Immunofluorescence staining with antibodies against acetylated histone H4 and 5-methylcytosine was carried out to investigate female chromatin remodelling throughout oocyte maturation and chromatin rearrangement involving both male and female genomes after fertilization. Oocyte cytoplasm remodels female chromatin in preparation of the fertilizing event and the subsequent chromatin rearrangement. Histone H4 are in fact progressively deacetylated whereas demethylating enzymes do not seem to be active over this period. The acetylase/deacetylase balance seems to be cell cycle dependent as female chromatin is deacetylated during maturation and reacetylated at telophase II stage both after fertilization and activation. On the contrary, DNA demethylation seems to be strictly selective. It is in fact confined to the remodelling of paternal genome after fertilization of mature oocytes as the ooplasm is not effective in demethylating either paternal chromatin in GVBD fertilized oocytes or maternal genome of partenogenetically activated oocytes. Surprisingly we induced maternal chromatin demethylation after fertilization by treating oocytes with a combination of a methyltransferase inhibitor, 5-Azacytidine (5-AzaC), and a reversible and specific inhibitor of histone deacetylase, Trichostatin A (TSA). This treatment likely induces a hyperacetylation of histones (thus favouring the access to demethylating enzymes by opening female chromatin structure) associated with a block of reparative methylation by inhibiting methytransferases. This manipulation of chromatin remodelling may have applications regarding the biological significance of aberrant DNA methylation.[...]
MATERNAL CHROMATIN REMODELLING DURING MATURATION AND AFTER FERTILIZATION IN MOUSE OOCYTES
MATTIOLI, Mauro
2004-01-01
Abstract
Immunofluorescence staining with antibodies against acetylated histone H4 and 5-methylcytosine was carried out to investigate female chromatin remodelling throughout oocyte maturation and chromatin rearrangement involving both male and female genomes after fertilization. Oocyte cytoplasm remodels female chromatin in preparation of the fertilizing event and the subsequent chromatin rearrangement. Histone H4 are in fact progressively deacetylated whereas demethylating enzymes do not seem to be active over this period. The acetylase/deacetylase balance seems to be cell cycle dependent as female chromatin is deacetylated during maturation and reacetylated at telophase II stage both after fertilization and activation. On the contrary, DNA demethylation seems to be strictly selective. It is in fact confined to the remodelling of paternal genome after fertilization of mature oocytes as the ooplasm is not effective in demethylating either paternal chromatin in GVBD fertilized oocytes or maternal genome of partenogenetically activated oocytes. Surprisingly we induced maternal chromatin demethylation after fertilization by treating oocytes with a combination of a methyltransferase inhibitor, 5-Azacytidine (5-AzaC), and a reversible and specific inhibitor of histone deacetylase, Trichostatin A (TSA). This treatment likely induces a hyperacetylation of histones (thus favouring the access to demethylating enzymes by opening female chromatin structure) associated with a block of reparative methylation by inhibiting methytransferases. This manipulation of chromatin remodelling may have applications regarding the biological significance of aberrant DNA methylation.[...]I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
