Page 20 - Academic Press Encyclopedia of Physical Science and Technology 3rd Molecular Biology

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Encyclopedia of Physical Science and Technology EN002G-104 May 17, 2001 20:53

812 Chromatin Structure and Modification

i.e., microscopically distinct compartments that appeared matin. The application of biophysical techniques in the
to represent chromosomes that condensed to distinct early 1970s led to significant progress in this field,
degrees: heterochromatic domains in chromosomes were and structural understanding improved accordingly, as a
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thus correctly deduced to contain silenced genes. 7 A X-ray crystal structure of the elementary subunit of
The complementary correlation between chromosome chromatin—the “nucleosome core particle” (146 bp of
decondensation and gene activation was provided by stud- DNA wrapped around the an octamer of core histones)—
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ies in insects, whose salivary glands contain giant chro- was described in 1984, and a 3.1 A resolution structure—in
mosomes that yield easily to microscopic examination. A 1991. Finally, in 1997, T. J. Richmond and colleagues pub-
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terminally differentiated tissue destined for destruction in lished a 2.8 A nucleosome core particle structure—thus,
the metamorphosis from a larva to the adult insect, the sali- we now understand the elementary composition and struc-
vary gland effects a very interesting solution to a demand ture of chromatin in very considerable detail. The next
for high protein production: it replicates its DNA without section describes the histone proteins, genetic evidence
intervening mitosis, and all the resulting DNA fibers for for their role in regulating transcription, work in the 1970s
each chromosome coalesce—side by side—into a macro- that led to the nucleosome hypothesis, and concludes with
scopic body termed “the polytene chromosome.” Their a description of the structure of the nucleosome.
size and certain other structural features have made them
invaluable model systems in biology. Most importantly,
cytologic analysis of changes in chromosome structure III. CHROMATIN STRUCTURE: THE
during larval development have revealed that at defined HISTONES AND THE NUCLEOSOME
timepoints, specific stretches of these giant chromosomes
decondense and form “puffs”—localized swellings. It was A. The Histones
correctly deduced quite early on that such decondensation
is related to the activation of genes that reside in those As discovered by Kossel early this century, the primary
stretches of the chromosome. protein residents of the nucleus are small highly basic pro-
In 1960, H. Clever and U. Karlsson combined their teinscalled“thehistones”(thereareapproximately36mil-
efforts of trying to determine, why the insect molting hor- lion nucleosomes, each containing nine histone molecules
mone, the steroid ecdysone, causes such dramatic mor- and 180 bp of DNA, in the nucleus of a human cell). Only
phological changes during insect development. The in- five different histones are sufficient to assemble chro-
jection of purified ecdysone into larvae of the midge matin: four core histones (H2A, H2B, H3, and H4; two
Chironomus lead to a premature and dramatic puffing of each are present in each nucleosome) and one linker
of specific stretches of polytene chromosomes: thus, hor- histone (H5; one per nucleosome). A technological Atlas
mone action was for the first time connected to regulation for molecular biology, gel electrophoresis through such
of gene activity and, importantly, to a localized alteration matrices as polyacrylamide and agarose has been an in-
(decondensation) of chromosome structure. A molecular valuable tool in studying the genome, and Fig. 1 shows a
mechanism, or correlate, of this striking phenomenon was denaturing (i.e., performed in the presence of detergent,
not obtained until studies in 1984 by K. Zaret and K. such as sodium dodecyl sulphate, SDS) polyacrylamide
Yamamoto, and subsequent experiments by T. Archer and gel on which the histones are resolved.
G. Hager (Section IV.A). The histones’ primary amino acid sequence offer sev-
Finally, 1964 saw the publication of a discovery whose eral glimpses into their function: these proteins are small
impact resonated only in 1996, but quite emphatically: (between 102 and 130 amino acids long) and very rich in
the observation by V. Allfrey, and A. E. Mirsky that hi- lysine or arginine. For example, of the 103 amino acids in
stone proteins are subjected to postranslational covalent human histone H4, 11 are lysine and 14 are arginine. This
modification via the acetylation and methylation of ly- has an immediate electrostatic implication for histone be-
sine residues in their NH 2 -terminal tails (Sections IV.A haviour in vivo: the pK a values for these amino acids’ side
and IV.C). Because the modifications reduce the positive chains (10.0 and 12.0, respectively) indicate that at phys-
charge of the histones (and thus have the potential to al- iological pH, the average histone H4 molecule carries ca.
ter the way histones interact with DNA), it was immedi- 24.99 positive charges on itself.
ately suspected they might have regulatory consequences. An interesting and informative aspect of histone biol-
Conclusive evidence to that effect was obtained in 1998 ogy is the extraordinary degree of sequence conservation
(Section IV.C). between these proteins across taxa: histone H4 in humans
Inherent technical limitations of cytological and bio- and in tomato (Lycopersicon esculentum) is identical in
chemical methods described could not illuminate the length and sequence with the exception of three highly
molecular structure of protein–DNA contacts within chro- conservative substitutions (i.e., Val 61 Ile). Thus, there