Microtubules and guard-cell morphogenesis in Zea mays L
Authors: Galatis B. 
Published: April 2021

ABSTRACT
In median paradermal planes of very young guard cells of Zea mays, numerous anticlinally oriented microtubules line densely the whole length of the ventral wall. In the external and internal regions of this wall, the subplasmalemmal microtubules are restricted to the middle of its length, where local thickenings start being deposited. In periclinal walls they were observed converging from their ends towards the thickenings. Few microtubules are present in the rest of the anticlinal walls. Before initiation of the thickenings, the parietal cytoplasm of the periclinal walls around the middle of the ventral wall contains a large number of microtubules diverging from this region and intimately associated with numerous dictyosome vesicles. Microtubule-organizing centres (MTOCs) of both periclinal and ventral walls seem to operate in these areas.
The thickening of the middle of the ventral wall is initially limited at its external and internal ends. In these regions, local pads of thickening are gradually deposited, also covering a significant part of the periclinal walls, particularly the external ones, while the microtubules around them proliferate. A number of microtubules are found at a significant distance from the thickenings. The mid-depth region of the ventral wall is obviously thickened before stomatai pore opening. In this region the microtubules also become confined around the thickening. Progressively, material is apposed to the unthickened mid-regions of the periclinal walls. In the latter, both microtubulesand microfibrils exhibit a clear radial arrangement around the margins of the ventral wall thickenings. The cytoplasm surrounding them possesses abundant smooth dictyosome vesicles, exhibiting intimate associations with microtubules, and some coated ones, both positive to the PA-TCH-SP reaction. The above wall differentiation is followed by stomatai pore opening, which commences initially from the internal and later from the external ventral wall thickenings and proceeds inwards; this process is also completed in dark-grown leaves.
During later stages of morphogenesis, the guard cells increase considerably in length, becoming thin in their middle region. The stomatai pore elongates, and the central canal is formed. The mid-canal microtubules do not initially exhibit a definite orientation; later they become parallel to the cell axis, an alignment followed by the microfibrils of the wall. The microtubules of the margins of the extending canal are more grouped and retain a rather consistent orientation. In the ventral wall they are usually oriented transversely to the leaf surface. In periclinal walls some of them are directed at an angle to the lateral walls bordering the terminal canal thickenings, and others radiate towards the bulbous ends of the guard cells. In this case, the microtubules also appear parallel to the microfibrils. An increased protoplasmic activity, especially marked by the proliferation of dictyosome and ER membranes, accompanies an intense thickening, initially of the periclinal and later of the ventral wall of the canal. The microtubules underlie the thickening ventral and periclinal walls, but are absent from the non-thickening lateral walls of the canal. Finally, the periclinal, the transverse, and a major part of the lateral walls of the bulbous ends of the guard cells become thickened. Microtubules again line these thickening wall regions.
The observations suggest that microtubules play a critical role in guard-cell morphogenesis in Zea mays. Apart from that, extensive cell elongation seems to be an essential shaping factor of the dumbell-shaped guard cells.