Transverse Sectioning of Mature Rice (Oryza sativa L.) Kernels for Scanning Electron Microscopy Imaging Using Pipette Tips as Immobilization Support.

Starch granules (SGs) exhibit different morphologies depending on the plant species, especially in the endosperm of the Poaceae family. Endosperm phenotyping can be used to classify genotypes based on SG morphotype using scanning electron microscopic (SEM) analysis. SGs can be visualized using SEM by slicing through the kernel (pericarp, aleurone layers, and endosperm) and exposing the organellar contents.

Plastid Envelope-Localized Proteins Exhibit a Stochastic Spatiotemporal Relationship to Stromules.

Unlabelled: Plastids in the viridiplantae sporadically form thin tubules called stromules that increase the interactive surface between the plastid and the surrounding cytoplasm. Several recent publications that report observations of certain proteins localizing to the extensions have then used the observations to suggest stromule-specific functions. The mechanisms by which specific localizations on these transient and sporadically formed extensions might occur remain unclear.

Chloroplast behaviour and interactions with other organelles in pavement cells.

Chloroplasts are a characteristic feature of green plants. Mesophyll cells possess the majority of chloroplasts and it is widely believed that, with the exception of guard cells, the epidermal layer in most higher plants does not contain chloroplasts. However, recent observations on have shown a population of chloroplasts in pavement cells that are smaller than mesophyll chloroplasts and have a high stroma to grana ratio.

Fluorescent Protein Aided Insights on Plastids and their Extensions: A Critical Appraisal.

Multi-colored fluorescent proteins targeted to plastids have provided new insights on the dynamic behavior of these organelles and their interactions with other cytoplasmic components and compartments. Sub-plastidic components such as thylakoids, stroma, the inner and outer membranes of the plastid envelope, nucleoids, plastoglobuli, and starch grains have been efficiently highlighted in living plant cells. In addition, stroma filled membrane extensions called stromules have drawn attention to the dynamic nature of the plastid and its interactions with the rest of the cell.

Mitochondrial pleomorphy in plant cells is driven by contiguous ER dynamics.

Mitochondria are pleomorphic, double membrane-bound organelles involved in cellular energetics in all eukaryotes. Mitochondria in animal and yeast cells are typically tubular-reticulate structures and several micro-meters long but in green plants they are predominantly observed as 0.2-1.

The myth of interconnected plastids and related phenomena.

Studies spread over nearly two and a half centuries have identified the primary plastid in autotrophic algae and plants as a pleomorphic, multifunctional organelle comprising of a double-membrane envelope enclosing an organization of internal membranes submerged in a watery stroma. All plastid units have been observed extending and retracting thin stroma-filled tubules named stromules sporadically. Observations on living plant cells often convey the impression that stromules connect two or more independent plastids with each other.

Simultaneous live-imaging of peroxisomes and the ER in plant cells suggests contiguity but no luminal continuity between the two organelles.

Transmission electron micrographs of peroxisomes in diverse organisms, including plants, suggest their close association and even luminal connectivity with the endoplasmic reticulum (ER). After several decades of debate de novo peroxisome biogenesis from the ER is strongly favored in yeasts and mammals. Unfortunately many of the proteins whose transit through the ER constitutes a major evidence for peroxisome biogenesis from the ER do not exhibit a similar localization in plants.

Organelle extensions in plant cells.

Cell walls lock each cell in a specific position within the supra-organization of a plant. Despite its fixed location, each cell must be able to sense alterations in its immediate environment and respond rapidly to ensure the optimal functioning, continued growth and development, and eventual long-term survival of the plant. The ultra-structural detail that underlies our present understanding of the plant cell has largely been acquired from fixed and processed material that does not allow an appreciation of the dynamic nature of sub-cellular events in the cell.

Differential coloring reveals that plastids do not form networks for exchanging macromolecules.

Stroma-filled tubules named stromules are sporadic extensions of plastids. Earlier, photobleaching was used to demonstrate fluorescent protein diffusion between already interconnected plastids and formed the basis for suggesting that all plastids are able to form networks for exchanging macromolecules. However, a critical appraisal of literature shows that this conjecture is not supported by unequivocal experimental evidence.

Green-to-red photoconvertible mEosFP-aided live imaging in plants.

Numerous subcellular-targeted probes have been created using a monomeric green-to-red photoconvertible Eos fluorescent protein for understanding the growth and development of plants. These probes can be used to create color-based differentiation between similar cells, differentially label organelle subpopulations, and track subcellular structures and their interactions. Both green and red fluorescent forms of mEosFP are stable and compatible with single colored FPs.

Color recovery after photoconversion of H2B::mEosFP allows detection of increased nuclear DNA content in developing plant cells.

Many higher plants are polysomatic whereby different cells possess variable amounts of nuclear DNA. The conditional triggering of endocycles results in higher nuclear DNA content (C value) that in some cases has been correlated to increased cell size. While numerous multicolored fluorescent protein (FP) probes have revealed the general behavior of the nucleus and intranuclear components, direct visualization and estimation of changes in nuclear-DNA content in live cells during their development has not been possible.

Correlated behavior implicates stromules in increasing the interactive surface between plastids and ER tubules.

Stromules are extended by plastids but the underlying basis for their extension and retraction had not been understood until recently. Our live-imaging aided observations on coincident plastid stromule branching and ER tubule dynamics open out new areas of investigation relating to these rapid subcellular interactions. The addendum provides a testable hypothesis on the formation of stromules, which argues against the need for new membrane incorporation and suggests that stromal extensions might result from a remodeling of the plastid envelope membrane in an ER aided manner.

Plastid stromule branching coincides with contiguous endoplasmic reticulum dynamics.

Stromules are stroma-filled tubules extending from plastids whose rapid extension toward or retraction from other plastids has suggested a role in interplastidic communication and exchange of metabolites. Several studies point to sporadic dilations, kinks, and branches occurring along stromule length but have not elucidated the underlying basis for these occurrences. Similarly, although specific details on interacting partners have been missing, a consensus viewpoint suggests that stromules increase the interactive surface of a plastid with its cytoplasmic surroundings.