Absence of Brca2 causes genome instability by chromosome breakage and loss associated with centrosome amplification

AbstractWomen heterozygous for mutations in the breast-cancer susceptibility genes BRCA1 and BRCA2 have a highly elevated risk of developing breast cancer [1]. BRCA1 and BRCA2 encode large proteins with no sequence similarity to one another. Although involvement in DNA repair and transcription has been suggested, it is still not understood how loss of function of these genes leads to breast cancer [2]. Embryonic fibroblasts (MEFs) derived from mice homozygous for a hypomorphic mutation (Brca2Tr2014) within the 3′ region of exon 11 in Brca2[3], or a similar mutation (Brca2Tr) [4], proliferate poorly in culture and overexpress the tumour suppressor p53 and the cyclin-dependent kinase inhibitor p21Waf1/Cip1. These MEFs have intact p53-dependent DNA damage G1–S [3,4] and G2–M checkpoints [4], but are impaired in DNA double-strand break repair [3] and develop chromosome aberrations [4]. Here, we report that Brca2Tr2014/Tr2014 MEFs frequently develop micronuclei. These abnormal DNA-containing bodies were formed through both loss of acentric chromosome fragments and by chromosome missegregation, which resulted in aneuploidy. Absence of Brca2 also led to centrosome amplification, which we found associated with the formation of micronuclei. These data suggest a potential mechanism whereby loss of BRCA2 may, within subclones, drive the loss of cell-cycle regulation genes, enabling proliferation and tumourigenesis

Collaborator of alternative reading frame protein (CARF) regulates early processing of pre-ribosomal RNA by retaining XRN2 (5′-3′ exoribonuclease) in the nucleoplasm

Collaborator of alternative reading frame protein (CARF) associates directly with ARF, p53, and/or human double minute 2 protein (HDM2), a ubiquitin-protein ligase, without cofactors and regulates cell proliferation by forming a negative feedback loop. Although ARF, p53, and HDM2 also participate in the regulation of ribosome biogenesis, the involvement of CARF in this process remains unexplored. In this study, we demonstrate that CARF associates with 5′-3′ exoribonuclease 2 (XRN2), which plays a major role in both the maturation of rRNA and the degradation of a variety of discarded pre-rRNA species. We show that overexpression of CARF increases the localization of XRN2 in the nucleoplasm and a concomitant suppression of pre-rRNA processing that leads to accumulation of the 5′ extended from of 45S/47S pre-rRNA and 5′-01, A0-1 and E-2 fragments of pre-rRNA transcript in the nucleolus. This was also observed upon XRN2 knockdown. Knockdown of CARF increased the amount of XRN2 in the nucleolar fraction as determined by cell fractionation and by immnocytochemical analysis. These observations suggest that CARF regulates early steps of pre-rRNA processing during ribosome biogenesis by controlling spatial distribution of XRN2 between the nucleoplasm and nucleolus

Vortex core switching by coherent excitation with single in-plane magnetic field pulses

The bistability of the core magnetization of nano-scaled magnets with a magnetic vortex configuration has great potential for data storage applications. To exploit this, reliable switching between the two possible states is needed. Time resolved x-ray microscopy was used to study the response of the vortex core to excitation pulses at sub-ns timescales and image the vortex core switching. A reliable switching process by coherent excitation with leading and trailing edges of in-plane magnetic field pulses was found and compared with micromagnetic simulations.Comment: 4 pages, 3 figure

Selective requirement of H2B N-Terminal tail for p14ARF-induced chromatin silencing

The N-terminal tail of histone H2B is believed to be involved in gene silencing, but how it exerts its function remains elusive. Here, we report the biochemical characterization of p14ARF tumor suppressor as a transcriptional repressor that selectively recognizes the unacetylated H2B tails on nucleosomes. The p14ARF–H2B tail interaction is functional, as the antagonistic effect of p14ARF on chromatin transcription is lost upon deletion or acetylation of H2B tails. Gene expression profiling and chromatin immunoprecipitation studies emphasize the significance of H2B deacetylation and p14ARF recruitment in establishing a repressive environment over the cell cycle regulatory genes. Moreover, HDAC1-mediated H2B deacetylation, especially at K20, constitutes an essential step in tethering p14ARF near target promoters. Our results thus reveal a hitherto unknown role of p14ARF in the regulation of chromatin transcription, as well as molecular mechanisms governing the repressive action of p14ARF

Parp1 facilitates alternative NHEJ, whereas Parp2 suppresses IgH/c-myc translocations during immunoglobulin class switch recombination

Immunoglobulin class switch recombination (CSR) is initiated by DNA breaks triggered by activation-induced cytidine deaminase (AID). These breaks activate DNA damage response proteins to promote appropriate repair and long-range recombination. Aberrant processing of these breaks, however, results in decreased CSR and/or increased frequency of illegitimate recombination between the immunoglobulin heavy chain locus and oncogenes like c-myc. Here, we have examined the contribution of the DNA damage sensors Parp1 and Parp2 in the resolution of AID-induced DNA breaks during CSR. We find that although Parp enzymatic activity is induced in an AID-dependent manner during CSR, neither Parp1 nor Parp2 are required for CSR. We find however, that Parp1 favors repair of switch regions through a microhomology-mediated pathway and that Parp2 actively suppresses IgH/c-myc translocations. Thus, we define Parp1 as facilitating alternative end-joining and Parp2 as a novel translocation suppressor during CSR

The pathology of familial breast cancer: The pathology of familial breast cancer How do the functions of BRCA1 and BRCA2 relate to breast tumour pathology?

Women with mutations in the breast cancer susceptibility genes, BRCA1 and BRCA2, have an increased risk of developing breast cancer. Both BRCA1 and BRCA2 are thought to be tumour suppressor genes since the wild type alleles of these genes are lost in tumours from heterozygous carriers. Several functions have been proposed for the proteins encoded by these genes which could explain their roles in tumour suppression. Both BRCA1 and BRCA2 have been suggested to have a role in transcriptional regulation and several potential BRCA1 target genes have been identified. The nature of these genes suggests that loss of BRCA1 could lead to inappropriate proliferation, consistent with the high mitotic grade of BRCA1-associated tumours. BRCA1 and BRCA2 have also been implicated in DNA repair and regulation of centrosome number. Loss of either of these functions would be expected to lead to chromosomal instability, which is observed in BRCA1 and BRCA2-associated tumours. Taken together, these studies give an insight into the pathogenesis of BRCA-associated tumours and will inform future therapeutic strategies