The two
principal mechanisms that regulate gene expression are DNA methylation and
histone deacetylation (Fig.1 a). Among
the amino acid residues on the histone proteins, lysine and argentine are
relatively in good proportion and they are vulnerable for methylation and other
enzymatic modifications (Table.1). DNA
methylation in mammals plays a crucial role in their cell-cycles, developmental
stages, X-chromosome inactivation, telomere length adjustment, gene silencing,
aging, carcinogenesis and a few human genetic disorders. Histone acetylation is associated with transcriptional
activation because the affinity of acetylated histone protein for DNA is
reduced and
chromatin package is thus relaxed (Fig.1 b).
Furthermore, there is a positive correlation between DNA methylation and
gene inactivity.
Does DNA
methylation cause the loss of transcriptional activity of a specific region of
chromatin? The same question was
addressed experimentally in that a DNA – demethylating agent viz.,
5-azacytidine brought about the reactivation of silenced endogenous genes viz.,
ribosomal RNA genes (13). In a few
ingenious protocols designed by Buschhausen et al.,(14) it was shown that the injected methylated and
non- methylated versions of the Herpes simplex virus thymidine kinase (HSVTK)
gene into the rodent cells were reported to be active during the first
8-hours. Later, the inhibitory effects
of DNA methylation were noticed on the methylated version of HSVTK. With these observations, it was hypothesized
that initially microinjected versions of TK were not fully incorporated into
the chromatin in the first 8-hours and that at a later stage the methylated
version became silent upon incorporation into the chromatin. The time-dependent repression of methylated
versions of DNA fragments was reported upon injection of m ethylated DNA
fragments into Xenopus oocytes (15).
Histone methylation
results in various transcriptional consequences depending on which histone is
affected. For example, histone H3
methylation at lysine 9 is associated with heterochromatin, a more compact
version of chromatin and ultimately silencing of genes. Additionally, inhibition
of histone deacetylation results in the re-expression of methylated copy of the
hygromycine resistance gene in the fungus, Neurospora
crassa (16) suggesting that deacetylation may either directly or indirectly
cause loss of DNA methylation. Even though
myeloid specific mouse M-lysozyme gene is methylated and silenced in
non-myeloid cells, upon treating them (fibroblasts, T-lymphocytes, etc) with
trichostatin-A (a specific inhibitor of histone deacetylse), the expression of
the same is revived. The level of
revival of M-lysozyme gene was comparable with that achieved by the
demethylating agent viz., 5-azacytidine.
In yet another incident, it is shown that the silencing of Metallothionein-1
gene is induced due to the methylation of CpG islands in mouse lymphosarcoma
P1798 cells (17). Thus, there are
multi-subunit chromatin-protein complexes prone to anchor a variety of
epigenetic factors which collectively and preferentially work together for the
awakening of genes.
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