Poly(ADP-ribose)-dependent regulation of DNA repair by the chromatin remodeling enzyme ALC1.

Posttranslational modifications play key roles in regulating chromatin plasticity. Although various chromatin-remodeling enzymes have been described that respond to specific histone modifications, little is known about the role of poly[adenosine 5'-diphosphate (ADP)-ribose] in chromatin remodeling. Here, we identify a chromatin-remodeling enzyme, ALC1 (Amplified in Liver Cancer 1, also known as CHD1L), that interacts with poly(ADP-ribose) and catalyzes PARP1-stimulated nucleosome sliding. Our results define ALC1 as a DNA damage-response protein whose role in this process is sustained by its association with known DNA repair factors and its rapid poly(ADP-ribose)-dependent recruitment to DNA damage sites. Furthermore, we show that depletion or overexpression of ALC1 results in sensitivity to DNA-damaging agents. Collectively, these results provide new insights into the mechanisms by which poly(ADP-ribose) regulates DNA repair.

Nucleosome dynamics.

In the 30 years since the discovery of the nucleosome, our picture of it has come into sharp focus. The recent high-resolution structures have provided a wealth of insight into the function of the nucleosome, but they are inherently static. Our current knowledge of how nucleosomes can be reconfigured dynamically is at a much earlier stage. Here, recent advances in the understanding of chromatin structure and dynamics are highlighted. The ways in which different modes of nucleosome reconfiguration are likely to influence each other are discussed, and some of the factors likely to regulate the dynamic properties of nucleosomes are considered.

p21WAF1/CIP1 regulates the p300 sumoylation motif CRD1 through a C-terminal domain independently of cyclin/CDK binding.

Although best known for its ability to inhibit Cyclin/Cdk complexes and the replication protein PCNA, p21(WAF1/CIP1) is a multifunctional protein that interacts with many cellular binding partners, including a number of transcriptional regulators. Previously, we characterized p21 derepression of the p300 sumoylation-dependent transcriptional repression domain, CRD1. Such repression domains are at least partially dependent upon recruitment of histone deacetylase (HDAC) complexes but the mechanism through which p21 selectively disrupts CRD1 activity remains unknown. Here, we demonstrate that distinct motifs in the C-terminus of p21 are required for regulation of p300 CRD1 function and that this effect does not correlate with Cyclin or PCNA binding. Through the creation of N-terminal glutathione-s-transferase fusion proteins, which also overcome the problems of instability that result from many p21 mutations, we investigated p21 binding to HDACs. Although p21 binds both Class I and Class II HDACs in vitro, only weak association with HDAC1 and 2 is seen in cells. Mutation of the p21 PCNA binding domain significantly increases this interaction suggesting that binding is mutually exclusive and only naturally occurs under certain conditions. Binding of HDACs also failed to correlate with CRD1 inducibility, suggesting that p21 targets other transcriptional repression complexes to mediate this effect.

Induction of transcription through the p300 CRD1 motif by p21WAF1/CIP1 is core promoter specific and cyclin dependent kinase independent.

The tumor suppressors p300 and CREB-binding protein (CBP) are both multifunctional transcriptional coactivators. We have previously found that the cyclin dependent kinase (CDK) inhibitor p21(WAF1/CIP1) can stimulate transactivation by p300 and CBP through inhibiting transcriptional repression by a discrete domain within these proteins termed CRD1. Given the large number of p300/CBP associated functions, it is unlikely that p21 regulates the expression of every gene under their control, however. Here we have investigated the factors that help determine this specificity. We have discovered that while CRD1 can repress the activity of p300 at multiple promoters, induction of transcription by p21 though this motif is highly variable. Analysis of this effect revealed that p21 inducibility is determined by sequences flanking the TATA box. Significantly, p21 regulation of CRD1 domain function is independent of Cyclin /CDK inhibition suggesting a novel function of this protein. p21 does not interact directly with the CRD1 motif, however. These results give further insight into how regulators of cell growth and tumorigenesis, such as p21, can specifically target and induce the expression of select groups of genes.