Genome folding principles uncovered in condensin-depleted mitotic chromosomes.

During mitosis, condensin activity is thought to interfere with interphase chromatin structures. To investigate genome folding principles in the absence of chromatin loop extrusion, we codepleted condensin I and condensin II, which triggered mitotic chromosome compartmentalization in ways similar to that in interphase. However, two distinct euchromatic compartments, indistinguishable in interphase, emerged upon condensin loss with different interaction preferences and dependencies on H3K27ac. Constitutive heterochromatin gradually self-aggregated and cocompartmentalized with facultative heterochromatin, contrasting with their separation during interphase. Notably, some cis-regulatory element contacts became apparent even in the absence of CTCF/cohesin-mediated structures. Heterochromatin protein 1 (HP1) proteins, which are thought to partition constitutive heterochromatin, were absent from mitotic chromosomes, suggesting, surprisingly, that constitutive heterochromatin can self-aggregate without HP1. Indeed, in cells traversing from M to G1 phase in the combined absence of HP1α, HP1ß and HP1γ, constitutive heterochromatin compartments are normally re-established. In sum, condensin-deficient mitotic chromosomes illuminate forces of genome compartmentalization not identified in interphase cells.

Genome folding principles revealed in condensin-depleted mitotic chromosomes.

During mitosis, condensin activity interferes with interphase chromatin structures. Here, we generated condensin-free mitotic chromosomes to investigate genome folding principles. Co-depletion of condensin I and II, but neither alone, triggered mitotic chromosome compartmentalization in ways that differ from interphase. Two distinct euchromatic compartments, indistinguishable in interphase, rapidly emerged upon condensin loss with different interaction preferences and dependence on H3K27ac. Constitutive heterochromatin gradually self-aggregated and co-compartmentalized with the facultative heterochromatin, contrasting with their separation during interphase. While topologically associating domains (TADs) and CTCF/cohesin mediated structural loops remained undetectable, cis-regulatory element contacts became apparent, providing an explanation for their quick re-establishment during mitotic exit. HP1 proteins, which are thought to partition constitutive heterochromatin, were absent from mitotic chromosomes, suggesting, surprisingly, that constitutive heterochromatin can self-aggregate without HP1. Indeed, in cells traversing from M- to G1-phase in the combined absence of HP1α, HP1ß and HP1γ, re-established constitutive heterochromatin compartments normally. In sum, "clean-slate" condensing-deficient mitotic chromosomes illuminate mechanisms of genome compartmentalization not revealed in interphase cells.

Altered cfDNA fragmentation profile in hypomethylated regions as diagnostic markers in breast cancer.

BACKGROUND: Breast cancer, the most common malignancy in women worldwide, has been proven to have both altered plasma cell-free DNA (cfDNA) methylation and fragmentation profiles. Nevertheless, simultaneously detecting both of them for breast cancer diagnosis has never been reported. Moreover, although fragmentation pattern of cfDNA is determined by nuclease digestion of chromatin, structure of which may be affected by DNA methylation, whether cfDNA methylation and fragmentation are biologically related or not still remains unclear. METHODS: Improved cfMeDIP-seq were utilized to characterize both cfDNA methylation and fragmentation profiles in 49 plasma samples from both healthy individuals and patients with breast cancer. The feasibility of using cfDNA fragmentation profile in hypo- and hypermethylated regions as diagnostic markers for breast cancer was evaluated. RESULTS: Mean size of cfDNA fragments (100-220 bp) mapped to hypomethylated regions decreased more in patients with breast cancer (4.60 bp, 172.33 to 167.73 bp) than in healthy individuals (2.87 bp, 174.54 to 171.67 bp). Furthermore, proportion of short cfDNA fragments (100-150 bp) in hypomethylated regions when compared with it in hypermethylated regions was found to increase more in patients with breast cancer in two independent discovery cohort. The feasibility of using abnormality of short cfDNA fragments ratio in hypomethylated genomic regions for breast cancer diagnosis in validation cohort was evaluated. 7 out of 11 patients were detected as having breast cancer (63.6% sensitivity), whereas no healthy individuals were mis-detected (100% specificity). CONCLUSION: We identified enriched short cfDNA fragments after 5mC-immunoprecipitation (IP) in patients with breast cancer, and demonstrated the enriched short cfDNA fragments might originated from hypomethylated genomic regions. Furthermore, we proved the feasibility of using differentially methylated regions (DMRs)-dependent cfDNA fragmentation profile for breast cancer diagnosis.

Novel lncRNAs with diagnostic or prognostic value screened out from breast cancer via bioinformatics analyses.

Background: Recent studies have shown that long non-coding RNAs (lncRNAs) may play key regulatory roles in many malignant tumors. This study investigated the use of novel lncRNA biomarkers in the diagnosis and prognosis of breast cancer. Materials and Methods: The database subsets of The Cancer Genome Atlas (TCGA) by RNA-seq for comparing analysis of tissue samples between breast cancer and normal control groups were downloaded. Additionally, anticoagulant peripheral blood samples were collected and used in this cohort study. The extracellular vesicles (EVs) from the plasma were extracted and sequenced, then analyzed to determine the expressive profiles of the lncRNAs, and the cancer-related differentially expressed lncRNAs were screened out. The expressive profiles and associated downstream-mRNAs were assessed using bioinformatics (such as weighted correlation network analysis (WGCNA), Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genome (KEGG) enrichments, Receiver-Operating Characteristic (ROC) curve and survival analysis, etc.) to investigate the diagnostic and prognostic values of these EV lncRNAs and their effectors. Results: In this study, 41 breast cancer-related lncRNAs were screen out from two datasets of tissue and fresh collected plasma samples of breast cancer via the transcriptomic and bioinformatics techniques. A total of 19 gene modules were identified with WGCNA analysis, of which five modules were significantly correlated with the clinical stage of breast cancer, including 28 lncRNA candidates. The ROC curves of these lncRNAs revealed that the area under the curve (AUC) of all candidates were great than 70%. However, eight lncRNAs had an AUC >70%, indicating that the combined one has a good diagnostic value. In addition, the results of survival analysis suggested that two lncRNAs with low expressive levels may indicate the poor prognosis of breast cancer. By tissue sample verification, C15orf54, AL157935.1, LINC01117, and SNHG3 were determined to have good diagnostic ability in breast cancer lesions, however, there was no significant difference in the plasma EVs of patients. Moreover, survival analysis data also showed that AL355974.2 may serve as an independent prognostic factor and as a protective factor. Conclusion: A total of five lncRNAs found in this study could be developed as biomarkers for breast cancer patients, including four diagnostic markers (C15orf54, AL157935.1, LINC01117, and SNHG3) and a potential prognostic marker (AL355974.2).