During the humoral immune response, B cells undergo a dramatic change in phenotype to enable antibody affinity maturation in germinal centers (GCs). Using genome-wide chromosomal conformation capture ...(Hi-C), we found that GC B cells undergo massive reorganization of the genomic architecture that encodes the GC B cell transcriptome. Coordinate expression of genes that specify the GC B cell phenotype—most prominently BCL6—was achieved through a multilayered chromatin reorganization process involving (1) increased promoter connectivity, (2) formation of enhancer networks, (3) 5′ to 3′ gene looping, and (4) merging of gene neighborhoods that share active epigenetic marks. BCL6 was an anchor point for the formation of GC-specific gene and enhancer loops on chromosome 3. Deletion of a GC-specific, highly interactive locus control region upstream of Bcl6 abrogated GC formation in mice. Thus, large-scale and multi-tiered genomic three-dimensional reorganization is required for coordinate expression of phenotype-driving gene sets that determine the unique characteristics of GC B cells.
Display omitted
•GC B cells feature massive unpacking of the arms of long chromosomes•The GC transcriptome is coordinated through GC-specific topologic reorganization•The BCL6 locus is a major hotspot for multilayered topological shifts in GC B cells•A GC B cell de novo highly interactive LCR specifies the GC phenotype
The formation of germinal center B cells is crucial to the production of high-affinity antibodies during the humoral immune response. Here, Bunting et al. provide evidence for multilayered reorganization of genomic architecture centered around BCL6 and an upstream locus control region, coordinating the transcriptional reprograming during the transition between naive and germinal center B cells.
The analysis of exonic DNA from prostate cancers has identified recurrently mutated genes, but the spectrum of genome-wide alterations has not been profiled extensively in this disease. We sequenced ...the genomes of 57 prostate tumors and matched normal tissues to characterize somatic alterations and to study how they accumulate during oncogenesis and progression. By modeling the genesis of genomic rearrangements, we identified abundant DNA translocations and deletions that arise in a highly interdependent manner. This phenomenon, which we term “chromoplexy,” frequently accounts for the dysregulation of prostate cancer genes and appears to disrupt multiple cancer genes coordinately. Our modeling suggests that chromoplexy may induce considerable genomic derangement over relatively few events in prostate cancer and other neoplasms, supporting a model of punctuated cancer evolution. By characterizing the clonal hierarchy of genomic lesions in prostate tumors, we charted a path of oncogenic events along which chromoplexy may drive prostate carcinogenesis.
Display omitted
► Interdependent DNA rearrangements may coordinately remodel prostate cancer genomes ► “Chromoplexy” defines a distinct class of complex structural rearrangements ► Multiple prostate cancer genes may be dysregulated coordinately ► Clonal evolution reveals paths of prostate cancer progression
Modeling the genesis of chromosomal rearrangements in prostate cancer genomes reveals that the chromosomal disarray found in a typical tumor may arise via a handful of discrete “chromoplexy” events during tumor development in which interdependent rearrangements shuffle DNA from five or more chromosomes.
Microsatellite instability (MSI) is characterized by the expansion or contraction of DNA repeat tracts as a consequence of DNA mismatch repair deficiency (MMRD). Accurate detection of MSI in cancer ...cells is important since MSI is associated with several cancer subtypes and can help inform therapeutic decisions. Although experimental assays have been developed to detect MSI, they typically depend on a small number of known microsatellite loci or mismatch repair genes and have limited reliability. Here, we report a novel genome-wide approach for MSI detection based on the global detection of insertions and deletions (indels) in microsatellites found in expressed genes. Our large-scale analyses of 20 cancer cell lines and 123 normal individuals revealed striking indel features associated with MSI: there is a significant increase of short microsatellite deletions in MSI samples compared to microsatellite stable (MSS) ones, suggesting a mechanistic bias of repair efficiency between insertions and deletions in normal human cells. By incorporating this observation into our MSI scoring metric, we show that our approach can correctly distinguish between MSI and MSS cancer cell lines. Moreover, when we applied this approach to primal tumor samples, our metric is also well consistent with diagnosed MSI status. Thus, our study offers new insight into DNA mismatch repair system, and also provides a novel MSI diagnosis method for clinical oncology with better reliability.
Transcription factors are frequently altered in leukaemia through chromosomal translocation, mutation or aberrant expression. AML1-ETO, a fusion protein generated by the t(8;21) translocation in ...acute myeloid leukaemia, is a transcription factor implicated in both gene repression and activation. AML1-ETO oligomerization, mediated by the NHR2 domain, is critical for leukaemogenesis, making it important to identify co-regulatory factors that 'read' the NHR2 oligomerization and contribute to leukaemogenesis. Here we show that, in human leukaemic cells, AML1-ETO resides in and functions through a stable AML1-ETO-containing transcription factor complex (AETFC) that contains several haematopoietic transcription (co)factors. These AETFC components stabilize the complex through multivalent interactions, provide multiple DNA-binding domains for diverse target genes, co-localize genome wide, cooperatively regulate gene expression, and contribute to leukaemogenesis. Within the AETFC complex, AML1-ETO oligomerization is required for a specific interaction between the oligomerized NHR2 domain and a novel NHR2-binding (N2B) motif in E proteins. Crystallographic analysis of the NHR2-N2B complex reveals a unique interaction pattern in which an N2B peptide makes direct contact with side chains of two NHR2 domains as a dimer, providing a novel model of how dimeric/oligomeric transcription factors create a new protein-binding interface through dimerization/oligomerization. Intriguingly, disruption of this interaction by point mutations abrogates AML1-ETO-induced haematopoietic stem/progenitor cell self-renewal and leukaemogenesis. These results reveal new mechanisms of action of AML1-ETO, and provide a potential therapeutic target in t(8;21)-positive acute myeloid leukaemia.
The processes of somatic hypermutation (SHM) and class switch recombination introduced by activation-induced cytosine deaminase (AICDA) at the Immunoglobulin (Ig) loci are key steps for creating a ...pool of diversified antibodies in germinal center B cells (GCBs). Unfortunately, AICDA can also accidentally introduce mutations at bystander loci, particularly within the 5' regulatory regions of proto-oncogenes relevant to diffuse large B cell lymphomas (DLBCL). Since current methods for genomewide sequencing such as Exon Capture and RNAseq only target mutations in coding regions, to date non-Ig promoter SHMs have been studied only in a handful genes. We designed a novel approach integrating bioinformatics tools with next generation sequencing technology to identify regulatory loci targeted by SHM genome-wide. We observed increased numbers of SHM associated sequence variant hotspots in lymphoma cells as compared to primary normal germinal center B cells. Many of these SHM hotspots map to genes that have not been reported before as mutated, including BACH2, BTG2, CXCR4, CIITA, EBF1, PIM2, and TCL1A, etc., all of which have potential roles in B cell survival, differentiation, and malignant transformation. In addition, using BCL6 and BACH2 as examples, we demonstrated that SHM sites identified in these 5' regulatory regions greatly altered their transcription activities in a reporter assay. Our approach provides a first cost-efficient, genome-wide method to identify regulatory mutations and non-Ig SHM hotspots.
During metastatic progression, circulating cancer cells become lodged within the microvasculature of end organs, where most die from mechanical deformation. Although this phenomenon was first ...described over a half-century ago, the mechanisms enabling certain cells to survive this metastasis-suppressive barrier remain unknown. By applying whole-transcriptome RNA-sequencing technology to isogenic cancer cells of differing metastatic capacities, we identified a mutation encoding a truncated form of the pannexin-1 (PANX1) channel, PANX1(1-89), as recurrently enriched in highly metastatic breast cancer cells. PANX1(1-89) functions to permit metastatic cell survival during traumatic deformation in the microvasculature by augmenting ATP release from mechanosensitive PANX1 channels activated by membrane stretch. PANX1-mediated ATP release acts as an autocrine suppressor of deformation-induced apoptosis through P2Y-purinergic receptors. Finally, small-molecule therapeutic inhibition of PANX1 channels is found to reduce the efficiency of breast cancer metastasis. These data suggest a molecular basis for metastatic cell survival on microvasculature-induced biomechanical trauma.
Emerging evidence suggests that chromatin adopts a nonrandom 3D topology and that the organization of genes into structural hubs and domains affects their transcriptional status. How chromatin ...conformation changes in diseases such as cancer is poorly understood. Moreover, how oncogenic transcription factors, which bind to thousands of sites across the genome, influence gene regulation by globally altering the topology of chromatin requires further investigation. To address these questions, we performed unbiased high-resolution mapping of intra- and interchromosome interactions upon overexpression of ERG, an oncogenic transcription factor frequently overexpressed in prostate cancer as a result of a gene fusion. By integrating data from genome-wide chromosome conformation capture (Hi-C), ERG binding, and gene expression, we demonstrate that oncogenic transcription factor overexpression is associated with global, reproducible, and functionally coherent changes in chromatin organization. The results presented here have broader implications, as genomic alterations in other cancer types frequently give rise to aberrant transcription factor expression, e.g., EWS-FLI1, c-Myc, n-Myc, and PML-RARα.
Copy number variants (CNVs) are a recently recognized class of human germ line polymorphisms and are associated with a variety of human diseases, including cancer. Because of the strong genetic ...influence on prostate cancer, we sought to identify functionally active CNVs associated with susceptibility of this cancer type. We queried low-frequency biallelic CNVs from 1,903 men of Caucasian origin enrolled in the Tyrol Prostate Specific Antigen Screening Cohort and discovered two CNVs strongly associated with prostate cancer risk. The first risk locus (P= 7.7 × 10⁻⁴, odds ratio = 2.78) maps to 15q21.3 and overlaps a noncoding enhancer element that contains multiple activator protein 1 (AP-1) transcription factor binding sites. Chromosome conformation capture (Hi-C) data suggested direct cis-interactions with distant genes. The second risk locus (P = 2.6 × 10⁻³ f odds ratio = 4.8) maps to the α-1,3-mannosyl-glycoprotein 4-β-N-acetylglucosaminyltransferase C (MGAT4C) gene on 12q21.31. In vitro cell-line assays found this gene to significantly modulate cell proliferation and migration in both benign and cancer prostate cells. Furthermore, MGAT4C was significantly overexpressed in metastatic versus localized prostate cancer. These two risk associations were replicated in an independent PSA-screened cohort of 800 men (15q21.3, combined P = 0.006; 12q21.31 , combined P = 0.026). These findings establish noncoding and coding germ line CNVs as significant risk factors for prostate cancer susceptibility and implicate their role in disease development and progression.
Adenosine-to-inosine RNA editing is crucial for generating molecular diversity, and serves to regulate protein function through recoding of genomic information. Here, we discover editing within ...Ca(v)1.3 Ca²⁺ channels, renown for low-voltage Ca²⁺-influx and neuronal pacemaking. Significantly, editing occurs within the channel's IQ domain, a calmodulin-binding site mediating inhibitory Ca²⁺-feedback (CDI) on channels. The editing turns out to require RNA adenosine deaminase ADAR2, whose variable activity could underlie a spatially diverse pattern of Ca(v)1.3 editing seen across the brain. Edited Ca(v)1.3 protein is detected both in brain tissue and within the surface membrane of primary neurons. Functionally, edited Ca(v)1.3 channels exhibit strong reduction of CDI; in particular, neurons within the suprachiasmatic nucleus show diminished CDI, with higher frequencies of repetitive action-potential and calcium-spike activity, in wild-type versus ADAR2 knockout mice. Our study reveals a mechanism for fine-tuning Ca(v)1.3 channel properties in CNS, which likely impacts a broad spectrum of neurobiological functions.
Abstract Background Certain patient demographics and histopathologic features are risk factors for papillary thyroid cancer (PTC) recurrence after initial treatment. Our objective was to determine ...whether very young age is associated with aggressive pathologic features in patients with papillary thyroid cancer. Materials and Methods A retrospective analysis was performed for PTC patients who underwent surgical treatment at the University of Michigan between 2006 and 2012. Patients with known distant metastases were excluded. Demographics, high-risk pathologic features (capsular or vascular invasion, extrathyroidal extension, lymph node metastases, extranodal extension), and disease recurrence were analyzed. Results 632 PTC patients were included in the analysis. Median age was 49 years (range 10–87). Tumors in patients <25 years of age had higher rates of extranodal extension ( p =0.002) compared to patients 25-44 years old. Patients <25 years had more vascular invasion ( p <0.001) and lymph node metastasis ( p =0.001) than tumors in patients between 45-75 years. Patients >75 years had higher rates of vascular invasion (p<0.001) and extrathyroidal extension (p=0.001) compared to patients 45-75 years and more extrathyroidal extension (p<0.001) than patients 25-44 years. There were no differences in tumor characteristics between the <25 and >75 age groups. Conclusions PTC patients <25 years of age or older than 75 years exhibit higher rates of aggressive histopathologic features compared to PTC patients between 25-75 years.
PDF
