Regulator Of Chromosome Condensation Research Articles

M6A-modified exosome-derived circHIF1α binding to KH domain of IGF2BP3 mediates DNA damage and arrests G1/S transition phase to resists bacterial infection in bacteremia

BackgroundAnimal and human health are seriously threatened by bacterial infections, which can lead to bacteremia and extremely high rates of morbidity and mortality. Recently, there have been reports indicating the involvement of exosomal circular RNAs (circRNAs) in a range of human disorders and tumor types. However, the role of exosomal circRNAs in bacterial infection remains elusive.MethodsWe extracted and identified exosomes from the culture medium of PIEC cells infected with or without Glaesserella parasuis. RNA sequencing analysis was performed on the exosomes to screen and identify circRNAs (circHIF1α) associated with Glaesserella parasuis infection. PIEC cells were infected with Staphylococcus aureus or Streptococcus suis 2 to further determine whether exosome-derived circHIF1α was the crucial circHIF1α associated with bacterial infections. The transmission process of exosomes and their circHIF1α between cells was clarified via exosome tracing and co-culture assay. Moreover, the mechanism of circHIF1α being packaged into exosomes was explored, and the effects of exosomes and their circHIF1α on cell proliferation, DNA damage and cell cycle were analyzed. In addition, the binding mode and site of interacting proteins with circHIF1α were further determined. In vivo and in vitro, the role of exosomes and their circHIF1α in host resistance to bacterial infection was confirmed.ResultsWe first discovered a new circHIF1α that was very stable and detectable, encapsulated into exosomes by hnRNPA2B1, and whose expression in exosomes of bacterially infected PIEC cells significantly decreased. Additionally, exosomal circHIF1α reduced bacterial infection both in vitro and in vivo and suppressed the growth of reception cells. Mechanistically, the circHIF1α interacted with the KH domain of IGF2BP3 in an m6A-modified manner, which mediated DNA damage to arrest the cells at the G1/S phase through the interaction between the regulator of Chromosome Condensation 2 (RCC2) and γ-H2AX protein. Exosomal circHIF1α is a unique therapeutic target for bacterial infection since this work highlights its critical function in fighting bacterial infection.

The chromosomal translocation t(1;6)(p35.3;p25.2), recurrent in chronic lymphocytic leukaemia, leads to RCC1::IRF4 fusion.

The chromosomal translocation t(1;6)(p35.3;p25.2) is a rare but recurrent aberration in chronic lymphocytic leukaemia (CLL). We report molecular characterization of 10 cases and show that this translocation juxtaposes interferon regulatory factor 4 (IRF4) on 6p25 with regulator of chromosome condensation 1 (RCC1) on 1p35. The breakpoints fell within the 5' untranslated regions of both genes, resulting in RCC1::IRF4 fusion transcripts without alterations of the protein-coding sequences. Levels of expression of both RCC1 and IRF4 proteins were not obviously deregulated. The cases showed other mutations typical of CLL and we confirm previously reported skewing towards the IGHV-unmutated subtype. RCC1::IRF4 fusion characterizes a rare subset of CLL.

The discovery of the RCC1::IRF4 Fusion in CLL patients with t(1;6)(p35.3;p25.2) chromosomal translocation.

Jayne etal. provide a molecular characterization of the t(1;6)(p35.3;p25.2) chromosomal translocation in patients with chronic lymphocytic leukaemia. They indicate that this translocation involves the gene encoding interferon regulatory factor 4 (IRF4) on chromosome 6p25.2 with the regulator of chromosome condensation 1 (RCC1) gene on chromosome 1p35.3. This translocations may have important prognostic value. Commentary on: Jayne etal. The chromosomal translocation t(1;6)(p35.3;p25.2), recurrent in chronic lymphocytic leukaemia leads to RCC1::IRF4 fusion. Br J Haematol 2024 (Online ahead of print). doi: 10.1111/bjh.19790.

Exploring Novel Genomic Loci and Candidate Genes Associated with Plant Height in Bulgarian Bread Wheat via Multi-Model GWAS.

In the context of crop breeding, plant height (PH) plays a pivotal role in determining straw and grain yield. Although extensive research has explored the genetic control of PH in wheat, there remains an opportunity for further advancements by integrating genomics with growth-related phenomics. Our study utilizes the latest genome-wide association scan (GWAS) techniques to unravel the genetic basis of temporal variation in PH across 179 Bulgarian bread wheat accessions, including landraces, tall historical, and semi-dwarf modern varieties. A GWAS was performed with phenotypic data from three growing seasons, the calculated best linear unbiased estimators, and the leveraging genotypic information from the 25K Infinium iSelect array, using three statistical methods (MLM, FarmCPU, and BLINK). Twenty-five quantitative trait loci (QTL) associated with PH were identified across fourteen chromosomes, encompassing 21 environmentally stable quantitative trait nucleotides (QTNs), and four haplotype blocks. Certain loci (17) on chromosomes 1A, 1B, 1D, 2A, 2D, 3A, 3B, 4A, 5B, 5D, and 6A remain unlinked to any known Rht (Reduced height) genes, QTL, or GWAS loci associated with PH, and represent novel regions of potential breeding significance. Notably, these loci exhibit varying effects on PH, contribute significantly to natural variance, and are expressed during seedling to reproductive stages. The haplotype block on chromosome 6A contains five QTN loci associated with reduced height and two loci promoting height. This configuration suggests a substantial impact on natural variation and holds promise for accurate marker-assisted selection. The potentially novel genomic regions harbor putative candidate gene coding for glutamine synthetase, gibberellin 2-oxidase, auxin response factor, ethylene-responsive transcription factor, and nitric oxide synthase; cell cycle-related genes, encoding cyclin, regulator of chromosome condensation (RCC1) protein, katanin p60 ATPase-containing subunit, and expansins; genes implicated in stem mechanical strength and defense mechanisms, as well as gene regulators such as transcription factors and protein kinases. These findings enrich the pool of semi-dwarfing gene resources, providing the potential to further optimize PH, improve lodging resistance, and achieve higher grain yields in bread wheat.

RCC1 regulation of subcellular protein localization via Ran GTPase drives pancreatic ductal adenocarcinoma growth

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal malignancy, with limited therapeutic options. Here, we evaluated the role of regulator of chromosome condensation 1 (RCC1) in PDAC. RCC1 functions as a guanine exchange factor for GTP-binding nuclear protein Ran (Ran) GTPase and is involved in nucleocytoplasmic transport. RCC1 RNA expression is elevated in PDAC tissues compared to normal pancreatic tissues and correlates with poor prognosis. RCC1 silencing by RNAi and CRISPR-Cas9 knockout (KO) results in reduced proliferation in 2-D and 3-D cell cultures. RCC1 knockdown (KD) reduced migration and clonogenicity, enhanced apoptosis, and altered cell cycle progression in human PDAC and murine cells from LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx1-Cre (KPC) tumors. Mechanistically, RCC1 KO shows widespread transcriptomic alterations including regulation of PTK7, a co-receptor of the Wnt signaling pathway. RCC1 KD disrupted subcellular Ran localization and the Ran gradient. Nuclear and cytosolic proteomics revealed altered subcellular proteome localization in Rcc1 KD KPC-tumor-derived cells and several altered metabolic biosynthesis pathways. In vivo, RCC1 KO cells show reduced tumor growth potential when injected as sub-cutaneous xenografts. Finally, RCC1 KD sensitized PDAC cells to gemcitabine chemotherapy treatment. This study reveals the role of RCC1 in pancreatic cancer as a novel molecular vulnerability that could be exploited to enhance therapeutic response.

CoCl2‐induced glioma hypoxia environment enhances CD47‐SIRPα to promote tumor immune evasion

AbstractElevated levels of tumor‐associated macrophages and microglia in the immune microenvironment of malignant gliomas promote tumor growth and progression. Although immune evasion has been implicated in these processes, the mechanisms underlying the regulation of CD47‐SIRPα‐mediated immune evasion under hypoxic conditions remain unclear. Therefore, this study aimed to explore the mechanisms through which CD47‐SIRPα modulates immune evasion in a cobalt chloride (CoCl2)‐induced hypoxic microenvironment of malignant gliomas. Human and mouse glioma cell lines were used to investigate immune evasion in vitro. After membrane protein extraction and nucleoplasmic isolation, we downregulated the expression levels of transport receptors to elucidate the regulation of CD47‐SIRPα transport. The CoCl2‐induced hypoxic microenvironment attenuated microglial phagocytosis in glioma cells. Enhanced CD47‐SIRPα interaction promoted immune evasion, which negatively affected tumor clearance. In addition, the intracellular transport of CD47 was promoted in the CoCl2‐induced hypoxic state. This process was regulated by regulator of chromosome condensation 1 (RCC1) and sortilin in the nuclear cytoplasm and cytoplasmic membrane, respectively. These results demonstrate the CD47‐SIRPα‐mediated immune escape mechanism of the CoCl2‐induced glioma hypoxic environment, which is associated with enhanced intracellular transport of CD47. Our findings provide a theoretical basis for the potential development of novel therapies targeting CD47‐SIRPα.

Abstract 5587: Regulator of chromosome condensation 1 (RCC1) as a novel therapeutic target in pancreatic ductal adenocarcinoma

Abstract Background and Introduction: Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease with limited treatment options. There is an urgent need for the identification of novel therapeutic targets for PDAC. The transport of molecular cargo between the nucleus and cytoplasm, facilitated by Ran-GTPase, is a critical process that dividing cells rely on to maintain their growth. Elevated nuclear Ran-GTP levels are produced by the activity of the Ran guanine nucleotide exchange factor (GEF) known as Regulator of Chromosome Condensation 1 (RCC1), which is found in the nucleus bound to chromatin. This creates a Ran-GTP gradient across the nuclear membrane, which maintains proper directional transport of proteins and RNA. To sustain proliferative signaling, cancer cells become reliant on high rates of nucleocytoplasmic transport through dysregulation of transport machinery. Methods: We examined the role of RCC1 in the biology of PDAC. The impact of RCC1 modulation on PDAC growth was evaluated using RNA interference and CRISPR-Cas9 in vitro and in vivo using PDAC cell lines and LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx1-Cre (KPC) tumor-derived cells. The broader impact of RCC1 silencing on PDAC-sustaining signaling was evaluated through RNA-sequencing and proteomics. Results: Transcriptomic sequencing of PDAC tissue (n=5,071) as well as analysis of publicly available data from the TCGA, CPTAC and GTEx revealed that RCC1 expression is higher in PDAC tissues compared to normal pancreas. Moreover, PDAC patients with higher RCC1 expression were more likely to have poorer outcomes. RCC1 silencing by RNAi and CRISPR-Cas9 resulted in reduced proliferation in 2D and 3D cultures and attenuated tumor growth in vivo. RCC1 KD decreased migration and colony formation, enhanced apoptosis, and altered the cell cycle in human and KPC mouse PDAC cells. Subcutaneous RCC1 KO cell line-derived xenografts show arrested growth. Subcellular Ran distribution was disrupted upon RCC1 KO, suggesting that nuclear Ran concentration is important for PDAC proliferation. Nuclear and cytosolic proteomic analysis revealed altered subcellular proteome in RCC1 KD KPC tumor-derived cells. Altered cytoplasmic protein pathways include several metabolic pathways, PI3K-AKT, and Hedgehog signaling. Nuclear enriched pathways include cell cycle, mitosis, metabolic processes, and RNA processing. RNA-seq of RCC1 KO cells showed widespread transcriptional alterations. Upstream of RCC1, c-MYC activates the RCC1-RAN axis, and RCC1 KO cells show differential sensitivity to c-MYC inhibitors. Finally, RCC1 KD resulted in the sensitization of PDAC cells to Gemcitabine. Molecular characterization of conditional Rcc1fl/fl KO transgenic model to study the role of RCC1 in PDAC in vivo is ongoing. Conclusions: Overall, our results show that RCC1 is involved in the regulation of PDAC growth and is a potential target for therapy. Citation Format: Sahar F. Bannoura, Husain Y. Khan, Amro Aboukameel, Md Hafiz Uddin, Rund Nimri, Eliza Beal, Seongho Kim, Mohammed Najeeb Al Hallak, Bassel El-Rayes, Philip A. Philip, Boris Pasche, Ramzi Mohammad, Asfar S. Azmi. Regulator of chromosome condensation 1 (RCC1) as a novel therapeutic target in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5587.

Abstract 3062: Unraveling the role of RCC2 in glioblastoma by mutational analysis and a murine model of glioma

Abstract Glioblastoma (GBM) is a brain malignancy characterized by being highly aggressive, lethal, yet treatment modalities remain limited. Recent studies have implicated Regulator of Chromosome Condensation 2 (RCC2), a guanine nucleotide exchange factor (GEF) for RalA, as a potential regulator in glioblastoma pathogenesis. RCC2 has been shown to play a role in promoting glucose metabolism through BACH1-dependent upregulation of hexokinase II (HK2)(1). In this study, we investigated the functional mechanism of RCC2 in gliomagenesis using two complementary approaches. First, the relevance of RCC2 structural regions on its downstream functions were examined. By superimposing the three-dimensional crystal structure of RCC2 onto RCC1 (Regulator of Chromosome Condensation 1), a related GEF, we identified several unique regions within the RCC1-like domain of RCC2. A panel of point, substitution, and deletion mutants were generated, and pulldown experiments were performed to assess their relative binding affinity with BACH1. Additional assays to assess their relative ability to modulate the expression of HK2 and other cellular parameters will be discussed. The structure-functional characterization of distinct regions within RCC2 may identify unique sequences within RCC2 that could have relevance to glucose metabolism, cell proliferation and cell motility. Additionally, we have established a murine glioma model utilizing a Pten heterozygous mice expressing a GFAP-Cre-recombinase transgene (Gfap-cre;Ptenfl/+). Glioma were induced by stereotaxically injected lentiviral constructs expressing either with HRasV12 oncogene alone or together with RCC2. Neurological symptoms were monitored over a period of 7 months, and histopathological analysis was performed to evaluate the morphological and molecular features of the tumors. We showed that co-expression of HRasV12 and RCC2 exhibited earlier emergence of neurological manifests in both male and female groups (Log-rank test, p<0.05) when compared to HRasV12 alone. Overall, our study provides insights into both the structural mechanism of RCC2 in gliomagenesis through mutational analysis of RCC2 and the functional relevance through a murine model of glioma. (1)Liu, T. et. al. Cancer Lett (2022) 549:215914. Funding: This work is supported by a General Research Fund grant (#14113519) from the University Grants Committee of Hong Kong. Citation Format: Wayne L.H. Ho, Penelope M.Y. Or, Andrew M. Chan. Unraveling the role of RCC2 in glioblastoma by mutational analysis and a murine model of glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3062.

Significance of RCC2, Rac1 and p53 Expression in Breast Infiltrating Ductal Carcinoma; An Immunohistochemical Study.

The regulator of chromosome condensation 2 (RCC2) and RAS-related C3 botulinum toxin substrate 1 (Rac1) have been implicated in the promotion of breast cancer cell proliferation and migration. The signaling pathway involving p53/RCC2/Rac1 has been proposed to contribute to the regulation of colon cancer metastasis. However, until now, this pathway has not been thoroughly investigated in breast cancer. This study seeks to explore the influence of immunohistochemical expression and the correlation among RCC2, Rac1, and p53 in breast infiltrating ductal carcinoma (IDC). Immunostaining was performed on 120 breast IDC specimens using RCC2, Rac1, and p53 antibodies. Statistical analyses were conducted to examine the correlations between these antibodies. A Positive expression of RCC2, Rac1, and p53 was observed in 116 (96.7%), 120 (100%), and 33 (27.5%) of the breast cancer cases, respectively. RCC2, Rac1, and p53 demonstrated association with poor prognostic parameters such as frequent mitoses, high Ki-67 status, positive lymphovascular invasion (LVI), and advanced tumor stage. A highly significant direct correlation was found between each immunohistochemical marker and the other two markers. Shorter overall survival was linked to multifocal tumors (P=0.017), advanced tumor stage (T3) (P=0.010), Luminal B subtype (P=0.015), progressive disease (P=0.003), positive Her2neu status (P=0.008), and metastasis to distant organs (P<0.001). However, RCC2, Rac1, and p53 did not exhibit a significant association with overall survival. The high expression levels of RCC2, Rac1, and p53 in breast IDC suggest their potential role in tumor behavior. The association of RCC2 and Rac1 with poor prognostic parameters may serve as predictive indicators for aggressive tumors, thus implying that targeted therapy could be beneficial in the treatment of breast cancer.

Comprehensive analysis of prognostic value and immune infiltration of Regulator of Chromosome Condensation 2 in lung adenocarcinoma.

Background: Lung adenocarcinoma (LUAD) incidence and mortality take the leading place of most malignancies. Previous studies have revealed the regulator of chromosome condensation 1 (RCC1) family members played an essential role during tumorigenesis. However, its biological functions in LUAD still need further investigation. Methods: Several databases were applied to explore potential effects of RCC1 family members on LUAD, such as Oncomine, GEPIA, and cBioPortal. Real-time PCR and immunohistochemistry were used to verify the expression of RCC2 in stage I LUAD. H1975 and A549 were selected to explore the biological function of RCC2 in cellular malignant phenotype. Results: The expressions of RCC1 and RCC2 showed marked differences in malignant tissue compared to lung tissue. The higher the expression levels of RCC1 or RCC2 in LUAD patients, the shorter their overall survival (OS). In normal lung tissues, RCC1 expression was highly enriched in alveolar cells and endothelial cells. Compare with RCC1, RCC2 expression in normal lung tissue was significantly enriched in macrophages, B cells and granulocytes. Additionally, RCC2 expression level was correlated with multiple immune cell infiltration in LUAD. Moreover, the mutation or different sCNA status of RCC2 exerted influence on multiple immune cell infiltration distribution. We found that the upregulation of RCC1 and RCC2 were obviously related to TP53 mutation. GSEA analysis revealed that RCC2 was involved in the process of DNA replication, nucleotide excision repair and cell cycle, which might affect tumor progression through P53 signaling pathway. We further elucidated that downregulation of RCC2 could dramatically repress the migration and invasion of LUAD cells. Conclusions: The study demonstrated that RCC1 and RCC2 expression were markedly increased in early-stage of LUAD. Patients with high expression of RCC1 or RCC2 had a worse prognosis. Based on our analysis, RCC1 and RCC2 might exert influence on LUAD process through DNA replication, nucleotide excision repair and cell cycle, as well as cells migration and invasion. Different from RCC1, RCC2 also involved in immune infiltration. These analyses provided a novel insight into the identification of diagnostic biomarker.

Long non-coding RNA (LncRNA) non-coding RNA activated by DNA damage (NORAD) knockdown alleviates airway remodeling in asthma via regulating miR-410-3p/RCC2 and inhibiting Wnt/β-catenin pathway

BackgroundAsthma is a chronic inflammatory disorder with high prevalence in childhood. Airway remodeling, an important structural change of the airways, is resulted from epithelial-mesenchymal transition. Long non-coding RNA non-coding RNA activated by DNA damage (NORAD) has been found to promote epithelial-mesenchymal transition in multiple cancers. This study aimed to analyze the role of NORAD in asthma, mainly focusing on epithelial-mesenchymal transition-mediated airway remodeling, and further explored the NORAD-miRNA-mRNA network. MethodsNORAD expression was analyzed in transforming growth factor-β1-induced BEAS-2B human bronchial epithelial cells and ovalbumin-challenged asthmatic mice. The influences of NORAD on the epithelial-mesenchymal transition characteristics and Wnt/β-catenin pathway activation were analyzed in vitro. The interactions between NORAD and miR-410-3p as well as miR-410-3p and regulator of chromosome condensation 2 were detected by dual-luciferase reporter assay and RNA pull-down assay. Rescue experiments using miR-410-3p antagonist and chromosome condensation 2 overexpression were used to confirm the mechanism of NORAD. Additionally, the role and mechanism of NORAD were further evaluated in asthmatic mice. ResultsNORAD expression was elevated in both asthmatic models. Knockdown of NORAD impeded spindle-like morphology changes, elevated E-cadherin expression, decreased N-cadherin expression, suppressed cell migration, and inactivated the Wnt/β-catenin pathway in transforming growth factor-β1-stimulated BEAS-2B cells. NORAD acted as a sponge of miR-410-3p to regulate chromosome condensation 2 expression. Rescue assays demonstrated that silencing of NORAD ameliorated transforming growth factor-β1-induced EMT via miR-410-3p/chromosome condensation 2/Wnt/β-catenin axis. In vivo, knockdown of NORAD led to the reduction of inflammatory cell infiltration and collagen deposition, suppression of IL-4, IL-13, transforming growth factor-β1 and immunoglobulin E production, decreasing of N-cadherin, chromosome condensation 2, β-catenin and c-Myc expression, but increasing of E-cadherin and miR-410-3p expression. ConclusionsSilencing of NORAD alleviated epithelial-mesenchymal transition-mediated airway remodeling in asthma via mediating miR-410-3p/chromosome condensation 2/Wnt/β-catenin pathway.

Impact of KIF4A on Cancer Stem Cells and EMT in Lung Cancer and Glioma

Kinesin family member 4A (KIF4A) belongs to the kinesin 4 subfamily of kinesin-related proteins and is involved in the regulation of chromosome condensation and segregation during mitotic cell division. The expression of KIF4A in various types of cancer, including lung, breast, and colon cancer, has been found to be associated with poor prognosis in cancer patients. However, the exact mechanism by which it promotes tumorigenesis is not yet understood. In osteosarcoma, the expression of KIF4A has been shown to be associated with cancer stem cells (CSCs), whereas in breast cancer, it is not associated with the maintenance of CSCs but regulates the migratory ability of cells. In this light, we identified phenotypic phenomena affecting the malignancy of cancer in lung cancer and glioma, and investigated the mechanisms promoting tumorigenesis. As a result, we demonstrated that KIF4A affected lung cancer stem cells (LCSCs) and glioma stem cells (GSCs) and regulated CSC signaling mechanisms. In addition, the migratory ability of cells was regulated by KIF4A, and epithelial-to-mesenchymal transition (EMT) marker proteins were controlled. KIF4A regulated the expression of the secretory factor plasminogen activator inhibitor-1 (PAI-1), demonstrating that it sustains cancer malignancy through an autocrine loop. Taken together, these findings suggest that KIF4A regulates CSCs and EMT, which are involved in cancer recurrence and metastasis, indicating its potential value as a novel therapeutic target and prognostic marker in lung cancer and glioma.

Genome-wide characterization of regulator of chromosome condensation 1 (RCC1) gene family in Artemisia annua L. revealed a conservation evolutionary pattern

BackgroundArtemisia annua is the major source for artemisinin production. The artemisinin content in A. annua is affected by different types of light especially the UV light. UVR8, a member of RCC1 gene family was found to be the UV-B receptor in plants. The gene structures, evolutionary history and expression profile of UVR8 or RCC1 genes remain undiscovered in A. annua.ResultsTwenty-two RCC1 genes (AaRCC1) were identified in each haplotype genome of two diploid strains of A. annua, LQ-9 and HAN1. Varied gene structures and sequences among paralogs were observed. The divergence of most RCC1 genes occurred at 46.7 – 51 MYA which overlapped with species divergence of core Asteraceae during the Eocene, while no recent novel RCC1 members were found in A. annua genome. The number of RCC1 genes remained stable among eudicots and RCC1 genes underwent purifying selection. The expression profile of AaRCC1 is analogous to that of Arabidopsis thaliana (AtRCC1) when responding to environmental stress.ConclusionsThis study provided a comprehensive characterization of the AaRCC1 gene family and suggested that RCC1 genes were conserved in gene number, structures, constitution of amino acids and expression profiles among eudicots.

DNA Methylation Landscape of Peripheral T-Cell Lymphomas Not Otherwise Specified

DNA methylation is an epigenetic modification catalyzed by DNA methyltransferases that regulate gene transcription in mammalian cells. DNA methylation in gene promoters is associated with repressive histone marks and contributes to gene repression. This process is involved in various physiological processes, including development, X chromosome inactivation, genomic imprinting, differentiation, and hematopoiesis. The global, genome-wide deregulation of methylation patterns contributes to the pathogenesis of immune disorders, hematologic malignancies, and cancer. Because aberrations in DNA methylation can deregulate spatiotemporal gene expression in a cell type-specific manner, genome-wide changes in this epigenetic modification may contribute to tumor development. Specifically, the downregulation of tumor suppressors through promoter hypermethylation is a widely accepted mechanism in cancer progression. Due to its distinct patterns in cancer, DNA methylation can be used as a diagnostic and predictive biomarker. Peripheral T-cell lymphomas, Not Otherwise Specified (PTCL-NOS), are a group of heterogeneous T-cell malignancies with diverse cytogenic, molecular, and morphological phenotypes. Despite large-scale gene expression profiling, the molecular features-and DNA methylation in particular-of these rare but aggressive malignancies remain poorly understood. To better understand the molecular landscape of PTCL-NOS, we performed global methylation profiling via Whole Genome Bisulfite Sequencing on 10 lymphomas and compared those to patterns seen in normal T-cells and Anaplastic Large Cell Lymphoma (ALCL). Our analysis revealed large-scale deregulation of methylation in all tumors at levels of both differentially methylated cytosines and regions. The dominant molecular change was hypomethylation, which was, on average, 7 times more frequent than hypermethylation. Hypomethylation was not limited to repetitive elements but was also seen in all genomic elements, including promoters, enhancers, gene bodies, and exons. Out of 1,286 genes whose promoters were consistently hypomethylated in Peripheral T-cell lymphomas (PTCL), 39 genes were associated with an increase in expression, suggesting that the loss of methylation may have contributed to their deregulation. This hypomethylated and overexpressed signature contained genes that might be involved in tumor progression or maintenance, such as Rac GTPase activating protein 1 ( RACGAP1), regulator of chromosome condensation 1 ( RCC1), anillin actin-binding protein ( ANLN), and others. Out of 578 genes whose promoters were consistently hypermethylated in PTCL, the expression of 56 genes was decreased. These included genes with tumor suppressor function, such as splicing factor 3b subunit 1 ( SF3B1) and lymphoid enhancer-binding factor 1 ( LEF1). Our analysis also identified differential methylation between PTCL-NOS and ALCL in various genomic features. For example, 71 hypo- and 59 hypermethylated promoters were found only in ALCL but not PTCL-NOS. Conversely, 326 hypo- and 254 hypermethylated promoters were observed only in PTCL-NOS. Analysis of gene expression also revealed specific changes in both PTCL subtypes, manifested by the activation of different signaling pathways as identified by Ingenuity Pathway Analysis. PTCL-NOS was characterized by the suppression of the SUMOylation pathway, PTEN, and EIF2 Signaling, and the activation of NF-κB and Notch signaling. ALCLs were characterized by the suppression of the Role of CHK Proteins in Cell Cycle Control, Erythropoietin, ERBB signaling, the Coronavirus Pathogenesis pathway, IGF-1 signaling, and the activation of Glutathione-mediated Detoxification and Triacylglycerol Biosynthesis. Altogether, we identified recurrent specific methylation and expression changes that could serve as the basis for the development of tools facilitating the diagnosis and prognosis of PTCL-NOS. In addition, our data may contribute to the identification of epigenetically-regulated drivers of tumor initiation, progression, and maintenance.

Multi-omic characterization of RCC1 expression and its association with molecular alterations, immune phenotypes, and cancer outcomes.

3128 Background: Regulator of Chromosome Condensation 1 (RCC1) is the only identified guanine nucleotide exchange factor for the Ras-related nuclear protein Ran and it functions in nuclear transport, cell cycle and DNA damage response. Overexpressed in several cancer types, RCC1 is associated with poor outcomes. We aim to investigate the role of aberrant RCC1 co-alterations and association with immune phenotypes and cancer outcomes. Methods: DNA (592-gene or whole exome)/RNA (whole transcriptome) sequencing was performed at Caris Life Sciences (Phoenix, AZ). Samples were stratified by RCC1 expression quartile thresholds (Q1:low, Q4:high) for small cell lung (SCLC, n=876), non-small cell lung (NSCLC, n=21603), gastric (GC, n=1908), pancreatic (PC, n=5071) and colorectal (CRC, n=14892) cancers. PD-L1+ expression was tested by IHC (22c3: ³1%; SP142: ³2+, 5%). TMB-High was defined as ³ 10 mutations/MB. Overall survival (OS) was calculated using Kaplan-Meier estimate. Statistical significance was determined using chi-square and Wilcoxon rank sum test and adjusted for multiple comparisons (*P&lt;0.05). Results: Median RCC1 expression was highest in SCLC (14.3 transcript per million [TPM]), followed by GC (9.9), NSCLC (9.9), CRC (9.8), and PC (6.9). TMB-high rates increased progressively with RCC1 expression in NSCLC (Q1-Q4 range: 31-41%*), GC (7-22%*) and CRC (3-13%*), consistent with dMMR/MSI-high rates in GC (5-18%*) and CRC (3-13%*). Similar trends were observed for PDL1+ rates in PC (Q1-Q4 range: 13-20%*), CRC (2-5%*) and NSCLC (55-58%*). In NSCLC, TP53 (Q1-Q4 range: 57-77%*), RB1 (7-13%*), and EGFR (10-15%*) mutations were seen more with higher RCC1 expression, while STK11 (15-10%*) and KRAS (33-21%*) mutations were seen less. In PC, TP53 mutations (Q1-Q4 range: 70-85%*) and MYC amplifications (1-4%*) were seen more with RCC1 expression, whereas ATM mutations were seen less (6-3%). In CRC, TP53 (Q1-Q4 range: 70-81%*) and APC (72-81%*) mutations were seen more with RCC1 expression, while GNAS mutations were seen less (4-2%*). In both SCLC and NSCLC, high RCC1 expression was associated with increased dendritic cell (5-6%* and .8-1.3%*, respectively) and NK cell fractions (5-6%* and 2.5-3.2%*) and decreased Treg fractions (1.8-1.3%* and 3.0-2.6%*). In pMMR/MSS CRC, high RCC1 expression was associated with increased dendritic cell (.6-.9%*), NK cell (3-4%*), neutrophil (6-8%*) and CD4 T cell fractions (1.1-1.2%*) and decreased CD8 T cell (.5-.4%*) and Treg fractions (1.9-1.7%*). High RCC1 expression was associated with worse OS in NSCLC (HR 1.3*), PC (HR 1.5*) and CRC (HR 1.3*), with a similar but not significant effect in SCLC (HR 1.2) and GC (HR 1.2). Conclusions: RCC1 expression is a negative prognostic marker in NSCLC, PC, and CRC. Further studies to investigate this at the molecular level may be a potential opportunity for novel targeted drug development.

Identification of Potential Biomarkers in Stomach Adenocarcinoma using Machine Learning Approaches

Background: Stomach adenocarcinoma (STAD) is a common cancer with poor clinical outcomes globally. Due to a lack of early diagnostic markers of disease, the majority of patients are diagnosed at an advanced stage. Objective: The aim of the present study is to provide some new insights into the available biomarkers for patients with STAD using bioinformatics. Methods: RNA-Sequencing and other relevant data of patients with STAD from The Cancer Genome Atlas (TCGA) database were evaluated to identify differentially expressed genes (DEGs). Then, Machine Learning algorithms were undertaken to predict biomarkers. Additionally, Kaplan–Meier analysis was used to detect prognostic biomarkers. Furthermore, the Gene Ontology and Reactome pathways, protein-protein interactions (PPI), multiple sequence alignment, phylogenetic mapping, and correlation between clinical parameters were evaluated. Results: The results showed 61 DEGs, and the key dysregulated genes associated with STAD are MTHFD1L (Methylenetetrahydrofolate dehydrogenase 1-like), ZWILCH (Zwilch Kinetochore Protein), RCC2 (Regulator of chromosome condensation 2), DPT (Dermatopontin), GCOM1 (GRINL1A complex locus 1), and CLEC3B (C-Type Lectin Domain Family 3 Member B). Moreover, the survival analysis reported ASPA (Aspartoacylase) as a prognostic marker. Conclusion: Our study provides a proof of concept of the potential value of ASPA as a prognostic factor in STAD, requiring further functional investigations to explore the value of emerging markers.

Conservative Protein RCC1 Is a New Component of Black Bands of &lt;i&gt;Drosophila&lt;/i&gt; &lt;i&gt;melanogaster&lt;/i&gt; Polytene Chromosomes

Previously, the RCC1 gene (Regulator of Chromosome Сondensation 1) was characterized, which is considered a regulator of chromosome condensation in the cell cycle. This gene encodes a nuclear protein whose amino acid sequence is highly conserved among all eukaryotes and consists of seven repeating units. We have shown that all the most prominent black bands of polytene chromosomes (about 250) and the chromocenter bind antibodies to this protein. We found that antibodies to the xenopus RCC1 protein specifically bind to the Drosophila and human RCC1 protein, while in Drosophila lines with under replication suppression, the relative amount of the RCC1 protein increases compared to the wild type.

Genome-Wide Identification and Expression Analysis of RCC1 Gene Family under Abiotic Stresses in Rice (Oryza sativa L.)

In plants, the essential roles played by the regulator of chromosome condensation 1 (RCC1) in diverse biological processes, including UV-B (ultraviolet-B radiation) response, hormonal signal transduction, cold tolerance and phenotypic plasticity, have been identified. No comprehensive study on the evolution and function of RCC1 gene family in rice has been carried out. A genome-wide analysis of this gene family is thus required. In this study, a total of 26 OsRCC1s unevenly distributed across 10 chromosomes were identified in rice. Based on their phylogenetic relationship and sequence composition, the OsRCC1 family could be classified into six groups. Members within the same group share a similar gene structure and protein motif/domain composition. Gene duplication analysis revealed that segmental duplication might be the main contributor to the expansion of the RCC1 gene family in rice. Several cis-regulatory elements (CREs) relevant to light, abscisic acid (ABA) and methyl jasmonate (MeJA) are abundant in the promoters of OsRCC1s. A large number of microRNA (miRNA) target sites were present in OsRCC1 mRNAs. Additionally, we used data from gene microarray and qRT-PCR to analyze the expression of OsRCC1 genes during various developmental stages and under abiotic stress conditions. OsRCC1s were found to be highly expressed in panicles and seeds, and most OsRCC1s were differentially expressed under abiotic stresses. Taken together, our study provides a systematic characterization of OsRCC1s and preliminarily explores their diversity as well as their biological functions. Evidence demonstrates that OsRCC1s may play vital roles in both development and abiotic stress response. The results presented here lay a foundation for further investigating the functions of OsRCC1s.

Genome-wide identification and expression analysis of the regulator of chromosome condensation 1 gene family in wheat (Triticum aestivum L.).

Wheat (Triticum aestivum L., 2n = 6x = 42, AABBDD) is the world's most widely cultivated crop and an important staple food for humans, accounting for one-fifth of calories consumed. Proteins encoded by the regulator of chromosome condensation 1 (RCC1) are highly conserved among eukaryotes and consist of seven repeated domains that fold into a seven-bladed propeller structure. In this study, a total of 76 RCC1 genes of bread wheat were identified via a genome-wide search, and their phylogenetic relationship, gene structure, protein-conserved domain, chromosome localization, conserved motif, and transcription factor binding sites were systematically analyzed using the bioinformatics approach to indicate the evolutionary and functional features of these genes. The expression patterns of 76 TaRCC1 family genes in wheat under various stresses were further analyzed, and RT-PCR verified that RCC1-3A (TraesCS3A02G362800), RCC1-3B (TraesCS3B02G395200), and RCC1-3D (TraesCS3D02G35650) were significantly induced by salt, cold, and drought stresses. Additionally, the co-expression network analysis and binding site prediction suggested that Myb-7B (TraesCS7B02G188000) and Myb-7D (TraesCS7D02G295400) may bind to the promoter of RCC1-3A/3B and upregulate their expression in response to abiotic stresses in wheat. The results have furthered our understanding of the wheat RCC1 family members and will provide important information for subsequent studies and the use of RCC1 genes in wheat.

A human pan-cancer system analysis of regulator of chromatin condensation 2

Regulation of chromosome condensation 2 (RCC2) is associated with the cell cycle and is a crucial regulator of the chromatin condensation 1 (RCC1) family. The members of this family were normally regulators in the process of DNA replication and nucleocytoplasmic transport. RCC2 overexpression may lead to tumor formation and poor prognosis in some tumors including breast cancer and lung adenocarcinoma. However, the possible role of RCC2 in tumor formation and its prognostic function remains unclear. In this study, expression analysis from databases including The Cancer Genome Atlas (TCGA) and Clinical Proteomic Tumor Analysis Consortium (CPTAC) were combined to perform the first integrative and comprehensive analysis of RCC2 in human pan-cancer. RCC2 was highly expressed in most tumors which may lead to a poor prognosis. RCC2 expression was associated with immune/stromal infiltration, immune checkpoints, tumor mutational burden, and microsatellite instability. Thus, RCC2 could be a novel biomarker for prognosis and a promising cancer therapy target.