Cleavage:
After fertilization, the development of a multicellular organism proceeds by a process called cleavage, a series of mitotic divisions whereby the enormous volume of egg cytoplasm is divided into numerous smaller, nucleated cells.
These cleavage-stage cells are called blastomeres. In most species (mammals being the chief exception), the rate of cell division and the placement of the blastomeres with respect to one another is completely under the control of the proteins and mRNAs stored in the oocyte by the mother.
The zygotic genome, transmitted by mitosis to all the new cells, does not function in early-cleavage embryos. Few, if any, mRNAs are made until relatively late in cleavage, and the embryo can divide properly even when chemicals are used experimentally to inhibit transcription.
During cleavage, however, cytoplasmic volume does not increase. Rather, the enormous volume of zygote cytoplasm is divided into increasingly smaller cells.
First the egg is divided in half, then quarters, then eighths, and so forth. This division of egg cytoplasm without increasing its volume is accomplished by abolishing the growth period between cell divisions (that is, the G1 and G2 phases of the cell cycle).
Meanwhile, the cleavage of nuclei occurs at a rapid rate never seen again (not even in tumor cells). A frog egg, for example, can divide into 37,000 cells in just 43 hours.
Mitosis in cleavage-stage Drosophila embryos occurs every 10 minutes for over 2 hours and in just 12 hours forms some 50,000 cells.
This dramatic increase in cell number can be appreciated by comparing cleavage with other stages of development.
Once consequence of this rapid cell division is that the ratio of cytoplasmic to nuclear volume gets increasingly smaller as cleavage progresses. In many types of embryos (such as those of Xenopus and Drosophila, but not those of C. elegans or mammals), this decrease in the cytoplasmic to nuclear volume ratio is crucial in timing the activation of certain genes.
For example, in the frog Xenopus laevis, transcription of new messages is not activated until after 12 divisions. At that time, the rate of cleavage decreases, the blastomeres become motile, and nuclear genes begin to be transcribed.
This stage is called the mid-blastula transition. It is thought that some factor in the egg is being titrated by the newly made chromatin, because the time of this transition can be changed by experimentally altering the ratio of chromatin to cytoplasm in the cell (Newport and Kirschner 1982a,b; Edgar et al. 1986).
Thus, cleavage begins soon after fertilization and ends shortly after the stage when the embryo achieves a new balance between nucleus and cytoplasm.
From fertilization to cleavage:
The transition from fertilization to cleavage is caused by the activation of mitosis promoting factor (MPF). MPF was first discovered as the major factor responsible for the resumption of meiotic cell divisions in the ovulated frog egg.
It continues to play a role after fertilization, regulating the biphasic cell cycle of early blastomeres. Blastomeres generally progress through a cell cycle consisting of just two steps: M (mitosis) and S (DNA synthesis) (Figure 8.2). Gerhart and co-workers (1984) showed that MPF undergoes cyclical changes in its level of activity in mitotic cells.
The MPF activity of early blastomeres is highest during M and undetectable during S. Newport and Kirschner (1984) demonstrated that DNA replication (S) and mitosis (M) are driven solely by the gain and loss of MPF activity.
Cleaving cells can be experimentally trapped in S phase by incubating them in an inhibitor of protein synthesis. When MPF is microinjected into these cells, they enter M.
Their nuclear envelope breaks down and their chromatin condenses into chromosomes. After an hour, MPF is degraded and the chromosomes return to S phase.
Mitosis-promoting factor contains two subunits. The large subunit is called cyclin B. It is this component that shows a periodic behavior, accumulating during S and then being degraded after the cells have reached M (Evans et al. 1983; Swenson et al. 1986).
Cyclin B is often encoded by mRNAs stored in the oocyte cytoplasm, and if the translation of this message is specifically inhibited, the cell will not enter mitosis (Minshull et al. 1989).
The presence of cyclin B depends upon its synthesis and its degradation. Cyclin B regulates the small subunit of MPF, the cyclin-dependent kinase.
This kinase activates mitosis by phosphorylating several target proteins, including histones, the nuclear envelope lamin proteins, and the regulatory subunit of cytoplasmic myosin.
This brings about chromatin condensation, nuclear envelope depolymerization, and the organization of the mitotic spindle.
Without cyclin, the cyclin-dependent kinase will not function. The presence of cyclin is controlled by several proteins that ensure its periodic synthesis and degradation.
In most species studied, the regulators of cyclin (and thus, of MPF) are stored in the egg cytoplasm. Therefore, the cell cycle is independent of the nuclear genome for numerous cell divisions.
These early divisions tend to be rapid and synchronous. However, as the cytoplasmic components are used up, the nucleus begins to synthesize them.
The embryo now enters the mid-blastula transition, in which several new phenomena are added to the biphasic cell divisions of the embryo. First, the growth stages (G1 and G2) are added to the cell cycle, permitting the cells to grow.
Before this time, the egg cytoplasm was being divided into smaller and smaller cells, but the total volume of the organism remained unchanged. Xenopus embryos add those phases to the cell cycle shortly after the twelfth cleavage. Drosophila adds G2 during cycle 14 and G1 during cycle 17 (Newport and Kirschner 1982a; Edgar et al. 1986).
Second, the synchronicity of cell division is lost, as different cells synthesize different regulators of MPF. Third, new mRNAs are transcribed. Many of these messages encode proteins that will become necessary for gastrulation.
If transcription is blocked, cell division will occur at normal rates and at normal times in many species, but the embryo will not be able to initiate gastrulation.
Patterns of embryonic cleavage:
In 1923, embryologist E. B. Wilson reflected on how little we knew about cleavage: “To our limited intelligence, it would seem a simple task to divide a nucleus into equal parts. The cell, manifestly, entertains a very different opinion.” Indeed, different organisms undergo cleavage in distinctly different ways.
The pattern of embryonic cleavage particular to a species is determined by two major parameters: the amount and distribution of yolk protein within the cytoplasm, and factors in the egg cytoplasm that influence the angle of the mitotic spindle and the timing of its formation.
The amount and distribution of yolk determines where cleavage can occur and the relative size of the blastomeres. When one pole of the egg is relatively yolk-free, the cellular divisions occur there at a faster rate than at the opposite pole.
The yolk-rich pole is referred to as the vegetal pole; the yolk concentration in the animal pole is relatively low. The zygote nucleus is frequently displaced toward the animal pole. In general, yolk inhibits cleavage.
At one extreme are the eggs of sea urchins, mammals, and snails. These eggs have sparse, equally spaced yolk and are thus isolecithal (Greek, “equal yolk”).
In these species, cleavage is holoblastic (Greek holos, “complete”). meaning that the cleavage furrow extends through the entire egg.
These embryos must have some other way of obtaining food. Most will generate a voracious larval form, while mammals get their nutrition from the placenta.
At the other extreme are the eggs of insects, fishes, reptiles, and birds. Most of their cell volumes are made up of yolk.
The yolk must be sufficient to nourish these animals. Zygotes containing large accumulations of yolk undergo meroblastic cleavage, wherein only a portion of the cytoplasm is cleaved.
The cleavage furrow does not penetrate into the yolky portion of the cytoplasm. The eggs of insects have their yolk in the center (i.e., they are centrolecithal), and the divisions of the cytoplasm occur only in the rim of cytoplasm around the periphery of the cell (i.e., superficial cleavage).
The eggs of birds and fishes have only one small area of the egg that is free of yolk (telolecithal eggs), and therefore, the cell divisions occur only in this small disc of cytoplasm, giving rise to the discoidal pattern of cleavage.
These are general rules, however, and closely related species can evolve different patterns of cleavage in a different environment.
However, the yolk is just one factor influencing a species' pattern of cleavage. There are also inherited patterns of cell division that are superimposed upon the constraints of the yolk.
This can readily be seen in isolecithal eggs, in which very little yolk is present. In the absence of a large concentration of yolk, four major cleavage types can be observed: radial holoblastic, spiral holoblastic, bilateral holoblastic, and rotational holoblastic cleavage. We will see examples of these cleavage patterns below when we take a more detailed look at the early development of four different invertebrate groups.
Different organisms undergo cleavage in different ways .The pattern of embryonic cleavage is determined by two major parameters ;
1) by the amount and distribution of yolk within the cytoplasm and
2) by the organization of the egg.
1. Role of yolk : The amount and distribution of yolk determines where cleavage can occur and also the relative size of the blastomeres . Yolk is a factor that controls the rate and pattern of cleavage . Yolk is concentrated toward the vegetal pole, while its concentration is relatively low towards the.animal pole .The zygote nucleus , therefore ,is frequently displaced towards the animal pole . Thus , when one pole of the egg is relatively yolk free, cleavage occurs at that pole at a faster rate than at the opposite pole . Generally , yolk inhibits cytokinesis , but it does not suppress mitotic division of the nucleus .
Depending upon the amount and distribution of yolk the following classificatory types of cleavage are seen :
a) Holoblastic cleavage (Greek : holos , complete) : When the amount of yolk is less , the cleavage furrow extends through the entire egg and completely divides it. This type of cleavage is called holoblastic cleavage and is generally of two types .
i) Equal holoblastic cleavage : In microlecithal and isolecithal (homolecithal) eggs the first few
holoblastic cleavages produce blastomeres of equal size and are thus referred to as equal holoblastic cleavage . examples: Amphioxus , marsupials and placental mammals .
ii) Unequal holoblastic cleavage : In mesolecithal and telolecithal eggs the cleavage furrows generally divide the egg into unequal sized blastomeres. The blastomeres towards the animal pole are smaller containing less amount of yolk and are called micromeres , while those towards the vegetal pole are larger, yolk laden and are called macromeres . Examples: Lower fishes and amphibians .
b) Meroblastic cleavage : Zygotes containing large accumulation of yolk undergo meroblastic or incomplete cleavage , where only a portion of the cytoplasm is divided . The small amount of active cytoplasm near the animal pole or on the periphery of the egg divides completely , while most of the yolky portion at the vegetal pole or central area of egg remains undivided. Depending on the distribution of yolk may be mainly of the following two types of meroblastic cleavage.
i) Discoidal : In case of macrolecithal and highly telolecithal eggs , the cleavage furrow remains restricted to the disc shaped active cytoplasm of animal pole and is called discoidal meroblastic cleavage . Examples: Elasmobranchs, bony fishes,reptiles , birds and monotreme mammals.
ii) Superficial : The centrolecithal eggs of insect have their yolk at the centre and the cleavage furrow occurs only at the rim of cytoplasm around the periphery of the cells . This type of cleavage is called superficial meroblastic cleavage . Example: Arthropods – particularly insects.
2. Organization of the egg : There are certain inherited patterns of cell division that are superimposed upon the yolk. These factors in the egg cytoplasm influences the angle of the mitotic spindle and the timing of its formation . This can be seen in isolecithal eggs, where the amount of yolk is very less. The following four major cleavage types can be observed – radial holoblastic, spiral holoblastic , bilateral holoblastic and rotational holoblastic cleavage.
i) Radial holoblastic cleavage : If each of the upper tier blastomeres lie over the corresponding blastomeres of the lower tier ,then the pattern of the blastomeres is radially symmetrical . This is called radial holoblastic cleavage and is seen in case of Echinoderms and Amphioxus .
ii) Spiral holoblastic cleavage : When all the blsatomeres of the upper tier are shifted in such a way , so that they come to lie not over the corresponding blastomeres at the vegetal pole , but over the junction between any two of the vegetal blastomeres . This arrangement arises not due to secondary shifting of the blastomeres , but because of oblique position of the mitotic spindles . Thus , right from the start the cleavage furrows are arranged in a sort of spiral . This type of cleavage is called spiral holoblastic cleavage and it is seen in annelids , molluscs and flat worms .
iii) Bilateral holoblastic cleavage : In some animals such as Tunicates cleavage initially is approximately of radial type . The second cleavage plane is slightly displaced towards the posterior side of the animal vegetal axis. It results in two large anterior blastomeres and two smaller posterior blastomeres . Thus , subsequent cleavage becomes symmetrical only at the median plane and so is described as bilateral holoblastic cleavage.
iv) Rotational holoblastic cleavage : This type of cleavage is seen in higher mammals where the first cleavage plane divides the egg into a larger and a smaller blastomere. The larger blastomere then begins to divide at a faster rate than the smaller one and this is maintained throughout resulting in rotational holoblastic cleavage . Although there is practically very less amount of yolk , the blastomere which results right from the first cleavage are unequal in size .
Probably some conditioning effect must be present in the eggs cytoplasm which determines the size of the blastomeres and the rate of later cleavages .
