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History and Context

1.1 From Cells to Their Nuclei

Until the last decades of the previous century, the history of the cell cycle is nearly the same as the history of the chromosome theory of heredity. And before chromosomes, we have to go back to when cells were first seen and then recognized as the unit of life. By the early seventeenth century, advances in lens grinding in the Netherlands allowed several individuals in Europe to experiment with and construct microscopes. Robert Hooke first used the word "cell" to describe what he saw when looking with his microscope at plant tissue. School children today have seen Hooke's famous drawings of cork sections. The drawings resemble a honeycomb, and they first appeared in Hooke's Micrographia in 1665. Hooke did not recognize that what he saw were the skeletal remains of all life's basic units. That realization did not happen until much later. In the decades after Hooke, numerous microscopists made a series of observations of cells. In 1719, looking at red cells from fish (which do not lose their nuclei as ours do), Antonie van Leeuwenhoek probably made the first drawings of the nucleus (1). The term "nucleus" was introduced in 1831 by Robert Brown (also of "Brownian" motion fame) when he used it to describe cells from orchids (1).

The discovery of the nucleus was highly significant. Not only because the nucleus contains the chromosomes (which had not been discovered at the time), but also because it is a visible cellular landmark. A marker that scientists can look for and monitor. Cellular landmarks, morphological or molecular, have driven and continue to drive cell cycle research. But just because a landmark is there does not necessarily mean that it may also make certain things happen. An example is the nucleolus and its role in the generation of new cells. Rudolf Wagner clearly described the nucleolus within the nucleus in 1835, which he assumed to be the "germinative spot" for cell formation. The nucleolus has important roles, but not in "seeding" new cells.

1.1.1 The Cell Theory

Theodor Schwann and Matthias Schleiden are usually credited with the formulation of key tenets of the cell theory. That the cell is the fundamental unit of life, and that all plants and animals are made of cells, each cell having a nucleus and a nucleolus (2). But it was not clear to them how new cells were made. They favored the idea that intracellular or extracellular matter crystallizes somehow into new cells (1, 2). Schleiden thought that new nuclei form without any relationship to preexisting ones. In essence, the views of Schwann and Schleiden on how new cells are made fell squarely into the realm of the spontaneous generation of life from nonliving matter, which was a popular theory at the time. Until Louis Pasteur put an end to spontaneous generation with his unambiguous col de cygnet (swan neck flask) experiment in 1859.

Explicit descriptions of cell division and the concept of new cells arising from preexisting ones came from Robert Remak (1, 2). Remak reached these conclusions from observations in multiple contexts, from red blood cells of chicken embryos to frog eggs immediately after fertilization (1). The splitting of eggs after fertilization had been observed by others before. But Remak's histological manipulations allowed him to visualize the membrane of the egg, and follow the origin of the embryonic cells from the fertilized egg. The continuity of all the cells in the embryo from one fertilized ancestor was now a settled issue. This placed cell division and the cell cycle as the basis of how multicellular organisms develop. Remak's data fit nicely with Pasteur's that there is no spontaneous generation of life. The notion that new cells arise only from cell division was opposed by many, including Rudolf Virchow. But Virchow changed his tune later. Virchow's famous aphorism omnis cellula e cellula (all cells arise only from preexisting cells) encapsulated succinctly and popularized a key part of the cell theory, a pillar of modern biology. Together with the other pillar of modern biology, Darwin's theory of evolution, we arrive at a stunning and profound conclusion: Starting with a single cell, all life that ever was, is, and will be on this planet results from cell division. Spend a moment to reflect on this. If you had any doubt that the topics we will discuss in this book are important, now is the time to put those doubts to rest.

1.1.2 Mitosis

By the mid-nineteenth century, the cell theory was established, and microscopy was getting better. Several individuals (including Remak) had documented distinct stages of nuclear division, including nuclear elongation in some cases, and nuclear dissolution in others. Wilhelm Hofmeister noticed that the nuclei of plant cells dissolved, but some nuclear material remained, in what he thought were coagulates, which then segregated into the nuclei of daughter cells. Although Hofmeister could not realize the biological role of what he was seeing, his descriptions were remarkably close to the stages of mitosis we recognize today (1). Scientists kept looking and looking under the microscope. They also processed and stained the cells with various techniques and dyes -all searching for crisp landmarks. Walther Flemming experimented with basic dyes and fixatives. To this day, pathologists use similar methods to look at cells in tissues. Flemming found that a nuclear substance, which was presumably acidic and negatively charged, stained very strongly with basic, positively charged dyes. He used the term chromatin (from the Greek "colored") to describe that substance. Flemming used salamander cells, because they were big and had big nuclei. As always, choosing the right experimental system for a research objective can reap enormous rewards.

Imagine you have a simple microscope, and you have figured out how to stain cells, with some nuclear substance being intensely stained. Assuming that it takes several hours to days for typical proliferating animal cells to divide, what would you expect to see? For the most part, not much. Each cell would be very similar to others and to itself, from its birth until it divides. Shortly before division, Flemming noticed that the colored nuclear substance was organized into threadlike structures (the Greek word for a thread is µ?t??/mitos), which were then distributed into the daughter cells, in a process he called mitosis (Figure 1.1).

The nuclear threads/filaments were named chromosomes a few years later by Waldeyer. Unlike others that had also seen thread-like structures forming and segregating, Flemming was the first to discover their splitting during mitosis lengthways (1). Flemming imaged and documented the complete series of events during mitosis, a fundamental cellular process, in all its glory: first, chromosomes appear, becoming denser and more compact over time, while at the same time the nuclear membrane disappears in most animal and plant cells (prophase); second, the compact chromosomes reach an equilibrium position at an "equator" position (metaphase); third, the chromosomes split lengthways and move toward the opposite poles of the cell (anaphase); fourth, the chromosomes arrive to the poles and the daughter nuclei appear, with reconstituted nuclear membrane around them (telophase).

Figure 1.1 Illustration of mitotic cells.

Source: (3) Walther Flemming (1882), F.C.W. Vogel.

Figure 1.2 The duration of the interphase is much longer than the mitotic stages in the cell cycle. In most cells, interphase is devoid of any morphological landmarks.

Now we can divide the cell cycle into two phases. The relatively short phase of mitosis ("M" phase) starts when threads appear in the nucleus and ends when two nuclei appear (Figure 1.2). Mitosis is typically followed very quickly with cytokinesis when the cell's cytoplasm segregates to two daughter cells, each with a nucleus. The rest of the cell cycle, which in most cells lasts a lot longer than the M phase, is called the interphase (Figure 1.2). During interphase nothing much appears to be happening. The dramatic visual landmarks of mitosis are hard to miss and continue to guide research about when and how the nucleus divides. But it would be many decades later, in the middle of the twentieth century, when interphase landmarks were discovered. Until then, scientists followed what they had, the mysterious chromosomes.

1.1.3 The Chromosome Theory of Heredity

When the chromosomes were first seen, their role was unknown. That the chromosomes carry the genetic information would not be clearly formulated until 1903 from Walter Sutton, and at about the same time from Theodor Boveri. The chromosome theory of heredity would be proven beyond doubt in the first decades of the twentieth century. Mendel's laws, which by 1900 were rediscovered, were abstract and could apply to any substance that carried the genetic information. As we will discuss later, the chromosomes do behave according to Mendel's laws. But what was the evidence that led to the idea that chromosomes carry the genetic information?

It was becoming more apparent that the nucleus, and not the cytoplasm, carried the genetic information. The unambiguous pieces of...