Morpho-histological, histochemical, and molecular evidences related to cellular reprogramming during somatic embryogenesis of the model grass Brachypodium distachyon

Abstract

The wild grass species Brachypodium distachyon (L.) has been proposed as a new model for temperate grasses. Among the biotechnological tools already developed for the species, an efficient induction protocol of somatic embryogenesis (SE) using immature zygotic embryos has provided the basis for genetic transformation studies. However, a systematic work to better understanding the basic cellular and molecular mechanisms that underlie the SE process of this grass species is still missing. Here, we present new insights at the morpho-histological, histochemical, and molecular aspects of B. distachyon SE pathway. Somatic embryos arose from embryogenic callus formed by cells derived from the protodermal-dividing cells of the scutellum. These protodermal cells showed typical meristematic features and high protein accumulation which were interpreted as the first observable steps towards the acquisition of a competent state. Starch content decreased along embryogenic callus differentiation supporting the idea that carbohydrate reserves are essential to morphogenetic processes. Interestingly, starch accumulation was also observed at late stages of SE process. Searches in databanks revealed three sequences available annotated as BdSERK, being two copies corresponding to SERK1 and one showing greater identity to SERK2. In silico analysis confirmed the presence of characteristic domains in a B. distachyon Somatic Embryogenesis Receptor Kinase genes candidates (BdSERKs), which suggests SERK functions are conserved in B. distachyon. In situ hybridization demonstrated the presence of transcripts of BdSERK1 in all development since globular until scutellar stages. The results reported in this study convey important information about the morphogenetic events in the embryogenic pathway which has been lacking in B. distachyon. This study also demonstrates that B. distachyon provides a useful model system for investigating the genetic regulation of SE in grass species.

High-Resolution Linkage Map With Allele Dosage Allows the Identification of Regions Governing Complex Traits and Apospory in Guinea Grass (Megathyrsus maximus)

“…In nucellar cells of apomictic genotypes of Poa pratensis, the SERK gene is involved in embryo sac development (Albertini et al, 2005). Recently, SERK was reported in Brachypodium distachyon grass as having a domain conserved among monocots and plays a prominent role in apomixis (Oliveira et al, 2017). Scarce studies have investigated the genes involved in the regulation of reproductive events in guinea grass, and these previous studies mainly utilized of transcriptome data (Toledo-Silva et al, 2013;Radhakrishna et al, 2018).…”

<b>Determination of endogenous IAA and carbohydrates during the induction and development of protocorm-like bodies of <i>Cattleya tigrina</i> A. Richard

“…Starch contents remained constant during the first 30 days in Acca sellowiana somatic embryo development, although these values significantly decreased compared to the initial inoculum (Cangahuala-Inocente et al, 2014). Starch content decrease may be related to its degradation to produce glucose as an energy source for various metabolic reactions that occur during cell vital metabolism growth and maintenance (Oliveira et al, 2017). In this context, the reduction in starch granules observed in dividing cells at the developmental stages of PLBs in C. tigrina suggests that these were required for the development of these structures.…”

In Vitro Tissue Culture in Brachypodium: Applications and Challenges

“…The molecular basis is completely obscure, but it may be due to cellular differentiation. Embryogenic cells have the typical features of undifferentiated cells such as a small or poorly developed vacuole, dense cytoplasm and a nucleus that usually has one and occasionally two nucleoli [98]. Mitoses are also occasionally present [99].…”

“…The cell wall composition also changes. An embryogenic callus that was formed by cells that had been derived from the protodermal-dividing cells of the scutellum [98] initially had typical meristematic features and a high level of protein accumulation, which is connected with the acquisition of a competent state [100]. Embryogenic calli have an extracellular matrix on the surface of the calli cells (ECMSN), which is composed of the arabinogalactan proteins (AGPs) and pectins [99].…”