RIKEN researchers discover a new pathway for glucosinolate degrading enzymes in Arabidopsis thaliana

A team of researchers led by Senior Technical Scientist Kiminori Toyooka, Technical Staff Tomomi Goto, and Masami Yokota Hirai, Unit Leader of the Mass Spectrometry and Microscopy Unit at the RIKEN Center for Sustainable Resource Science, has utilized advanced electron microscopy techniques to uncover new insights into the transport of β‐Glucosidase in the lateral root cap cells of Arabidopsis thaliana.

The team discovered that β‐Glucosidases are transported directly to the vacuoles via endoplasmic reticulum (ER) bodies (ER organelle), a differentiated structure that buds from the ER without the need for transport through Golgi bodies.

Previous research has shown that in Arabidopsis leaves, the mixing of β‐glucosidase in the ER bodies and glucosinolates in the vacuoles releases a disease‐repellent substance when the vacuole breaks down due to insect damage. However, the mechanism of this process in roots was not well understood.

To address this gap in understanding, the research team took high‐resolution, TEM images of whole root tips from Arabidopsis thaliana shoots prepared using a high‐pressure freezing technique. As a result, they found numerous ER bodies in the lateral root caps, measuring 2‐4 micrometers in size. Three‐dimensional analysis using serial section scanning electron microscopy revealed that the ER bodies have an elongated, sausage‐like structure. The team then marked ER bodies in ring and longitudinal cross‐sections of whole root tips in the TEM imagery. They found that the ER bodies were present only in the lateral root caps and were frequently in contact with the vacuoles.

In addition, genetically transformed Arabidopsis plants labeled with red fluorescent protein for PYK10 (a type of β‐glucosidase), a primary enzyme in the ER body in leaves that has an ER‐retention signal, were used to analyze root tips prepared using the high‐pressure freezing technique and correlative light‐electron microscopy (CLEM). The study's results showed that the PYK10 red fluorescent protein was present in the ER bodies of the lateral root cap's inner and middle layer cells. From the inner to outermost layer cells of the lateral root cap, the ER bodies and vacuoles are frequently in contact, and the ER bodies protrude into the vacuoles. PYK10 does not localize to Golgi bodies or other cell organelles, suggesting that ER bodies in the lateral root caps transport PYK10 directly from the endoplasmic reticulum to the vacuoles without the need for transport through Golgi bodies.

Toyooka noted, 'When imaging ER bodies with fluorescently labeled enzymes using electron microscopy, we faced challenges in achieving high temporal resolution with CLEM while preserving fluorescence. Moving forward, we aim to study vacuole collapse resulting from ER bodies and vacuoles fusing and explore the connection between root morphogenesis and soil microorganisms in the rhizosphere.'

■ Glucosinolate: A specialized metabolite produced primarily by cruciferous plants. Also called mustard oil glycosides, when hydrolyzed by β‐glucosidase, they are converted into volatile substances such as isothiocyanate (mustard oil), which has insect‐repellent and carcinogenic effects.

■ β‐glucosidase: Enzyme that cuts β‐glucoside (sugar) bonds. It releases sugars from sugar‐bound compounds.

Journal Information
Publication: Plant & Cell Physiology
Title: Endoplasmic Reticulum Bodies in the Lateral Root Cap are Involved in the Direct Transport of Beta‐Glucosidase to Vacuoles
DOI: 10.1093/pcp/pcac177