In this article we will discuss about the mechanism of changes occurs in the shape of cells during morphogenesis.
Electron microscopic studies of cells undergoing changes in shape have shown that a peculiar group of cytoplasmic structures are involved in these processes. They are the microtubules and microfilaments. The microtubules are seen as long, straight, un-branched rods, 200 to 250 Å in diameter, which appear hollow in cross sections.
In cells not undergoing changes in shape the microtubules may be observed to traverse the cytoplasm at random. In cells which are elongating in a certain direction, however, the microtubules are arranged in parallel bundles running in the direction in which the cell is elongating.
Bundles of such microtubules are prominent in the narrowed “necks” of the bottle-shaped cells of the amphibian blastopore, in the cells immigrating from the primitive streak in birds, and in the cells of the neural plate. Bunches of microtubules may be seen along the length of outgrowing processes of nerve cells, and they are also involved in mitosis, as the fibers of the achromatic spindle.
It was originally with the spindle fibers of mitosis that the microtubules were found to be very sensitive to certain poisons, namely, colchicine (an alkaloid extracted from plants of the lily family) and also vinblastine sulfate. In cells treated with these poisons the microtubules disintegrate, and as a result the movements of the mitotic chromosomes stop, and mitosis is arrested in metaphase.
When applied to growing nerve cell processes, colchicine causes the collapse of the axon, which shortens, loses its rectilinear shape, and may eventually be completely withdrawn into the cell body. When the chick primitive streak is treated with colchicine, the elongating cells shorten, and the primitive groove straightens out.
The same happens with the elongating cells of the neural plate. These experiments show that the microtubules are involved in changes in the shape of the cells and particularly in the elongation of cells and the maintenance of the elongated form. The collapse of the outgrowing nerve cell processes suggests that the microtubules have a supporting or skeletal function.
This explanation should not, however, be understood in too narrow a sense. Although the microtubules appear to be able to increase in length, either by growth at the ends or by intussusception along their whole length, and thus to provide continuous support to elongating parts of the cells, it would be premature to conclude that the microtubules actually “push” the protruding or elongating parts.
Rather it may be suggested that the microtubules in some way lead the flow of cytoplasm in the direction which is indicated by the oriented arrangement of the microtubule bundles. This appears to be the role of microtubules in mitosis, in which chromosomes attached to the spindle fibers are drawn to the opposite poles of the achromatic figure.
The poisons which disrupt microtubules and prevent elongation of cells do not affect, to any notable extent, other processes in the cell; in particular, they do not prevent synthetic processes from occurring. Their influence on microtubules thus appears to be a direct one.
The other cell organoid as taking part in changes in cell form, the microfilaments, are much finer threadlike bodies, which are 30 to 60 Å in diameter. They appear to be contractile elements, and accordingly bundles of microfilaments are found in positions where their contraction could cause a narrowing or constriction of parts of the cells.
In cells undergoing cleavage (the egg during its first division and subsequently during the division of blastomeres), the microfilaments appear in the form of a “contractile ring” just underneath the cleavage furrow. As the “ring” narrows, the cleavage furrow cuts deeper and eventually separates the two daughter blastomeres. Colchicine has no effect on microfilaments, but these organoids are highly sensitive to another poison, cytochalazin B, a cyclic organic compound which is extracted from a mold, Helminthosporium dematoideum.
Cytochalazin B reversibly destroys the microfilaments, transforming bundles of these fibers into granular amorphous cytoplasm. When cleaving eggs are treated with this poison, the contractile ring consisting of the microfilaments is destroyed, cleavage stops immediately, and the cleavage furrows are reversed, and that is, they become smoothed out again.
Systems of microfilaments have been found to participate in changes in the shape of cells which are instrumental in producing folding of epithelial layers. In neural plate cells, in addition to the microtubules, which are responsible for elongation of the cells, there are bundles of microfilaments which are arranged parallel to the distal surfaces of the cells. If sections of the neural plate are made parallel to its surface, in each cell the microfilaments can be seen to be arranged in a ring around the cell’s distal end.
The bundle of filaments works as a “purse string,” literally pulling together the distal part of the cell and transforming it into a narrowed “neck.” A similar mechanism must be involved in the narrowing of the outer ends of cells and in their transformation into bottle-shaped cells in the blastopore during gastrulation. This conclusion is supported by the fact that cytochalazin B stops invagination of the archenteron in sea urchins and in the worm Urechis.
Most convincing experiments regarding the role of microfilaments in morphogenesis have been performed on oviducal glands in chickens. In immature birds the oviducts are simple tubes consisting of cuboidal epithelium. At sexual maturity, in response to estrogen, numerous oviducal glands are formed as simple out-pocketing’s of the oviducal epithelium.
The epithelium of the oviduct, however, is already responsive to estrogen in 7- to 12-day-old chicks. If they are injected with estrogen, their oviducts start developing glands within 24 hours. Thus, the change in shape of the cells and the resulting morphogenetic process can be studied under accurately controlled conditions, especially with respect to its timing.
Even before the cells in the gland rudiment start changing in shape, a layer of microfilaments appears close to the luminal surface of the cells. Contraction of the microfilaments pulls the luminal surface of the cells together. The cells become cone-shaped, with the narrow ends facing the cavity of the oviduct, and the gland becomes discernible as a pocket-like depression in the oviducal wall.
To prove that the microfilaments are, in fact, causing the invagination of the gland, the oviduct is excised and kept in vitro in a medium containing cytochalazin B. As a result, the microfilaments disintegrate and become a granular mass, the cells change back to the cuboidal shape, and the depression which was to be the oviducal gland becomes smoothed out.
The action of the cytochalazin B is reversible. If the oviducts are again placed in cytochalazin-free medium, the bundles of microfilaments reappear. Reversal of cytochalazin B action can occur while protein synthesis in the affected cells-is suppressed by another metabolic poison (cycloheximide). Thus, the formation of the microfilaments does not involve a synthesis of new proteins but is the result of a reorganization of the existing materials in the competent cells.
As with hair rudiments, once the oviducal glands take shape, their further development depends on the processes of growth and cell division setting in. This phase of development can then be suppressed by exposing the gland rudiments to a chemical which blocks DNA synthesis (hydroxyurea).