Topological reorganization and functional alteration of distinct genomic components in gallbladder cancer.

Altered three-dimensional architecture of chromatin influences various genomic regulators and subsequent gene expression in human cancer. However, knowledge of the topological rearrangement of genomic hierarchical layers in cancer is largely limited. Here, by taking advantage of in situ Hi-C, RNA-sequencing, and chromatin immunoprecipitation sequencing (ChIP-seq), we investigated structural reorganization and functional changes in chromosomal compartments, topologically associated domains (TADs), and CCCTC binding factor (CTCF)-mediated loops in gallbladder cancer (GBC) tissues and cell lines. We observed that the chromosomal compartment A/B switch was correlated with CTCF binding levels and gene expression changes. Increased inter-TAD interactions with weaker TAD boundaries were identified in cancer cell lines relative to normal controls. Furthermore, the chromatin short loops and cancer unique loops associated with chromatin remodeling and epithelial-mesenchymal transition activation were enriched in cancer compared with their control counterparts. Cancer-specific enhancer-promoter loops, which contain multiple transcription factor binding motifs, acted as a central element to regulate aberrant gene expression. Depletion of individual enhancers in each loop anchor that connects with promoters led to the inhibition of their corresponding gene expressions. Collectively, our data offer the landscape of hierarchical layers of cancer genome and functional alterations that contribute to the development of GBC.

Long-term bone remodeling of maxillary anterior teeth with post-treatment alveolar bone defect in adult patients with maxillary protrusion: a prospective follow-up study.

BACKGROUND: Alveolar bone defects, particularly palatal bone dehiscence (PBD) and labial bone fenestration (LBF), occur frequently as a result of retraction of the maxillary anterior teeth. The study aims to explore the long-term bone remodeling of maxillary anterior teeth in adult patients with post-orthodontic treatment PBD and LBF. MATERIALS AND METHODS: The study includes 24 adult patients with maxillary protrusion (8 males, 16 females) who were treated with extraction of four first premolars and had alveolar bone defects (PBD or LBF) in maxillary anterior teeth following orthodontic treatment. Cone-beam computed tomography imaging measurements were obtained before (T1), after (T2) orthodontic treatment, and after at least 1-year removable thermoplastic retainer retention (T3). The maxillary anterior teeth with PBD or LBF at T2 were divided into the PBD or LBF groups, respectively. The labial and palatal alveolar bone height (ABH), alveolar bone thickness (ABT), and movement of maxillary anterior teeth were measured during retraction (T2-T1) and retention (T3-T2) periods. RESULTS: The incidence of PBD and LBF in maxillary anterior teeth significantly increased after orthodontic treatment and decreased during the retention period. In the PBD group, the palatal ABH of all maxillary anterior teeth significantly increased from T1 to T2 but decreased from T2 to T3. The ABT of the maxillary central incisor and canine significantly increased on the palatal side and decreased on the labial side during the retention period. In the LBF group, the labial ABT of the maxillary central incisor at the apical level showed a significant decrease from T1 to T2, followed by an increase from T2 to T3. In both groups, the maxillary central incisor showed significant labial movement, with a relative intrusion during the retention period. CONCLUSION: For adult patients with maxillary protrusion, the alveolar bone defect of maxillary anterior teeth caused by orthodontic retraction significantly improved during the retention period, indicating good long-term bone remodeling. Our findings suggest that a combination of spontaneous reorientation of maxillary anterior teeth and bone remodeling contributed to alveolar bone covering in these patients.

A novel three-dimensional quantitative assessment method for abnormal root morphology of the maxillary premolars in vivo on cone-beam computed tomography.

BACKGROUND: Previous studies have described and recorded abnormal root morphology; however, most of these studies were based on two-dimensional periapical or panoramic radiographs, and only a few studies have quantified it. We aimed to combine two-dimensional periapical radiographs and three-dimensional cone-beam computed tomography (CBCT) to conduct qualitative judgments and quantitative analyses of normal and conical roots, and explore the clinical diagnostic method of normal and conical roots based on intraoral radiographs and CBCT. METHODS: The conical root was identified visually on periapical radiographs as the clinical gold standard. All teeth were divided into the cone-rooted teeth (CRT) or normal-rooted teeth (NRT) groups. Furthermore, differences in root length (RL), root surface area (RSA), and root volume (RV) of conical and normal roots in the maxillary premolars on CBCT were compared. Receiver operator characteristic curves were generated, and the area under the curve (AUC) and cut-off values were calculated to evaluate the diagnostic value of RV, RSA, RV/RL, and RSA/RL. RESULTS: The RSAs of NRT and CRT were 236.88 ± 27.93 mm2 and 207.98 ± 27.80 mm2, respectively (P = 0.000). The mean RV in the CRT group was lower than that in the NRT group, and the difference was statistically significant (253.40 ± 41.98 mm3 vs. 316.93 ± 49.89 mm3, P = 0.000). The RSA and RV of conical roots in single root premolars were 12.29% and 19.33% less than those of normal roots, respectively. The AUC values of RSA/RL and RV/RL were 0.87 and 0.89, respectively, and the best cut-off values were 19.61 for RSA/RL (if RSA/RL was < 19.61, the teeth were considered CRT) and 24.05 for RV/RL (if RV/RL was < 24.05, the teeth were considered CRT). CONCLUSIONS: CBCT has significant diagnostic value in the clinical evaluation of conical roots. RSA/RL and RV/RL were the best parameters with the largest AUC and high sensitivity and specificity.

MiR-4733-5p promotes gallbladder carcinoma progression via directly targeting kruppel like factor 7.

Gallbladder carcinoma (GBC) is highly aggressive with poor prognosis. Accumulating reports show that miRNAs play critical roles in tumor progression. Previous studies have identified several miRNAs that promoted or inhibited GBC cell proliferation and/or metastasis. Here, we used the Gene Expression Omnibus (GEO) dataset to identify dysregulated miRNAs in GBC, followed by validating the upregulation of the miR-4733-5p and downregulation of kruppel-like factor 7 (KLF7) in GBC biopsies by quantitative real-time PCR (RT-qPCR), in situ hybridization (ISH) staining, and immunohistochemistry (IHC) assays. GBC cell proliferation and invasion capacities mediated by miR-4733-5p were evaluated by a series of function assays in vitro, including CCK-8, colony formation assay, wound healing assay and transwell assay. Xenograft tumor model found that miR-4733-5p promoted GBC tumor growth in vivo. This study clarified that miR-4733-5p was upregulated in GBC and promoted GBC cell proliferation via directly binding to 3' untranslated region (UTR) of KLF, which was downregulated and prohibited the proliferation and migration of GBC cells.

Ring finger protein 6 enhances chemo-resistance by transcriptionally activating proliferating cell nuclear antigen expression and attenuating DNA damage in lung adenocarcinoma.

The E3 ubiquitin ligase RING finger protein 6 (RNF6) is elevated in several cancers, including prostate and colorectal cancers. Here, we extended the finding of elevated RNF6 expression levels and its association with poor prognosis in patients with lung adenocarcinoma (LUAD). Genome-wide RNA sequencing in H3255 cells with RNF6 knockdown, followed by analysis of differentially expressed genes using Clusters of Orthologous Groups and gene set enrichment analysis revealed aberrations in genes related to DNA repair, especially double-strand break (DSB) repair. RNF6 knockdown increased γH2AX foci, a biomarker for DSBs in H3255 and A549 LUAD cells, and enhanced DNA damage induced by chemotherapy in cisplatin-resistant A549/CDDP cells. In a series of experiments in cultured cells, as well as in nude mice carrying xenografts, RNF6 knockdown restored the sensitivity of A549/CDDP cells to cisplatin treatment. Mechanistically, RNA sequencing in RNF6-knockdown cells revealed the significant downregulation of proliferating cell nuclear antigen (PCNA), an oncogene that promotes DNA repair. Re-chromatin immunoprecipitation assay results suggested the formation of a RNF6-TCF4 complex that binds to the PCNA promoter to activate its transcription. Downregulation of RNF6 reduced TCF4 recruitment to PCNA promoters in H3255 and A549 cells, indicating that RNF6 regulates PCNA transcription to a certain extent by regulating TCF4 binding to PCNA promoters. The collective results implicate RNF6 overexpression as a molecular target in the management of cisplatin-resistant LUAD.

Parallel and Intertwining Threads of Domestication in Allopolyploid Cotton.

The two cultivated allopolyploid cottons, Gossypium hirsutum and Gossypium barbadense, represent a remarkable example of parallel independent domestication, both involving dramatic morphological transformations under selection from wild perennial plants to annualized row crops. Deep resequencing of 643 newly sampled accessions spanning the wild-to-domesticated continuum of both species, and their allopolyploid relatives, are combined with existing data to resolve species relationships and elucidate multiple aspects of their parallel domestication. It is confirmed that wild G. hirsutum and G. barbadense were initially domesticated in the Yucatan Peninsula and NW South America, respectively, and subsequently spread under domestication over 4000-8000 years to encompass most of the American tropics. A robust phylogenomic analysis of infraspecific relationships in each species is presented, quantify genetic diversity in both, and describe genetic bottlenecks associated with domestication and subsequent diffusion. As these species became sympatric over the last several millennia, pervasive genome-wide bidirectional introgression occurred, often with striking asymmetries involving the two co-resident genomes of these allopolyploids. Diversity scans revealed genomic regions and genes unknowingly targeted during domestication and additional subgenomic asymmetries. These analyses provide a comprehensive depiction of the origin, divergence, and adaptation of cotton, and serve as a rich resource for cotton improvement.

The Gossypium longicalyx Genome as a Resource for Cotton Breeding and Evolution.

Cotton is an important crop that has made significant gains in production over the last century. Emerging pests such as the reniform nematode have threatened cotton production. The rare African diploid species Gossypium longicalyx is a wild species that has been used as an important source of reniform nematode immunity. While mapping and breeding efforts have made some strides in transferring this immunity to the cultivated polyploid species, the complexities of interploidal transfer combined with substantial linkage drag have inhibited progress in this area. Moreover, this species shares its most recent common ancestor with the cultivated A-genome diploid cottons, thereby providing insight into the evolution of long, spinnable fiber. Here we report a newly generated de novo genome assembly of G. longicalyx This high-quality genome leveraged a combination of PacBio long-read technology, Hi-C chromatin conformation capture, and BioNano optical mapping to achieve a chromosome level assembly. The utility of the G. longicalyx genome for understanding reniform immunity and fiber evolution is discussed.

Genomic insights into divergence and dual domestication of cultivated allotetraploid cottons.

BACKGROUND: Cotton has been cultivated and used to make fabrics for at least 7000 years. Two allotetraploid species of great commercial importance, Gossypium hirsutum and Gossypium barbadense, were domesticated after polyploidization and are cultivated worldwide. Although the overall genetic diversity between these two cultivated species has been studied with limited accessions, their population structure and genetic variations remain largely unknown. RESULTS: We resequence the genomes of 147 cotton accessions, including diverse wild relatives, landraces, and modern cultivars, and construct a comprehensive variation map to provide genomic insights into the divergence and dual domestication of these two important cultivated tetraploid cotton species. Phylogenetic analysis shows two divergent groups for G. hirsutum and G. barbadense, suggesting a dual domestication processes in tetraploid cottons. In spite of the strong genetic divergence, a small number of interspecific reciprocal introgression events are found between these species and the introgression pattern is significantly biased towards the gene flow from G. hirsutum into G. barbadense. We identify selective sweeps, some of which are associated with relatively highly expressed genes for fiber development and seed germination. CONCLUSIONS: We report a comprehensive analysis of the evolution and domestication history of allotetraploid cottons based on the whole genomic variation between G. hirsutum and G. barbadense and between wild accessions and modern cultivars. These results provide genomic bases for improving cotton production and for further evolution analysis of polyploid crops.

Small interfering RNAs from bidirectional transcripts of GhMML3_A12 regulate cotton fiber development.

Natural antisense transcripts (NATs) are commonly observed in eukaryotic genomes, but only a limited number of such genes have been identified as being involved in gene regulation in plants. In this research, we investigated the function of small RNA derived from a NAT in fiber cell development. Using a map-based cloning strategy for the first time in tetraploid cotton, we cloned a naked seed mutant gene (N1 ) encoding a MYBMIXTA-like transcription factor 3 (MML3)/GhMYB25-like in chromosome A12, GhMML3_A12, that is associated with fuzz fiber development. The extremely low expression of GhMML3_A12 in N1 is associated with NAT production, driven by its 3' antisense promoter, as indicated by the promoter-driven histochemical staining assay. In addition, small RNA deep sequencing analysis suggested that the bidirectional transcriptions of GhMML3_A12 form double-stranded RNAs and generate 21-22 nt small RNAs. Therefore, in a fiber-specific manner, small RNA derived from the GhMML3_A12 locus can mediate GhMML3_A12 mRNA self-cleavage and result in the production of naked seeds followed by lint fiber inhibition in N1 plants. The present research reports the first observation of gene-mediated NATs and siRNA directly controlling fiber development in cotton.

Genetic basis for glandular trichome formation in cotton.

Trichomes originate from epidermal cells and can be classified as either glandular or non-glandular. Gossypium species are characterized by the presence of small and darkly pigmented lysigenous glands that contain large amounts of gossypol. Here, using a dominant glandless mutant, we characterize GoPGF, which encodes a basic helix-loop-helix domain-containing transcription factor, that we propose is a positive regulator of gland formation. Silencing GoPGF leads to a completely glandless phenotype. A single nucleotide insertion in GoPGF, introducing a premature stop codon is found in the duplicate recessive glandless mutant (gl2gl3). The characterization of GoPGF helps to unravel the regulatory network of glandular structure biogenesis, and has implications for understanding the production of secondary metabolites in glands. It also provides a potential molecular basis to generate glandless seed and glanded cotton to not only supply fibre and oil but also provide a source of protein for human consumption.

Sequencing of allotetraploid cotton (Gossypium hirsutum L. acc. TM-1) provides a resource for fiber improvement.

Upland cotton is a model for polyploid crop domestication and transgenic improvement. Here we sequenced the allotetraploid Gossypium hirsutum L. acc. TM-1 genome by integrating whole-genome shotgun reads, bacterial artificial chromosome (BAC)-end sequences and genotype-by-sequencing genetic maps. We assembled and annotated 32,032 A-subgenome genes and 34,402 D-subgenome genes. Structural rearrangements, gene loss, disrupted genes and sequence divergence were more common in the A subgenome than in the D subgenome, suggesting asymmetric evolution. However, no genome-wide expression dominance was found between the subgenomes. Genomic signatures of selection and domestication are associated with positively selected genes (PSGs) for fiber improvement in the A subgenome and for stress tolerance in the D subgenome. This draft genome sequence provides a resource for engineering superior cotton lines.

Identification of centromeric regions on the linkage map of cotton using centromere-related repeats.

Centromere usually contains high-copy-number retrotransposons and satellite repeats, which are difficult to map, clone and sequence. Currently, very little is known about the centromere in cotton. Here, we sequenced a bacterial artificial chromosome (BAC) mapping to the centromeric region and predicted four long-terminal-repeat (LTR) retrotransposons. They were located in the heterochromatic centromeric regions of all 52 pachytene chromosomes in Gossypium hirsutum. Fiber-FISH mapping revealed that these retrotransposons span an area of at least 1.8Mb in the centromeric region. Comparative analysis showed that these retrotransposons generated similar, strong fluorescent signals in the D progenitor Gossypium raimondii but not in the A progenitor Gossypium herbaceum, suggesting that the centromere sequence of tetraploid cotton might be derived from the D progenitor. Centromeric regions were anchored on 13 chromosomes of D-genome sequence. Characterization of these centromere-related repeats and regions will enhance cotton centromere mapping, sequencing and evolutionary studies.