Rearrangement of T Cell genome architecture regulates GVHD.

WAPL, cohesin's DNA release factor, regulates three-dimensional (3D) chromatin architecture. The 3D chromatin structure and its relevance to mature T cell functions is not well understood. We show that in vivo lymphopenic expansion, and alloantigen-driven proliferation, alters the 3D structure and function of the genome in mature T cells. Conditional deletion of WAPL, cohesin's DNA release factor, in T cells reduced long-range genomic interactions and altered chromatin A/B compartments and interactions within topologically associating domains (TADs) of the chromatin in T cells at baseline. WAPL deficiency in T cells reduced loop extensions, changed expression of cell cycling genes and reduced proliferation following in vitro and in vivo stimulation, and reduced severity of graft-versus-host disease (GVHD) following experimental allogeneic hematopoietic stem cell transplantation. These data collectively characterize 3D genomic architecture of T cells in vivo and demonstrate biological and clinical implications for its disruption by cohesin release factor WAPL.

Human CD206+ macrophages associate with diabetes and adipose tissue lymphoid clusters.

Increased adipose tissue macrophages (ATMs) correlate with metabolic dysfunction in humans and are causal in development of insulin resistance in mice. Recent bulk and single-cell transcriptomics studies reveal a wide spectrum of gene expression signatures possible for macrophages that depends on context, but the signatures of human ATM subtypes are not well defined in obesity and diabetes. We profiled 3 prominent ATM subtypes from human adipose tissue in obesity and determined their relationship to type 2 diabetes. Visceral adipose tissue (VAT) and s.c. adipose tissue (SAT) samples were collected from diabetic and nondiabetic obese participants to evaluate cellular content and gene expression. VAT CD206+CD11c- ATMs were increased in diabetic participants, were scavenger receptor-rich with low intracellular lipids, secreted proinflammatory cytokines, and diverged significantly from 2 CD11c+ ATM subtypes, which were lipid-laden, were lipid antigen presenting, and overlapped with monocyte signatures. Furthermore, diabetic VAT was enriched for CD206+CD11c- ATM and inflammatory signatures, scavenger receptors, and MHC II antigen presentation genes. VAT immunostaining found CD206+CD11c- ATMs concentrated in vascularized lymphoid clusters adjacent to CD206-CD11c+ ATMs, while CD206+CD11c+ were distributed between adipocytes. Our results show ATM subtype-specific profiles that uniquely contribute to the phenotypic variation in obesity.

TCF7L2 silencing results in altered gene expression patterns accompanied by local genomic reorganization.

Canonical Wnt signaling is crucial for intestinal homeostasis as TCF4, the major Wnt signaling effector in the intestines, is required for stem cell maintenance. The capability of TCF4 to maintain the stem cell phenotype is contingent upon ß-catenin, a potent transcriptional activator, which interacts with histone acetyltransferases and chromatin remodeling complexes. We used RNAi to explore the influence of TCF4 on chromatin structure (Hi-C) and gene expression (RNA sequencing) across a 72-hour time series in colon cancer. We found that TCF4 reduction results in a disproportionate up-regulation of gene expression, including a powerful induction of SOX2. Integration of RNA sequencing and Hi-C data revealed a TAD boundary loss, which occurred concomitantly with the over-expression of a cluster of CEACAM genes on chromosome 19. We identified EMT and E2F as the 2 most deregulated pathways upon TCF4 depletion and LUM, TMPO, and AURKA as highly influential genes in these networks using measures of centrality. Results from gene expression, chromatin structure, and centrality analyses were integrated to generate a list of candidate transcription factors crucial for colon cancer cell homeostasis. The top ranked factor was c-JUN, an oncoprotein known to interact with TCF4 and ß-catenin, confirming the usefulness of this approach.

Single Chromosome Aneuploidy Induces Genome-Wide Perturbation of Nuclear Organization and Gene Expression.

Chromosomal aneuploidy is a defining feature of carcinomas and results in tumor-entity specific genomic imbalances. For instance, most sporadic colorectal carcinomas carry extra copies of chromosome 7, an aneuploidy that emerges already in premalignant adenomas, and is maintained throughout tumor progression and in derived cell lines. A comprehensive understanding on how chromosomal aneuploidy affects nuclear organization and gene expression, i.e., the nucleome, remains elusive. We now analyzed a cell line established from healthy colon mucosa with a normal karyotype (46,XY) and its isogenic derived cell line that acquired an extra copy of chromosome 7 as its sole anomaly (47,XY,+7). We studied structure/function relationships consequent to aneuploidization using genome-wide chromosome conformation capture (Hi-C), RNA sequencing and protein profiling. The gain of chromosome 7 resulted in an increase of transcript levels of resident genes as well as genome-wide gene and protein expression changes. The Hi-C analysis showed that the extra copy of chromosome 7 is reflected in more interchromosomal contacts between the triploid chromosomes. Chromatin organization changes are observed genome-wide, as determined by changes in A/B compartmentalization and topologically associating domain (TAD) boundaries. Most notably, chromosome 4 shows a profound loss of chromatin organization, and chromosome 14 contains a large A/B compartment switch region, concurrent with resident gene expression changes. No changes to the nuclear position of the additional chromosome 7 territory were observed when measuring distances of chromosome painting probes by interphase FISH. Genome and protein data showed enrichment in signaling pathways crucial for malignant transformation, such as the HGF/MET-axis. We conclude that a specific chromosomal aneuploidy has profound impact on nuclear structure and function, both locally and genome-wide. Our study provides a benchmark for the analysis of cancer nucleomes with complex karyotypes.

Biomaterial Scaffolds Recruit an Aggressive Population of Metastatic Tumor Cells In Vivo.

For most cancers, metastasis is the point at which clinical treatment shifts from curative intent to extending survival. Biomaterial implants acting as a synthetic premetastatic niche recruit metastatic cancer cells and provide a survival advantage, and their use as a diagnostic platform requires assessing their relevance to disease progression. Here, we showed that scaffold-captured tumor cells (SCAF) were 30 times more metastatic to the lung than primary tumor (PT) cells, similar to cells derived from lung micrometastases (LUNG). SCAF cells were more aggressive in vitro, demonstrated higher levels of migration, invasion, and mammosphere formation, and had a greater proportion of cancer stem cells than PT. SCAF cells were highly enriched for gene expression signatures associated with metastasis and had associated genomic structural changes, including globally enhanced entropy. Collectively, our findings demonstrate that SCAF cells are distinct from PT and more closely resemble LUNG, indicating that tumor cells retrieved from scaffolds are reflective of cells at metastatic sites. SIGNIFICANCE: These findings suggest that metastatic tumor cells captured by a biomaterial scaffold may serve as a diagnostic for molecular staging of metastasis.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/79/8/2042/F1.large.jpg.