Two-ended recombination at a Flp-nickase-broken replication fork.

Replication fork collision with a DNA nick can generate a one-ended break, fostering genomic instability. The opposing fork's collision with the nick could form a second DNA end, enabling conservative repair by homologous recombination (HR). To study mechanisms of nickase-induced HR, we developed the Flp recombinase "step arrest" nickase in mammalian cells.

Two-ended recombination at a Flp-nickase-broken replication fork.

Collision of a replication fork with a DNA nick is thought to generate a one-ended break, fostering genomic instability. Collision of the opposing converging fork with the nick could, in principle, form a second DNA end, enabling conservative repair by homologous recombination (HR). To study mechanisms of nickase-induced HR, we developed the Flp recombinase "step arrest" nickase in mammalian cells.

BRCA1-IRIS promotes human tumor progression through PTEN blockade and HIF-1α activation.

is an established breast and ovarian tumor suppressor gene that encodes multiple protein products whose individual contributions to human cancer suppression are poorly understood. BRCA1-IRIS (also known as "IRIS"), an alternatively spliced product and a chromatin-bound replication and transcription regulator, is overexpressed in various primary human cancers, including breast cancer, lung cancer, acute myeloid leukemia, and certain other carcinomas. Its naturally occurring overexpression can promote the metastasis of patient-derived xenograft (PDX) cells and other human cancer cells in mouse models.

BRCA1/FANCD2/BRG1-Driven DNA Repair Stabilizes the Differentiation State of Human Mammary Epithelial Cells.

An abnormal differentiation state is common in BRCA1-deficient mammary epithelial cells, but the underlying mechanism is unclear. Here, we report a convergence between DNA repair and normal, cultured human mammary epithelial (HME) cell differentiation. Surprisingly, depleting BRCA1 or FANCD2 (Fanconi anemia [FA] proteins) or BRG1, a mSWI/SNF subunit, caused HME cells to undergo spontaneous epithelial-to-mesenchymal transition (EMT) and aberrant differentiation.

An acetylation switch in p53 mediates holo-TFIID recruitment.

Posttranslational modifications mediate important regulatory functions in biology. The acetylation of the p53 transcription factor, for example, promotes transcriptional activation of target genes including p21. Here we show that the acetylation of two lysine residues in p53 promotes recruitment of the TFIID subunit TAF1 to the p21 promoter through its bromodomains.

Mechanistic insights into maintenance of high p53 acetylation by PTEN.

Earlier studies have shown that PTEN regulated p53 protein stability both in a phosphatase-dependent manner through antagonizing Akt-Mdm2 pathway and in a phosphatase-independent manner through interacting with p53. In this study, we report that PTEN forms a complex with p300 in the nucleus and plays a role in maintenance of high p53 acetylation in response to DNA damage. Furthermore, p300 is required for nuclear PTEN-regulated cell cycle arrest.

Purification of acetyl-p53 using p300 co-infection and the baculovirus expression system.

As cells persist in their environment, they are exposed to harmful agents that can damage their genomic DNA. When DNA becomes damaged, p53, a tumor suppressor, is stabilized and acts as a transcription factor to cause either cell cycle arrest or apoptosis. Strict p53 regulatory mechanisms have been well characterized relative to phosphorylation and dephosphorylation, but acetylation of p53 in response to DNA damage has also been shown to participate in p53 function.

Insights into selective activation of p53 DNA binding by c-Abl.

As a transcription factor, p53 recognizes a specific consensus DNA sequence and activates the expression of the target genes involved in either growth arrest or apoptosis. Despite our wealth of knowledge on the genes that are targeted by p53 in growth arrest and apoptosis, relatively little is known about the promoter specificity triggered by p53 in these processes. Here we show that interaction with c-Abl stabilized p53 tetrameric conformation, and as a consequence c-Abl stimulated p53 DNA binding only when all quarter binding sites (a perfect binding sequence) on p53-responsive promoters were present.

Phosphorylation on Thr-55 by TAF1 mediates degradation of p53: a role for TAF1 in cell G1 progression.

The largest subunit of TFIID, TAF1, possesses an intrinsic protein kinase activity and is important for cell G1 progression and apoptosis. Since p53 functions by inducing cell G1 arrest and apoptosis, we investigated the link between TAF1 and p53. We found that TAF1 induces G1 progression in a p53-dependent manner.

PTEN tumor suppressor regulates p53 protein levels and activity through phosphatase-dependent and -independent mechanisms.

We show in this study that PTEN regulates p53 protein levels and transcriptional activity through both phosphatase-dependent and -independent mechanisms. The onset of tumor development in p53(+/-);Pten(+/-) mice is similar to p53(-/-) animals, and p53 protein levels are dramatically reduced in Pten(-/-) cells and tissues. Reintroducing wild-type or phosphatase-dead PTEN mutants leads to a significant increase in p53 stability.