Question

The 2006 research paper by Paredez et al (full reference details below) provided direct evidence linking a role for microtubules in regulating cellulose microfibril deposition in the plant cell wall. Describe the major experimental findings reported in this paper and the significance of these findings to resolve long standing models that attempted to explain a role for cortical microtubules in controlling the direction of cellulose microfibril deposition. In explaining the experimental strategy used in this research, what was the rationale for using a CesA6 mutant line as part of their experimental approach?

Reference: Paredez, Somerville and Ehrhardt (2006) Visualization of cellulose synthase demonstrates functional association with microtubules. Science 312: 1491-1495.

Answer 1

Major findings

1. The process of cellulose deposition in living cells was demonstrated in a transgenic Arabidopsis having a fusion of a functional yellow fluorescent protein and cellulose synthase (CESA).
2. Confocal microscopy shows movement and alignment of CESA complexes in the plasma membrane. The CESA complexes are arranged at constant rates in linear tracks. This observation is similar to that of cortical microtubules. The movement of microtubules is bidirectional.
3. Cellulose synthase complexes containing CESA6 are organized in the cell membrane by a functional association with cortical microtubules.
4. If polymerization of microtubules is inhibited, distribution and movement pattern of CESA complexes changes in the membrane. This means that a direct correlation occurs between cellulose deposition by the cytoskeleton and movement of CESA complexes and cortical microtubules.
5. The distribution and movement of YFP:CESA6 is regulated by microtubules. A high level of coordination occurs between YFP:CESA6 and microtubules during reorganization events.
6. CESA localization and guidance is linked to spatial and temporal coupling with microtubules. Each cortical microtubule shows a lateral interaction with the cytosolic domains of CESA complexes. This helps in organization of CESA in linear arrays on either side of the microtubule. The arrangement suggests possible mechanisms for bidirectional movement of microtubules along CESA tracks.
7. Direct interaction of microtubules of CESA protein or linker proteins is not required for motility. The motive force for complex motility is provided primarily by cellulose polymerization.
8. Localization of CESA protein occurs in one type of tissue. CESA proteins do not have localization patterns but are tightly coupled to the cytoskeleton. The behavior of these proteins varies within different organs.
9. Microtubules confer orientation on the movement of the CESA complexes but are not required for motility of CESA.

Cortical microtubules play a major role in formation of cellulose synthase complex that help in the organization of CESA6 complexes. Each cortical microtubule forms a lateral interaction with the cytosolic domain of CESA complex, leading to organization of two linear arrays of CESA, one on either side of the microtubule. The bidirectional movement of microtubules guided by CESA tracks. Cortical microtubules influence the distribution and guidance of CESA rosettes. In the absence of cortical microtubules, the movement of CESA complexes is self organized or occurs by the action of a second extrinsic organizational mechanism.