New Technique Reveals Plant's Defense Mechanism

Researchers at the Danforth Plant Science Center and the University of Tennessee, Knoxville, working with Tessa Burch-Smith, PhD, are breaking new ground in their investigation of how viruses, essential molecules, and information are transferred between cells in plants.

In a recent study, they showed how callose, a carbohydrate polymer, is deposited when plants are responding to infection, controlling the structure known as Plasmo Desmata (PD), which connects neighboring cells in leaves and other organs.

Their work clears the path for a more comprehensive understanding of the functioning of the plant immune system by comparing various approaches to precisely quantify callose accumulation surrounding the microscopic PD channels. The research was published in the journal Molecular Plant-Microbe Interactions.

Glucose molecules form the polymer cellulose, which is necessary to control intercellular trafficking through plasmodesmata (PD). By eliminating callose at PD or, in the opposite scenario, by increasing callose accumulation at PD to prevent intercellular trafficking during infection, pathogens modify PD-localized proteins to facilitate intercellular trafficking.

Salicylic acid and other plant defense hormones control intercellular trafficking and PD-localized proteins during immune defense reactions like systemic acquired resistance.

Plant callose deposition at photoperiod (PD) has become a widely used technique to evaluate possible molecular trafficking between cells during plant immunity. There is no accepted method for taking these measurements, even though they are very popular.

To determine which of three commonly used methods for identifying and quantifying PD callose by aniline blue staining was most effective in the Nicotiana benthamiana leaf model, first author Amie Sankoh, PhD, and her undergraduate colleague Joseph Adjei conducted a comparison.  Amie and Joseph mainly used American Sign Language to communicate because they were deaf.

According to their findings, the most accurate technique for measuring callose deposition in fixed tissue involved the use of fluorescent microscopy and aniline blue staining. The results of comparing manual and semi-automated image analysis workflows were found to be similar, with the semi-automated workflow yielding a wider distribution of data points.

We were surprised at how different the reliability of the different methods for detecting callose could be. We think this work will greatly improve consistency in experiments across labs.”

Amie Sankoh PhD, Study First Author, University of Tennessee

The Danforth Center's Advanced Bioimaging Laboratory was utilized in this investigation. The team intends to look into how callose levels at PD fluctuate during infection with different hormones using the protocol and analysis they have found. These investigations may pinpoint critical windows for manipulating PD to impede the spread of infection and stop plant disease.