No abstract

The three-dimensional organization of chromosomes within the nucleus and its dynamics during differentiation are largely unknown. To visualize this process in molecular detail, we generated high-resolution maps of genome-nuclear lamina interactions during subsequent differentiation of mouse embryonic stem cells via lineage-committed neural precursor cells into terminally differentiated astrocytes. This reveals that a basal chromosome architecture present in embryonic stem cells is cumulatively altered at hundreds of sites during lineage commitment and subsequent terminal differentiation. This remodeling involves both individual transcription units and multigene regions and affects many genes that determine cellular identity. Often, genes that move away from the lamina are concomitantly activated; many others, however, remain inactive yet become unlocked for activation in a next differentiation step. These results suggest that lamina-genome interactions are widely involved in the control of gene expression programs during lineage commitment and terminal differentiation.

Chromatin is important for the regulation of transcription and other functions, yet the diversity of chromatin composition and the distribution along chromosomes is still poorly characterized. By integrative analysis of genome-wide binding maps of 53 broadly selected chromatin components in Drosophila cells, we show that the genome is segmented into five principal chromatin types that are defined by unique, yet overlapping combinations of proteins, and form domains that can extend over >100 kb. We identify a repressive chromatin type that covers about half of the genome and lacks classic heterochromatin markers. Furthermore, transcriptionally active euchromatin consists of two types that differ in molecular organization and H3K36 methylation, and regulate distinct classes of genes. Finally, we provide evidence that the different chromatin types help to target DNA-binding factors to specific genomic regions. These results provide a global view of chromatin diversity and domain organization in a metazoan cell.

Transcriptional induction of cell-cycle regulatory proteins ensures proper timing of subsequent cell-cycle events. Here we show that the Forkhead transcription factor FoxM1 regulates expression of many G2-specific genes and is essential for chromosome stability. Loss of FoxM1 leads to pleiotropic cell-cycle defects, including a delay in G2, chromosome mis-segregation and frequent failure of cytokinesis. We show that transcriptional activation of cyclin B by FoxM1 is essential for timely mitotic entry, whereas CENP-F, another direct target of FoxM1 identified here, is essential for precise functioning of the mitotic spindle checkpoint. Thus, our data uncover a transcriptional cluster regulated by FoxM1 that is essential for proper mitotic progression.

No abstract

The evidence for T-cell-mediated regression of human cancers such as non-small-cell lung carcinoma, renal cell carcinoma, and-in particular-melanoma after immunotherapy is

Protein ubiquitination and deubiquitination are dynamic processes implicated in the regulation of numerous cellular pathways. Monoubiquitination of the Fanconi anemia (FA) protein FANCD2 appears to be critical in the repair of DNA damage because many of the proteins that are mutated in FA are required for FANCD2 ubiquitination. By screening a gene family RNAi library, we identify the deubiquitinating enzyme USP1 as a novel component of the Fanconi anemia pathway. Inhibition of USP1 leads to hyperaccumulation of monoubiquitinated FANCD2. Furthermore, USP1 physically associates with FANCD2, and the proteins colocalize in chromatin after DNA damage. Finally, analysis of crosslinker-induced chromosomal aberrations in USP1 knockdown cells suggests a role in DNA repair. We propose that USP1 deubiquitinates FANCD2 when cells exit S phase or recommence cycling after a DNA damage insult and may play a critical role in the FA pathway by recycling FANCD2.

Women carrying germ-line mutations in BRCA1 are strongly predisposed to developing breast cancers with characteristic features also observed in sporadic basal-like breast cancers. They appear as high-grade tumors with high proliferation rates and pushing borders. On the molecular level, they are negative for hormone receptors and ERBB2, display frequent TP53 mutations, and express basal epithelial markers. To study the role of BRCA1 and P53 loss of function in breast cancer development, we generated conditional mouse models with tissue-specific mutation of Brca1 and/or p53 in basal epithelial cells. Somatic loss of both BRCA1 and p53 resulted in the rapid and efficient formation of highly proliferative, poorly differentiated, estrogen receptor-negative mammary carcinomas with pushing borders and increased expression of basal epithelial markers, reminiscent of human basal-like breast cancer. BRCA1-and p53-deficient mouse mammary tumors exhibit dramatic genomic instability, and their molecular signatures resemble those of human BRCA1-mutated breast cancers. Thus, these tumors display important hallmarks of hereditary breast cancers in BRCA1-mutation carriers.mouse models ͉ conditional knockout G erm-line mutations in the human breast cancer susceptibility gene BRCA1 are responsible for 40% to 50% of hereditary breast cancers and confer increased risk for development of ovarian, colon, and prostate cancers (1, 2). BRCA1 has been implicated in various cellular processes, including maintenance of genome integrity, DNA replication and repair, chromatin remodeling, and transcriptional regulation (3, 4). Although the exact mechanism of mammary tumor suppression by BRCA1 remains largely unknown, cells with dysfunctional BRCA1 show defects in survival and proliferation, increased radiosensitivity, chromosomal abnormalities, G 2 /M checkpoint loss, and impaired homologous recombination repair (5).BRCA1-mutated breast cancers that arise in women with germline mutations in BRCA1 are high-grade, hormone receptornegative breast carcinomas with frequent mutation of TP53 (4, 6). They also possess a basal-like phenotype as defined by the expression of markers that are typical for basal/myoepithelial cells, such as the basal cytokeratins (CKs) CK5/6 and CK14 (7). Indeed, strong molecular similarities are observed between hereditary BRCA1-mutated breast cancers and sporadic basal-like breast carcinomas (8,9). This phenotypic overlap has led to the hypothesis that sporadic basal-like cancers may have defects in BRCA1-related pathways, such as the amplification of EMSY and the methylation of BRCA1 and FANCF (10).Despite the fact that several mouse strains with conventional or conditional mutations in Brca1 have been generated (11), no good mouse model for BRCA1-mutated basal-like breast cancer has been developed so far. Most conventional Brca1 knockouts are embryonic-lethal when bred to homozygosity, yet heterozygous ⌬11 allele, which encodes BRCA1-⌬11, a naturally occurring splice variant of Brca1 (19). Mouse mammary tumor models ba...