Transcriptionally repressive state

A system such as this can provide a powerful mechanism for transcriptional control. While bound to nucleosomes, PARP-1 promotes a transcriptionally repressed state, but one that is simultaneously poised for activation because of the allosteric activating effect of nucleosomes on PARP-1 enzymatic activity. In the presence of NAD+, PARP-1 can autoPARylate and release from nucleosomes, shifting the chromatin to a more transcriptionally active conformation. PARC can reset the system by cleaving the PAR chains from PARP-1, allowing PARP-1 to re-bind the nucleosomes and re-establish a transcriptionally repressed state. 

There are, however, a number of alternative explanations for these results and the following conclusions that were drawn (1) a transcriptionally repressive state develops by the two-cell stage and (2) the role of the enhancer is to relieve this inhibition. For example, the plasmid could be lost during the course of the first mitosis and hence not available for transcription in the newly formed two-cell embryos different promoter elements may be used by the cleavage-arrested one-cell embryos and the two-cell embryos the transcriptional capacities may differ in the cleavage-... 

Figure 4. Demonstration of the development of a transcriptionally repressive state by nuclear transplantation. In the first experimental protocol, the injected nucleus was transplanted to an S-phase-arrested, one-cell embryo that was then analyzed at a time that corresponded to the mid-late two-cell stage. In the second protocol, the injected nucleus was transplanted to an S-phase-arrested, two-cell embryo that was analyzed at a time that corresponded to the four-cell stage, whereas in the third protocol the injected nucleus was transplanted to a two-cell blastomere that divided. In the last protocol, the injected two-cell blastomere in G2 was transplanted to an S-phase-arrested, one-cell embryo that was then at a time that corresponded to the mid-late two-cell stage. The data are expressed relative to the amount of activity observed for the tk promoter-containing reporter gene in the S-phase-arrested one-cell embryos, and were taken from Henery et al., 1995.

In toto, these results suggest that a transcriptionally repressive state develops following ZGA (see sections VII and VIII for further discussion) and that the primary function of an enhancer is to relieve this repression, in particular for genes with weak promoters,... 

The developmental acquisition of a requirement for an enhancer for efficient reporter gene expression suggests that a transcriptionally repressive state develops during the one- to two-cell transition. This repression, as manifested by a requirement for the enhancer, was not observed in aphidicolin-treated, cleavage-arrested embryos in which the plasmid was injected into the male pronucleus (Figure 3). The extent of BrUTP incorporation by S-phase-arrested, one-cell embryos was also consistent with the development of a transcriptionally repressive state (Aoki et al., 1997) (Figure 5). The total amount of incorporation of BrUTP by the two pronuclei of such S-phase-arrested embryos was about four times that of a two-cell blasto-mere in G2 when expressed on a per chromosome basis. 

Figure 5. Role of DNA replication in the development of a transcriptionally repressive state. BrUTP incorporation of membrane-permeabilized embryos was conducted on two-cell embryos in G2 (2C) or S-phase-arrested, one-cell embryos that were analyzed at a time that corresponded to G2 of the two-cell stage. Both the male (aph M) and female (aph F) were analyzed. Transcription was also analyzed at a time that corresponded to C2 of the two-cell stage for S-phase-arrested, two-cell embryos (aphidicolin added after the first round of replication aph 2C) or in two-cell embryos treated with trapoxin (trap). The data are expressed relative to the amount of BrUTP incorporated by a two-cell blastomere nucleus in C2. (Data taken from Aoki et al., 1997).

The expression of a somatic form of histone HI could constitute one component of the transcriptionally repressive state. The biogenesis of a mature nucleosome is associated with the formation of transcriptionally repressive chromatin. While the association of DNA with histones H3 and H4 can generate a nucleosomelike structure, the DNA can still readily interact with transcription factors. This ability is diminished with the further incorporation of histones H2A and H2B that results in the formation of a nucleosome that can exclude the binding of basic components of the transcription machinery (e.g., RNA polymerase II). The formation of a truly transcriptionally repressive chromatin structure is brought about by the addition of histone HI, which can be incorporated only after the addition of histones H2A and H2B and causes chromatin to condense into a 30-nm fiber (Bouvet et al., 1994). The requirement for an enhancer for efficient promoter utilization in zygotic nuclei, but not the pronuclei, could reflect the developmental acquisition of chromatin con-... 

In summary, the reciprocal decrease in histone acetylation coupled with an increase in histone HI expression could, in principle, promote the conversion of transcriptionally permissive chromatin into a transcriptionally repressed form and hence serve as the foundation for the development of the transcriptionally repressed state that is first observed at the two-cell stage. The second round of DNA replication may also contribute to the development of this state by displacing transcriptionally productive complexes and thus providing an opportunity for the assembly of a mature, transcriptionally repressive chromatin structure. 

The wide spectrum of biological activities exhibited by the sirtuins linking transcriptional repression and activation, metabolic state and stress response strongly... 

Fig. 1.40. Model of repression and activation of transcription. The figure illustrates various mechanisms of repression of transcription, a) genes are in a generally repressed states in inactive chromatin. In a first phase of activation the chromatin is restrnctured. b) The promoter is now accessible for the binding of the basal transcription factors and for RNA polymerase II. c) An initiation complex is formed that contains the central components of the transcription apparatns, bnt which enables transcription only at a low rate, d) the binding of repressors to the transcription initiation complex can prevent fnrther activation of transcription at this step, e) the binding of transcription activators to their DNA elements leads to activation of transcription, f) an active repression is affected by proteins that bind seqnence specifically to DNA elements and in their DNA-bound form inhibit the transcritption preventing interactions with the transcription apparatus.

The amino terminal tads of the four core histones contain lysines that are acetylated by HATs and deacetylated by HDACs. The histone octamer (H2A, H2B, H3, H4)2 is represented as a cylinder wrapped by DNA. It is thought that neutrahzation of the positive charges on the histone tads results in alterations of the nucleosome structure that may lead to a higher mobdity of the nucleosome and/or an improved accessibdity of the bound DNA, with accompanying changes in chromatin structure, chromatin hierarchy and transcription. In most, but not ad cases, deacetylation correlates with the repressed state and acetylation correlates with the transcriptionady active state. 

In the case of the T3R and RAR specific repressor proteins have been identified which bind to the receptor and mediate an inhibition of transcription (Hoerlein et al, 1995). The repressor proteins are also termed co-repressors Among the repressors are found proteins with histone deacetylase activity (see 1.4.6). The RXR heterodimers possibly stabilize the repressed state of chromatin by recruiting a histone deacetylase to the chromatin. 

Despite this elaborate binding complex, repression is not absolute. Binding of the Lac repressor reduces the rate of transcription initiation by a factor of 10s. If the 02 and 03 sites are eliminated by deletion or mutation, the binding of repressor to CL alone reduces transcription by a factor of about 102. Even in the repressed state, each cell has a few molecules of /3-galactosidase and galactoside permease, presumably synthesized on the rare occasions when the repressor transiently dissociates from the operators. This basal level of transcription is essential to operon regulation. 

Fig. 1.32 Pathways of repression of transcription. The figure illustrates various mechanisms of repression of transcription, a) Repressors can induce a generally repressed state in chromatin which is incompatible with transcription. To allow transcription at all, the repressed state must be relieved, b) Repressors can target the transcription complex...

Fig. 4.11 Model of repression and activation of T3R. In the absence of the T3 hormone, a hetero-dimeric RXR-T3R receptor is bound at the T3-re-sponsive element,TRE, establishing a basal repressed state. The repressed state is maintained by recruitment of corepressor complexes containing histone deacetylase activity. X refers to potential unidentified cofactors (possibly chromatin remodeling complexes or SRBs) which help to keep the promotor-bound basal transcription apparatus in...

The activity of cyclin E-CDK2 complexes is mainly directed toward two substrates, the pRb protein and the inhibitor p27KIP1. Phosphorylation of pRb inactivates pRb and releases the transcription factor E2F from a repressed state, allowing transcription of key proteins for Gj/S transition. The gene for cyclin E is also induced by transcription... 

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