Lauren Butz

9 October 2003

Cell Biology, Swann

Chromatin: Structure and Function

Chromatin is found inside the nucleus of a cell, and is composed of proteins and DNA. DNA makes up 40 percent of chromatin, while proteins make up the remaining 60 percent (Raven & Johnson, 1999). Two types of chromatin, heterochromatin and euchromatin, can be observed using electron or light microscopy based on the degree of chromosomal condensation in the nucleus. Heterochromatin is highly condensed chromatin, and is inactive in DNA transcription, whereas euchromatin is much less coiled and is the site of DNA transcription. The mass of DNA that makes up chromatin contains 8 x 10⁹ base pairs in a human cell and would stretch to approximately three meters long if removed from the nucleus (Mathews, van Holde, & Ahern, 1999). Because each human chromosome contains 140 million nucleotides, it is necessary that the components of the chromosomes be highly condensed to fit inside the nucleus. The manner in which all this material fits inside the nucleus of a cell occurs through chromatin packaging.

The basic building block of chromatin is double helix DNA wrapped around an octamer histone complex, called a nucleosome. Nucleosomes were first described by Ada and Donald Olins, who initially called the nucleosomes “ν– bodies” (Klug & Cummings, 2002). A nucleosome is composed of a core of eight histone molecules, two each of the histones H2A, H2B, H3, and H4, and one histone of H1 or H5, a central strand of DNA of 147 base pairs, and linker DNA (Widom, 1998). Histone structure and function has been studied using electron and atomic force microscopy, cryoelectron microscopy, and neutron scattering (Widom, 1998). Individual histones are small polypeptides, composed primarily of the amino acids arginine and lysine, which give each histone a positive charge. H1 is lysine-rich, H2A and H2B are slightly lysine-rich, and H3 and H4 are arginine-rich (Klug & Cummings, 2002). The four histone types associate to form an octamer histone to which the DNA binds to create the completed nucleosome (Figure 1).

Figure 1. Schematic representation of a nucleosome, containing the central DNA, link DNA, and a core of four types of histones— two each of H2A, H2B, H3, and H4 (Junqueira & Carneiro, 2003).

This core particle structure was studied by Timothy Richmond in 1997 using X-ray crystallography at 2.8Å, a great improvement over the 7Å resolution used in 1984. Observations show that the four histone types occur as two types of tetramers: two H2A/H2B tetramers and one H3-H4 tetramere subunit on each histone complex (Klug & Cummings, 2002). The four histone groups, H2A, H2B, H3, and H4, are connected by a histone fold (Widom, 1998). There is an interlocking protein fold that occurs between the two H3-H4 heterodimers, similar in structure to that which occurs between the H2A-H2B heterodimers (Widom, 1998). This common histone fold lends evidence that each histone protein has a common ancestor.

The central histone core has 121 base pairs attached to it, with two tails each of 10-13 base pairs projecting from either end. Each individual histone dimer organizes approximately 27- 28 base pairs, with four base pairs between each dimer (Widom, 1998). The amino acid tail of each histone interacts with the tails of other histones and the DNA that binds to the histone complex, such that the tails bracket turns of the DNA on the core histones (Widom, 1998). In vitro studies have shown that nucleosomes are dynamic and have the ability to move, or “slide,” on the DNA double helix (Messerman, Pennings, & Bradbury, 1992). There are millions of such histone complexes that compose each strand of chromatin, each of which is connected to the subsequent nucleosome with the “linker” DNA. The structure of H1, which was determined using nuclear magnetic resonance, and the location, determined though neutron scattering, shows that it is an 80 amino acid globular domain (Widom, 1998). Each H1 histone has a long C-terminus and a shorter N-terminal domain.

Because the DNA double helix is so long, it must be tightly coiled to fit inside the nucleus. The first level of packaging involves the wrapping of 147 base pairs of DNA around the nucleosome, thus reducing the physical length of the DNA sevenfold (Murray & Hunt, 1993). To do this, the DNA double helix coils a 1.7 left-handed turn around the nucleosome and links to subsequent nucleosomes by the linker DNA (Mathews, van Holde, & Ahern, 1999). The net positive charge of the nucleosome, given by the amino acids arginine and lysine, attracts the negatively charged phosphate groups of the DNA, an important characteristic for the formation of chromatin (Rave & Johnson, 1999). The phosphate groups on the DNA bind to the nucleosomes with salt bridges and hydrogen bonds (Widom, 1998). The wrapping and bending of the DNA around the histone complex is not uniform throughout the chromatin strand.

In the second level of chromatin packaging, the individual strands of chromatin are coiled on an axis into a solenoid shape, which is 30nm in diameter and is composed of millions of nucleosomes (Figure 2).

Figure 2. Levels of chromatin packaging, ranging from the 2nm DNA double helix, 11nm nucleosome, 30nm chromatin fibril, the 300nm and 700nm condensed chromatin, and 1400nm metaphase chromosome (Junqueira & Carneiro, 2003).

Through atomic free microscopy, it can be see that each turn of the chromatin strand consists of six nucleosome complexes (Widom, 1998). Serial thin sectioning electron microscopy shows that the 30nm chromatin fibrils condense further and organize into 60-80nm fibers, which then undergo helical coiling to produce 100-130nm strands, called chromatids (Widom, 1998).

Chromatin structure is directly correlated to its function. The process of cell division relies on the initial condensation of chromatin, which protects the DNA during cell division. Each structural component of chromatin, the DNA, histones, and linker DNA, is important in chromatin function.

It is undetermined whether the H1 histone is essential to chromatin function, though as the following summarized experiments show, H1 does influence chromatin functions. The H1 histone plays a regulatory role in the entry and exit angles of the linker DNA. Nuclear magnetic resonance shows that the H1 histone binds to the DNA at two points (Draves, Lowary, & Widom, 1992). Experiments have also shown that the entry and exit angles of the DNA occur closer together on the nucleosome when H1 is present than when H1 is absent (Widom, 1998). H1 is also important in the histone folding mechanism that occurs in each nucleosome. In nucleosomes in which the trypsin-sensitive tail domains and H1 histone have been removed, the linker DNA does not bend correctly and the histones do not undergo normal cation-dependent folding (Garcia-Ramirez, Dong, & Ausio 1992). It is also possible that histone H1 functions as an initiator of the condensation of chromatin. During chromatin condensation of early prophase, histone H1 is phosphorylated on serine and threonine (Murray & Hunt, 1993). It is currently unknown what the purpose of this phosphorylation is.

The 10-13 base pair linker DNA associated with each nucleosome functions as a connection between adjacent nucleosomes and influences chromatin fiber dynamics. The histone functions as a stabilizer of the DNA that connects adjacent nucleosomes and of the solenoid coil (Simpson, 1978). Without this stabilization, the nucleosomes would move uninhibited on the chromatin fiber. The sliding of the nucleosomes, in fact, is enhanced at higher temperatures and ionic concentrations, but does occur at lower temperature and ionic concentrations (Widom, 1998).

At present, on-going experimentation is examining the specific functions of the H1 histone and the linker DNA. Through further scientific exploration, it is hoped that the exact structure and function of all the components of chromatin will someday be known.

Sources

Draves, P.H., Lowry, P.T., & Widom, J. (1992). Cooperative binding of the globular domain of H5 to DNA. Journal of Molecular Biology, 255, 1105-1121.

Garcia-Ramirez, M., Dong, F., & Ausio, J. (1992). The role of histone tails in the folding of oligionucleosomes depleted of histone H1. Journal of Biological Chemistry, 267, 19587-19595.

Junqueira, L.C., & Carneiro, J. (2003). Basic histology: Text & atlas (10^th ed.). New York: McGraw-Hill.

Klug, W.S., & Cummings, M.R. (2002). Essentials of genetics (4^th ed.). Upper Saddle River, NJ: Prentice Hall.

Mathews, C.K., van Holde, K.E., & Ahern, K.G. (1999). Biochemistry (3^rd ed.). San Francisco: Benjamin/Cummings.

Messerman, G., Pennings, S., Bradbury, E.M. (1992). Mobile nucleosomes—A general behavior. EMOB J. 11:251-259.

Murray, A., & Hunt, T. (1993). The cell cycle: An introduction. New York: Oxford University Press.

Raven, P.H., & Johnson, G.B. (1999). Biology. Boston: WCB/McGraw-Hill.

Simpson, R.T. (1978). Structure of the chromatosome, a chromatin core particle containing 160 base pairs of DNA and all the histones. Biochemistry, 17, 5524- 5531.

Widom, J. (1998). Structure, dynamics, and function of chromatin in vitro. Annual Review of Biophysics and Bimolecular Structure, 27, 285-327.