related to graft (in)compatibility, we used four clones of two grapevine cultivars that show different compatibility behavior when grafted onto the same rootstock. Several physiological parameters, internal anatomy of the graft union, chlorophyll fluorescence, and pigment contents of homo- and heterografts were monitored in a nursery-grafting context. The measurements highlighted enhanced performance of the heterografts due to rooting difficulties of Vitis vinifera homografts. This suggests that in viticulture, homografts should only be used as compatibility controls regarding qualitative attributes. By observing the internal anatomy of the union, we found that grapevines might require longer times for graft healing than anticipated. While Affinity Coefficients were not informative to assess incompatibility, leaf chlorophyll concentration analysis proved to be a more sensitive indicator of stress than the analysis of chlorophyll fluorescence. Overall, we conclude that graft take correlated best with callus formation at the graft junction three weeks after grafting.

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Tedesco et al. Figure 2a_Characterisation of grafts

Fig. 2a: Internal characterization of the graft union. (a) Example images of the category A to E charactering the internal graft unions. Category A represents a perfect union in which the graft line is almost invisible. Category B shows few structural imperfections and/or slight discontinuities between wood and bark or cambial invaginations. Category C is characterized by bark discontinuities and D by wood discontinuities. Category E includes broken/unattached unions and/or unions with dead tissue in proximity of the union line.

Eleftheria Saplaoura, Valentina Perrera, Friedrich Kragler

Journal of visualized experiments – JoVE, 2020, doi:10.3791/61231

Secondary base modifications on RNA, such as m5C, affect the structure and function of the modified RNA molecules. Methylated RNA Immunoprecipitation and sequencing (MeRIP-seq) is a method that aims to enrich for methylated RNA and ultimately identify modified transcripts. Briefly, sonicated RNA is incubated with an antibody for 5-methylated cytosines and precipitated with the assistance of protein G beads. The enriched fragments are then sequenced and the potential methylation sites are mapped based on the distribution of the reads and peak detection. MeRIP can be applied to any organism, as it does not require any prior sequence or modifying enzyme knowledge. In addition, besides fragmentation, RNA is not subjected to any other chemical or temperature treatment. However, MeRIP-seq does not provide single-nucleotide prediction of the methylation site as other methods do, although the methylated area can be narrowed down to a few nucleotides. The use of different modification-specific antibodies allows MeRIP to be adjusted for the different base modifications present on RNA, expanding the possible applications of this method.

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Fig. 1: RNA samples are incubated with an antibody for 5-methylated cytosines and the complexes are pulled down with protein G magnetic beads that capture the antibodies along with the bound RNA. The eluted RNA samples are analyzed by deep sequencing and qRT-PCR.

2019

Lei Yang, Valentina Perrera, Eleftheria Saplaoura, Federico Apelt, Mathieu Bahin, Amira Kramdi, Justyna Olas, Bernd Mueller-Roeber, Ewelina Sokolowska, Wenna Zhang, Runsheng Li, Nicolas Pitzalis, Manfred Heinlein, Shoudong Zhang, Auguste Genovesio, Vincent Colot, Friedrich Kragler

Current Biology, 2019, doi: 10.1016/j.cub.2019.06.042

In plants, transcripts move to distant body parts to potentially act as systemic signals regulating development and growth. Thousands of messenger RNAs (mRNAs) are transported across graft junctions via the phloem to distinct plant parts. Little is known regarding features, structural motifs, and potential base modifications of transported transcripts and how these may affect their mobility. We identified Arabidopsis thaliana mRNAs harboring the modified base 5-methylcytosine (m5C) and found that these are significantly enriched in mRNAs previously described as mobile, moving over graft junctions to distinct plant parts. We confirm this finding with graft-mobile methylated mRNAs TRANSLATIONALLY CONTROLLED TUMOR PROTEIN 1 (TCTP1) and HEAT SHOCK COGNATE PROTEIN 70.1 (HSC70.1), whose mRNA transport is diminished in mutants deficient in m5C mRNA methylation. Together, our results point toward an essential role of cytosine methylation in systemic mRNA mobility in plants and that TCTP1 mRNA mobility is required for its signaling function.

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