Research

Correct growth and development in plants is inextricably linked with the ability of cells to produce and/or modify their cell walls. Thus, a key challenge in plant biology is to understand the structure of cell walls and identify the mechanisms and proteins that are required to synthesize and modify them.

The epidermal cells of the Arabidopsis seed coat undergo a complex differentiation process in which there is sequential synthesis of pectinaceous mucilage followed by secondary cell wall production. As a result, at maturity these cells contain a donut-shaped pocket of mucilage subtended by a volcano-shaped secondary cell wall. Wetting of mature seeds leads to the release of the hydrated mucilage gel, a process that appears to require cell wall weakening through modifying enzymes.

The Western Lab uses these mucilage secretory cells as a model to study the processes and regulation of cell wall production and modification during plant development. In addition, the polar secretion of pectin during mucilage production also makes this system a good model with which to dissect the mechanisms of targeted secretion in plants.

A Preliminary Model for the Regulation of Mucilage Secretory Cell Differentiation

Recent cloning of the MUCILAGE-MODIFIED4 (MUM4 = RHM2) gene, mutants of which have reduced amounts of mucilage, revealed that it encodes a putative rhamnose synthase. Similar phenotypes have been found for loss-of-function mutations in the developmental regulator APETALA2 (AP2) and the epidermal cell differentiation factors TRANSPARENT TESTA1 (TTG1), GLABRA2 (GL2) and TTG2, as well as MYB61. Transcriptional analysis has allowed the construction of a preliminary model for the regulation of mucilage production in Arabidopsis seed coats. At least two pathways are required for mucilage production. One pathway involves the regulation of rhamnose synthesis (MUM4) whereby TTG1 and AP2 activate GL2 expression, which in turn leads to increased transcription of MUM4. Another one or more pathways exist that appear to be regulated by the TTG2 and MYB61 transcription factors.

Filling in the Model:

1. Identification of MUM EHANCERS (MEN)
The above model for the regulation mucilage production in the mucilage secretory cells is a preliminary framework wherein only the basic regulation of MUM4 expression/rhamnose synthesis has been elucidated. Other players acting in the parallel pathways have yet to be determined, either at the biosynthetic level or other regulators acting along with, upstream or downstream of TTG2 and MYB61. In addition, there may be further genes acting at other levels/pathways that have not yet been considered. In order to identify further genes acting in mucilage secretory cell differentiation, we are performing an enhancer/suppressor screen in the sensitized background of mum4 mutants, which produce only a small amount of mucilage. A number of putative enhancer lines have been identified that are undergoing analysis.


Enhancer

mum4

Suppressor


2. Reverse Genetics
The differentiation of mucilage secretory cells requires the biosynthesis and secretion of large quantities of pectins and other complex carbohydrates. The sequencing and annotation of the Arabidopsis genome, not to mention a number of public functional genomics resources, allows the identification of candidate genes. Knockout mutants can be obtained and studied for the effect on mucilage secretory cell differentiation, while the gene expression in the cells can be studied with a variety of techniques.

PRAIRIE and PATCHY, Two Novel Genes Required for Mucilage Secretory Cell Differentiation

prairie (pra) mutants resemble those of MUM4, TTG1, GL2, TTG2 and MYB61. Preliminary genetic and molecular analyses place PRA in a pathway independent from those involving MUM4 and MYB61.

patchy (pty) is a novel member of an unexpected set of mutants that are affected in mucilage release from mature seeds. pty mutants release mucilage only from a random set of cells, though they appear to make normal mucilage. Recent cloning of PTY reveals that it encodes a putative cell wall modification enzyme that presumably acts in cell wall weakening to aid mucilage release upon seed hydration.


Cell Biology of Mucilage Secretory Cell Differentiation

A fascinating question is the regulation of intracellular morphogenesis in mucilage secretory cell differentiation. At maturity, the mucilage is found in a donut shape at the top of the cell, sitting on a volcano-shaped cell wall. How does this occur?

Targeted accumulation of mucilage in a ring at the top of the cell probably requires a combination of polar secretion and changes in plasma membrane-cell wall attachment in these areas. Formation of the tight cytoplasmic column reflected in the shape of the secondary cell wall appears to be at least partially due to the accumulation of mucilage (causing vacuolar constriction by displacement and/or osmotically induced loss of water).

 

 

Last update: July 25, 2012