Cyclin-dependent Kinase Inhibitor Gene Research Articles

Comprehensive Analysis of the Significance of Cyclin Dependent Kinase Inhibitor Genes in the Prognosis of Cervical Cancer

Comprehensive Analysis of the Significance of Cyclin Dependent Kinase Inhibitor Genes in the Prognosis of Cervical Cancer

CDKN2A deletion and radiation sensitivity in IDH-mutant astrocytomas.

2041 Background: IDH-mutant astrocytomas represent the most prevalent primary tumors in younger adults (<45yo). A substantial minority of these tumors exhibit the deletion of CDKN2A (Cyclin-Dependent Kinase Inhibitor Gene 2A). Homozygous deletion of this gene– which encodes the tumor suppressor protein p16 – is associated with a malignant phenotype and poorer prognosis in IDH-mutant astrocytoma. CDKN2A deletions are primarily observed in tumors that have received radiation therapy (RT), suggesting a potential mechanistic relationship between RT and deletion. This observation gives rise to two alternative hypotheses: either RT is causing DNA damage in irradiated cells, leading to the induction of subclonal CDKN2A deletion, or a subset of cells already possesses the CDKN2A deletion and is resistant to RT, resulting in the treatment selecting for the subsequent emergence of these cells. Here, we sought to ascertain the influence of CDKN2A deletion on IDH-mutant astrocytoma cellular response to RT. Methods: The antitumor effect of RT was evaluated in vitro using patient-derived IDH-mutant astrocytoma cells, MGG152. A homozygous CDKN2A deletion was engineered in this parental line, creating a paired line to permit an isogenic comparison between CDKN2A intact versus CDKN2A deleted tumor cells. A photon irradiator was used to deliver a range of clinically-relevant RT doses spanning from 0 to 20 gray (Gy). Differences in cell viability after a predetermined incubation period were examined and relevant statistical comparisons performed. Results: Dose-response curves in response to radiation therapy, comparing CDKN2A intact and deleted cell lines, were generated (Table). Both CDKN2A intact and deleted cell lines exhibited significant reductions in viability following radiation exposure at and above 2Gy (p < 0.001). CDKN2A-deleted cells displayed a small but significantly greater sensitivity than CDKN2A-intact at 1, 2, and 5Gy (Table). Conclusions: Our findings offer evidence indicating that p16 loss does not bestow radioresistance in IDH-mutant astrocytomas. These findings align with a hypothesis that radiotherapy (RT) might promote DNA damage, and consequently, subclonal CDKN2A deletion, in irradiated IDH-mutant tumors. [Table: see text]

N6-Methyladenosine modified circ-NAB1 modulates cell cycle and epithelial-mesenchymal transition via CDKN3 in endometrial cancer.

Endometrial cancer (EC) is a common malignant tumor in the female reproductive system. Circular RNAs (circRNAs) and N6-methyladenosine (m6A) modification are widely involved in cancer progression. Nevertheless, the cross-talk between circ-NAB1 and m6A as well as the biological functions of circ-NAB1 in EC remain unclear. Circ-NAB1 was observed to be upregulated in EC tissues and cells by RT-qPCR. MeRIP and RNA pull-down assays were utilized for detecting the m6A modification of circ-NAB1. The interaction between circ-NAB1 and RNAs was also detected. Colony formation, transwell, flow cytometry, and western blot were utilized for measuring EC cell behaviors. Mechanically, we proved the m6A demethylase alkylation repair homolog protein 5 (ALKBH5) can mediate circ-NAB1 expression through an m6A-YTHDF2-dependent manner. Circ-NAB1 overexpression can promote cell proliferation, migration, invasion, epithelial-mesenchymal transition (EMT) process, and cell cycle through functional assays. Circ-NAB1 knockdown exerts the opposite function on EC cells. Furthermore, we proved that circ-NAB1 can sponge miR-876-3p to upregulate the target gene cyclin-dependent kinase inhibitor 3 (CDKN3) in EC cells. CDKN3 overexpression can reverse the impacts of circ-NAB1 depletion on EC cell behaviors. Collectively, we proved that ALKBH5-mediated m6A modification of circ-NAB1 promoted EMT process and cell cycle in EC via targeting the miR-876-3p/CDKN3 axis.

EZH2 inhibition dampens autoantibody production in lupus by restoring B cell immune tolerance.

The aim of this study was to elucidate the role of enhancer of zeste homolog 2 (EZH2) in the breakdown of B cell immune tolerance and production of autoantibodies in systemic lupus erythematosus (SLE), and to explore the therapeutic effects of EZH2 inhibition on lupus. Peripheral blood mononuclear cells (PBMCs) were collected from new-onset SLE patients for flow cytometric analysis. Pristane-induced lupus mice were constructed, and the EZH2 inhibitor was administrated by intraperitoneal injection to treat lupus mice. Blood and urine were collected from lupus mice to detect autoantibodies and proteinuria, and renal pathology scores were assessed. Mouse spleen B cells were sorted with magnetic beads and subjected to flow cytometric apoptosis detection, real time quantitative PCR (RT-qPCR), and western blotting (WB). EZH2 expression was elevated in diverse B-cell subsets in both SLE patients and pristane-induced lupus mice. The EZH2 inhibitor attenuated lupus-like symptoms and dampened autoantibody production in pristane-induced lupus mice. Inhibition of EZH2 also reduced autoantibody secretion by plasma cells from lupus patients. Mechanistically, EZH2 mediated the impaired apoptosis of autoreactive B cells and the differentiation of autoantibody producing plasma cells by inhibiting multiple cyclin-dependent kinase inhibitor (CKI) genes. EZH2 mediated the breakdown of B-cell peripheral immune tolerance by inhibiting CKI genes and participated in the generation of autoantibodies in SLE. EZH2 inhibition could serve as a promising drug intervention for the treatment of SLE.

IGF2 interacts with the imprinted gene Cdkn1c to promote terminal differentiation of neural stem cells.

Adult neurogenesis is supported by multipotent neural stem cells (NSCs) with unique properties and growth requirements. Adult NSCs constitute a reversibly quiescent cell population that can be activated by extracellular signals from the microenvironment in which they reside in vivo. Although genomic imprinting plays a role in adult neurogenesis through dose regulation of some relevant signals, the roles of many imprinted genes in the process remain elusive. Insulin-like growth factor 2 (IGF2) is encoded by an imprinted gene that contributes to NSC maintenance in the adult subventricular zone through a biallelic expression in only the vascular compartment. We show here that IGF2 additionally promotes terminal differentiation of NSCs into astrocytes, neurons and oligodendrocytes by inducing the expression of the maternally expressed gene cyclin-dependent kinase inhibitor 1c (Cdkn1c), encoding the cell cycle inhibitor p57. Using intraventricular infusion of recombinant IGF2 in a conditional mutant strain with Cdkn1c-deficient NSCs, we confirm that p57 partially mediates the differentiation effects of IGF2 in NSCs and that this occurs independently of its role in cell-cycle progression, balancing the relationship between astrogliogenesis, neurogenesis and oligodendrogenesis.

A Human iPSC Model Reveals Specific Upregulation of Linker Histone H1-0 in ETV6::RUNX1+ Preleukemia and B Cell Precursor Acute Lymphoblastic Leukemia

The ETV6::RUNX1 translocation is the most common chromosomal rearrangement associated with pediatric B cell precursor acute lymphoblastic leukemia (BCP-ALL). As previously shown, ETV6::RUNX1 leads to a distinct clinically silent preleukemic state in utero that manifests in impaired early B cell differentiation. Acquisition of secondary oncogenic mutations gives rise to overt leukemia in 0.2% of children carrying ETV6::RUNX1. In-depth characterization of ETV6::RUNX1+ preleukemia is essential to gain an understanding of leukemic transformation and develop preventive strategies. Here, we identify dysregulation of the epigenetic regulator H1-0 in ETV6::RUNX1+ preleukemia and BCP-ALL. Utilizing CRISPR/Cas9 genome editing, we generated monoclonal ETV6::RUNX1+ human induced pluripotent stem cell (hiPSC) lines with stable ETV6::RUNX1 expression driven by the endogenous ETV6 promoter. To assess the preleukemic setting mediated by ETV6::RUNX1, we performed RNA sequencing (RNA-seq) analysis of ETV6::RUNX1+ and matching wild-type (WT) hiPSCs of two donors. We identified 20 differentially expressed genes in ETV6::RUNX1+ hiPSCs of both donors compared to the respective WT hiPSCs (up: 15, down: 5, absolute fold change ≥ 2, p ≤ 0.05). Interestingly, our RNA-seq analysis revealed consistent upregulation of H1-0 (mean: 2.43-fold), a linker histone involved in chromatin remodeling, cell proliferation and differentiation. Elevated H1-0 levels in the ETV6::RUNX1+ hiPSC lines were confirmed by quantitative real-time PCR and immunoblot (Figure 1A). To determine H1-0 levels in ETV6::RUNX1+ B cell precursors, we analyzed published gene expression microarray and RNA-seq data sets. Both ETV6::RUNX1+ CD34+ cells (n = 10) derived from umbilical cord blood (Polak et al. Haematologica 2019) and ETV6::RUNX1+ CD34+CD19+ cells (n = 4) generated by hematopoietic differentiation of hiPSCs (Böiers et al. Developmental Cell 2018) showed significant upregulation of H1-0 compared to the respective control cells lacking ETV6::RUNX1 (unpaired t-test, p = 0.0087 and p = 0.0076, respectively). Additionally, we differentiated ETV6::RUNX1+ and WT hiPSCs into CD34+CD45+ hematopoietic progenitor cells and performed single-cell RNA-seq analysis. We identified coexpression of H1-0 with cyclin-dependent kinase inhibitor genes CDKN1A and CDKN1B, indicating a slow or non-cycling phenotype of H1-0+ cells. Next, we examined H1-0 mRNA levels in gene expression data sets of pediatric and adult leukemia patients (St. Jude PeCan database (n = 876), Jerchel et al. Haematologica 2018 (n = 654), MILE study (n = 1299)). H1-0 expression was found to be significantly elevated in ETV6::RUNX1+ BCP-ALL patient samples compared to other leukemia entities, especially TCF3::PBX1+ ALL (ordinary one-way ANOVA, p ≤ 0.0001), KMT2A rearranged ALL (p ≤ 0.0001) and chronic lymphoblastic leukemia (CLL, p ≤ 0.0001). Strikingly, H1-0 expression was 22.37-fold lower in CLL compared to ETV6::RUNX1+ BCP-ALL (Figure 1B). This observation prompted us to investigate H1-0 expression in B cell developmental stages using published data sets. Interestingly, we found that H1-0 levels decrease during early B cell development in both normal bone marrow (Black et al. Nucleic Acids Research 2018) and umbilical cord blood-derived cells (Novershtern et al. Cell 2011). Additionally, analysis of single-cell RNA-seq data of early B cell precursors derived from healthy bone marrow (Mehtonen et al. Genome Medicine 2020) and fetal liver (Developmental Cell Atlas Newcastle University) revealed that relative numbers of H1-0 expressing cells per differentiation state decreased along the pseudotime B cell development trajectory. Taken together, our study identifies H1-0 upregulation as a specific alteration present already at the ETV6::RUNX1+ preleukemic state. High H1-0 expression was associated with slower proliferation and increased stemness. This phenotype might explain the persistence of in utero generated preleukemic ETV6::RUNX1+ clones residing in the bone marrow of approximately 5% of healthy children for years. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Gene polymorphisms of cyclin-dependent kinase inhibitor and matrix metalloproteinase-9 in Sudanese patients with esophageal squamous cell carcinoma.

The polymorphisms of the cyclin-dependent kinase inhibitor (CDKN1A) gene and matrix metalloproteinase-9 (MMP9) gene may increase one's susceptibility to malignancies. In this study, the association of the single nucleotide polymorphisms (SNPs) CDKN1A rs1059234 c.70C>T at the 3' untranslated region and MMP9 rs17576 (c.836A>G, p.Gln279Arg) with esophageal squamous cell carcinoma (ESCC) in Sudanese individuals were investigated. A case-control study involving age- and gender-matched groups were conducted in a cancer center in eastern Sudan (Gadarif) between April and October 2020. The case group consisted of ESCC patients, whereas the control group comprised healthy subjects. Polymerase chain reaction-restriction fragment length polymorphism was performed for the genotyping of the CDKN1A rs1059234 and MMP9 rs17576 SNPs. The genotyping results were confirmed by Sanger sequencing. The genotype distributions for CDKN1A rs1059234 and MMP9 rs17576 were in agreement with the Hardy-Weinberg equilibrium. The variant allele T in CDKN1 rs1059234 c.70C>T was significantly more prevalent in the ESCC patients than in the healthy controls [51.3% vs. 19.2%; OR=4.4; 95% CI (2.6-7.4); p < 0.001]. Moreover, in CDKN1A rs1059234, the genotype TC + TT [76.9% vs. 38.4%; OR=5.3; 95% CI (2.6-10.7); p < 0.001] was more frequent in the cases than in the controls, and it was significantly associated with ESCC risk. In MMP9 rs17576, the variant allele G was also significantly prevalent in the cases relative to the controls, and it was significantly associated with increased ESCC risk in the cases compared with the controls [27.5% vs. 1.9%; OR=19.4; 95%CI (5.8-64.1); p < 0.001]. Both genotypes containing the allele G (AG + GG) were the most common genotypes in the cases [48.7% vs. 3.8%; OR=23.7; 95%CI (6.8-81.7); p < 0.001], and they significantly increased the risk of ESCC. A significant increase in ESCC risk is associated with the SNPs CDKN1A rs1059234 and MMP9 rs17576.

High-Level MYCN-Amplified RB1-Proficient Retinoblastoma Tumors Retain Distinct Molecular Signatures.

PurposeRetinoblastomas are malignant eye tumors diagnosed in young children. Most retinoblastomas are genetically characterized by biallelic inactivation of the RB1 gene. However, 1.5% of tumors demonstrate high-level amplification of the proto-oncogene MYCN. Patients with MYCN-amplified RB1-proficient retinoblastoma receive a diagnosis at an earlier age and show a clinically and histologically more malignant phenotype. This study aimed to identify genome-wide molecular features that distinguish this subtype from other retinoblastomas.DesignCohort study.ParticipantsForty-seven retinoblastoma tumors, comprising 36 RB1–/–, 4 RB1+/–, and 7 RB1+/+ tumors. In total, 5 retinoblastomas displayed high-level MYCN amplification, with 3 being RB1+/+, 1 being RB1+/–, and 1 being RB1–/–.MethodsIntegrated analysis, based on gene expression, methylation, and methylation-expression correlations, was performed to identify distinct molecular components of MYCN-amplified RB1-proficient retinoblastomas compared with other retinoblastoma subtypes. The methylation and methylation-expression correlation analysis was initially conducted within a subset of samples (n = 15) for which methylation profiles were available. The significant findings were cross-validated in the entire cohort (n = 47) and in publicly available data.Main Outcome MeasuresDifferentially expressed genes/pathways, differentially methylated genes, and methylation-driven differential gene expression.ResultsA large number of genes (n = 3155) were identified with distinct expression patterns in MYCN-amplified RB1-proficient retinoblastomas. The upregulated and downregulated genes were associated with translation and cell-cycle processes, respectively. Methylation analysis revealed distinct methylated patterns in MYCN-amplified RB1-proficient tumors, many of which showing significant impact on gene expression. Data integration identified a 40-gene expression signature with hypermethylated state resulting in a significant downregulation in MYCN-amplified RB1-proficient retinoblastomas. Cross-validation using the entire cohort and the public domain expression data verified the overall lower expression of these genes not only in retinoblastomas with a MYCN-amplified RB1-proficient background, but also in MYCN-amplified neuroblastomas. These include the metabolism-associated TSTD1 gene and the cyclin-dependent kinase inhibitor gene CDKN2C.ConclusionsMYCN-amplified RB1-proficient retinoblastomas display significantly distinct molecular features compared with other retinoblastomas, including a set of 40 hypermethylation-driven downregulated genes. This gene set can give insight into the biology of MYCN-amplified retinoblastomas and may help us to understand the more aggressive clinical behavior.

Loss of the transcription repressor ZHX3 induces senescence-associated gene expression and mitochondrial-nucleolar activation.

Cellular senescence is accompanied by metabolic and epigenomic remodeling, but the transcriptional mechanism of this process is unclear. Our previous RNA interference-based screen of chromatin factors found that lysine methyltransferases including SETD8 and NSD2 inhibited the senescence program in cultured fibroblasts. Here, we report that loss of the zinc finger and homeobox protein 3 (ZHX3), a ubiquitously expressed transcription repressor, induced senescence-associated gene expression and mitochondrial–nucleolar activation. Chromatin immunoprecipitation–sequencing analyses of growing cells revealed that ZHX3 was enriched at the transcription start sites of senescence-associated genes such as the cyclin-dependent kinase inhibitor (ARF-p16INK4a) gene and ribosomal RNA (rRNA) coding genes. ZHX3 expression was consistently downregulated in cells with replicative or oncogene-induced senescence. Mass spectrometry-based proteomics identified 28 proteins that interacted with ZHX3, including ATP citrate lyase and RNA metabolism proteins. Loss of ZHX3 or ZHX3-interaction partners by knockdown similarly induced the expression of p16INK4a and rRNA genes. Zhx3-knockout mice showed upregulation of p16INK4a in the testes, thymus and skeletal muscle tissues, together with relatively short survival periods in males. These data suggested that ZHX3 plays an essential role in transcriptional control to prevent cellular senescence.

Immune system parameters in chronic opisthorchiasis patients related to genes polymorphisms associated with developing ischemic heart disease

The aim of the study was to identify a relationship between gene polymorphisms associated with developing coronary heart disease and pathogenetic features of opisthorchiasis invasion. There were compared laboratory parameters assessing the immune system functioning (lymphocyte immunophenotyping evaluated by flow cytometry, serum immunoglobulin and cytokine concentrations, non-specific resistance indices) in patients with chronic opisthorchiasis (caused by Opisthorchis felineus) of various genotypes with/without mutations in loci associated with predisposition to developing coronary heart disease. It was found that opisthorchiasis invasion leads to altered immune response in patients bearing mutations in genes encoding adenosine monophosphate deaminase 1, hypoxia induced factor 1 alpha, and apolipoprotein E affecting relevant protein functions compared to those without minor alleles. Upon that, patients carrying mutation in the adenosine monophosphate deaminase 1 gene, turned out to have lowered cellular (decreased count of cytotoxic lymphocytes) and humoral immunity (decreased immunoglobulin A level). Patients with opisthorchiasis bearing a rare allele in the hypoxia-induced factor 1 alpha gene were featured with activated adaptive immunity and reduced activation of innate immunity. Moreover, mutated apolipoprotein E gene was coupled to decreased activity of cellular immune response, which exacerbates the course of opisthorchiasis invasion, whereas minor allele of the rs1333049 polymorphism in the cyclin-dependent kinase inhibitor gene results in no significantly altered immune response. Thus, gene-related dysfunction of certain proteins may allow opisthorchiasis invasion to elicit multidirectional changes in immune response, so that approaching the normal range of diverse immune-related parameters during chronic parasitic invasion may not necessarily be interpreted as a “protective” effect exerted by any of gene polymorphisms. The most prominent changes in immune response of patients with chronic opisthorchiasis were detected in case of polymorphism in the factor-induced hypoxia-1 alpha gene. The data obtained suggest that the aforementioned genetic polymorphisms may predispose to manifestation of certain opisthorchiasis clinical forms.

The SNP rs7865618 of 9p21.3 locus emerges as the most promising marker of coronary artery disease in the southern Indian population

Development of coronary artery disease (CAD) is primarily due to the process of atherosclerosis, however the prognosis of CAD depends on pleiotropic effects of the genes located at 9p21.3 region. Genome wide association studies revealed association of variants in this region with CAD pathology. However, specific marker in predicting CAD development or progression is not yet identified. In the present study, 35 SNPs at 9p21.3 region, located in the cyclin dependent kinase inhibitor (CDKN2A/CDKN2B) genes, were genotyped among 350 CAD cases and 480 controls from the southern Indian population of Hyderabad using fluidigm nanofluidic SNP genotyping system and the data were analyzed using PLINK and R softwares. Of the 35 SNPs analysed, only one SNP, rs7865618, was found to be highly significantly associated with CAD, even after correction for multiple testing (p = 0.008). The AG and GG genotypes of this SNP conferred 3.08 and 1.93 folds increased risk for CAD respectively. In particular, this SNP was significantly associated with severe anatomic (triple vessel disease p = 0.023) and phenotypic (acute coronary syndrome p = 0.007) categories of CAD. Pair wise SNP interaction analysis between the SNPs of 9p21.3 and 11q23.3 regions revealed significantly increased risk of three SNPs of 11q23.3 region that were not associated individually, in conjunction with rs7865618 of 9p21.3.

MDS1-EVI1 Complex (MECOM) Activation Promotes Leukemic Stem Cell Quiescence By Upregulating CDKN1C/P57Kip2

Background: Activation of MDS1-EVI1 Complex (MECOM) via proviral insertion at its gene locus (Morishita et al., 1988) or through chromosomal rearrangements at 3q26 (Suzukawa et al., 1994; Tang et al., 2019) results in marked overexpression of EVI1, and is predominantly associated with myeloid malignancies (Secker-Walker et al., 1995), especially high-risk acute myeloid leukemia (AML) (Barjesteh van Waalwijk van Doorn-Khosrovani et al., 2003). We previously showed that EVI1 overexpression skews hematopoiesis toward the myeloid lineage via the upregulation of a master regulator of myelopoiesis PU.1 encoded by SPI1 (SFPI1) (Ayoub et al., 2018), which explains the high cooccurrence of 3q abnormalities in myeloid malignancies. However, the role of EVI1 in reduced survival and high rates of relapse in AML patients is still unknown, and therapies specific for EVI1+ AML are absent due to the lack of EVI1 specific targets. Here we describe a novel mechanism of action of EVI1 in controlling leukemic stem cell quiescence by upregulating an essential member of the cyclin-dependent kinase inhibitor (cip/kip) gene family: CDKN1C/P57Kip2. Results: We first identified the upregulation of CDKN1C in the presence of EVI1 overexpression using RNA-seq on sorted hematopoietic stem and progenitor leukemia cells from our previously published EVI1+ leukemia mouse model (EVI1TO) (Ayoub et al., 2018). Since CDKN1C/ P57 expression has been linked to high relapse rates in AML patients following chemotherapy (Radujkovic et al., 2016), we confirmed the upregulation of CDKN1C using RNA-seq on sorted populations (mononucleated cells (MNCs), leukemia stem cells (LSCs), and minimal residual disease (MRD)) from activated-MECOM AML patients in comparison to nonactivated-MECOM high risk AML patients (67 samples from 27 patients). Additionally, we found EVI1 overexpression produces a reversible block in differentiation and engraftment that can be rescued by ablating EVI1 overexpression or by additional mutations. In an effort to identify EVI1-induced transcriptional regulations for CDKN1C, we performed chromatin immunoprecipitation and sequencing (ChIP-seq) and assayed for transposase accessible Chromatin (ATAC-seq) using our in vivo and in vitro EVI1 overexpression models. ChIP-seq for EVI1 showed a binding site for EVI1 located 300 kb from CDKN1C transcription start site (TSS), and ATAC-seq in EVI1 overexpressing AML showed an open chromatin pattern at the CDKN1C TSS only in the presence of EVI1 overexpression. We are presently investigating the molecular mechanism involved in the transcriptional regulation of CDKN1C in the presence of EVI1 overexpression. Conclusions: Our data suggest a correlation between EVI1 and CDKN1C expression in high risk AML with 3q abnormalities and provide insights into a potential molecular mechanism for MECOM activation in controlling leukemic stem cell quiescence and resistance to chemotherapy by upregulating CDKN1C. Disclosures Andreeff: Daiichi-Sankyo; Jazz Pharmaceuticals; Celgene; Amgen; AstraZeneca; 6 Dimensions Capital: Consultancy; Daiichi-Sankyo; Breast Cancer Research Foundation; CPRIT; NIH/NCI; Amgen; AstraZeneca: Research Funding; Centre for Drug Research &amp; Development; Cancer UK; NCI-CTEP; German Research Council; Leukemia Lymphoma Foundation (LLS); NCI-RDCRN (Rare Disease Clin Network); CLL Founcdation; BioLineRx; SentiBio; Aptose Biosciences, Inc: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding.

The CDK Inhibitor SIAMESE Targets Both CDKA;1 and CDKB1 Complexes to Establish Endoreplication in Trichomes.

Endoreplication, also known as endoreduplication, is a modified cell cycle in which DNA is replicated without subsequent cell division. Endoreplication plays important roles in both normal plant development and in stress responses. The SIAMESE (SIM) gene of Arabidopsis (Arabidopsis thaliana) encodes a cyclin-dependent kinase (CDK) inhibitor that plays a central role in establishing endoreplication, and is the founding member of the SIAMESE-RELATED (SMR) family of plant-specific CDK inhibitor genes. However, there has been conflicting evidence regarding which specific cyclin/CDK complexes are inhibited by SIM in vivo. In this work, we use genetic evidence to show that SIM likely inhibits both CDKA;1- and CDKB1-containing CDK complexes in vivo, thus promoting endoreplication in developing Arabidopsis trichomes. We also show that SIM interacts with CYCA2;3, a binding partner of CDKB1;1, via SIM motif A, which we previously identified as a CDK-binding motif. By contrast, SIM motif C, which has been indicated as a cyclin binding motif in other contexts, appears to be relatively unimportant for interaction between SIM and CYCA2;3. Together with earlier results, our work suggests that SIM and other SMRs likely have a multivalent interaction with CYC/CDK complexes.

Selected CDKN2A and MDM2 polymorphisms in oral cavity cancer.

The head and neck squamous cell carcinoma (HNSCC) is an aggressive human malignancy diagnosed in more than 800 000 new cases worldwide, and mostly arises in the oral cavity, oropharynx, larynx, hypopharynx, and nasopharynx. The study presented here aimed to determine a possible association of rs11515 and rs3088440 gene polymorphisms in the CDKN2A gene (cyclin dependent kinase inhibitor 2A), as well as rs769412 and rs937283 in the MDM2 gene (murine double minute 2), with oral cavity cancer in a sample of Polish population; CDKN2A is crucial in regulating the cell cycle while MDM2 is an oncogene involved in regulating cell proliferation and apoptosis. The study included 95 primary tumor samples following surgical resection from patients, and the control group consisted of 100 healthy individuals. DNA samples were genotyped by employing the 5' nuclease assay for allelic discrimination using TaqMan SNP Genotyping Assays (Applied Biosystems, USA). There was no significant association between any of the polymorphism (rs11515, rs3088440, rs769412 and rs937283) and the oral cavity cancer risk. We found that the AA homozygotes for rs3088440 were significantly more frequent in the control group (OR=0.046, p<0.0001). In addition, the GG genotype of rs769412 was not found in any group. We found no influence of the examined genotypes on clinicopathological parameters, such as T, N and grading values in patients with oral cavity cancer. The results of this study indicate that none of the investigated polymorphisms were associated with the risk of oral cavity cancer in the examined sample of the Polish population.

Cyclin-Dependent Kinase Inhibitor Gene TaICK1 acts as a Potential Contributor to Wheat Male Sterility induced by a Chemical Hybridizing Agent.

Heterosis has been widely accepted as an effective strategy to increase yields in plant breeding. Notably, the chemical hybridization agent SQ-1 induces male sterility in wheat, representing a critical potential tool in hybrid seed production. However, the mechanisms underlying the male sterility induced by SQ-1 still remain poorly understood. In this study, a cyclin-dependent kinase inhibitor gene, TaICK1, which encodes a 229 amino acid protein, was identified as a potential contributor to male sterility in common wheat. The expression of TaICK1 was upregulated during the development of anthers in Xinong1376 wheat treated with SQ-1. Meanwhile, the seed setting rate was found to be significantly decreased in TaICK1 transgenic rice. Furthermore, we identified two cyclin proteins, TaCYCD2;1 and TaCYCD6;1, as interactors through yeast two-hybrid screening using TaICK1 as the bait, which were validated using bimolecular fluorescence complementation. Subcellular localization revealed that the proteins encoded by TaICK1, TaCYCD2;1, and TaCYCD6;1 were localized in the cell nucleus. The expression levels of TaCYCD2;1 and TaCYCD6;1 were lower in Xinong1376 treated with SQ-1. A further analysis demonstrated that the expression levels of OsCYCD2;1 and OsCYCD6;1 were lower in transgenic TaICK1 rice lines as well. Taken together, these results suggest that the upregulation of TaICK1, induced by SQ-1, may subsequently suppress the expression of TaCYCD2;1 and TaCYCD6;1 in anthers, resulting in male sterility. This study provides new insights into the understanding of SQ-1-induced wheat male sterility, as well as the developmental mechanisms of anthers.

C-Myc Inhibitor 10074-G5 Induces Murine and Human Hematopoietic Stem and Progenitor Cell Expansion and HDR Modulator Rad51 Expression.

c-Myc plays a major role in the maintenance of glycolytic metabolism and hematopoietic stem cell (HSC) quiescence. Targeting modulators of HSC quiescence and metabolism could lead to HSC cell cycle entry with concomitant expansion. Here we show that c-Myc inhibitor 10074-G5 treatment leads to 2-fold increase in murine LSKCD34low HSC compartment post 7 days. In addition, c-Myc inhibition increases CD34+ and CD133+ human HSC number. c-Myc inhibition leads to downregulation of glycolytic and cyclindependent kinase inhibitor (CDKI) gene expression ex vivo and in vivo. In addition, c-Myc inhibition upregulates major HDR modulator Rad51 expression in hematopoietic cells. Besides, c-Myc inhibition does not alter proliferation kinetics of endothelial cells, fibroblasts or adipose-derived mesenchymal stem cells, however, it limits bone marrow derived mesenchymal stem cell proliferation. We further demonstrate that a cocktail of c-Myc inhibitor 10074-G5 along with tauroursodeoxycholic acid (TUDCA) and i-NOS inhibitor L-NIL provides a robust HSC maintenance and expansion ex vivo as evident by induction of all stem cell antigens analyzed. Intriguingly, the cocktail of c-Myc inhibitor 10074-G5, TUDCA and L-NIL improves HDR related gene expression. These findings provide tools to improve ex vivo HSC maintenance and expansion, autologous HSC transplantation and gene editing through modulation of HSC glycolytic and HDR pathways.

Role of p15(INK4B) Methylation in Patients With Myelodysplastic Syndromes: A Systematic Meta-Analysis

BackgroundTumor suppressor gene cyclin-dependent kinase inhibitor 2B (p15(INK4B)) methylation has been frequently reported in myelodysplastic syndromes (MDS). However, the association between p15(INK4B) methylation and MDS remains elusive. Thus, this meta-analysis was first conducted to evaluate the clinical significance of p15(INK4B) methylation in MDS. Materials and MethodsEligible studies were identified via an online electronic databases search. The overall odds ratios (ORs) or hazard ratios (HRs) and 95% confidence intervals (CIs) were calculated. ResultsTwenty-eight studies published between 1997 and 2017 were identified, including 1205 MDS patients and 243 nontumor controls. No evidence of heterogeneity was found in our study. p15(INK4B) methylation was significantly elevated in MDS compared with nontumor controls (OR, 10.37; P < .001). In addition, p15(INK4B) methylation was significantly higher in advanced MDS than in early MDS (OR, 4.70; P < .001) and was linked to an unfavorable overall survival (multivariate analysis: HR, 1.78; 95% CI, 1.23-2.71). Subgroup analyses on the basis of ethnicity and detection method showed that the results remained significant in different subgroups (all Ps < .05). ConclusionOur findings suggest that p15(INK4B) methylation might play an important role in the development, progression, and poor prognosis of MDS. More prospective studies with larger study populations are needed.

A Polymorphic Variant in p19Arf Confers Resistance to Chemically Induced Skin Tumors by Activating the p53 Pathway

Identification of the specific genetic variants responsible for the increased susceptibility to familial or sporadic cancers is important. Using a forward genetics approach to map such loci in a mouse skin cancer model, we previously identified a strong genetic locus, Stmm3, conferring resistance to chemically induced skin papillomas on chromosome 4. Here, we report the cyclin-dependent kinase inhibitor gene Cdkn2a/p19Arf as a major responsible gene for the Stmm3 locus. We provide evidence that the function of Stmm3 is dependent on p53 and that p19ArfMSM confers stronger resistance to papillomas than p16Ink4aMSMinvivo. In addition, we found that genetic polymorphism in p19Arf between a resistant strain, MSM/Ms (Val), and a susceptible strain, FVB/N (Leu), alters the susceptibility to papilloma development, malignant conversion, and the epithelial-mesenchymal transition. Moreover, we demonstrated that the p19ArfMSM allele more efficiently activates the p53 pathway than the p19ArfFVB allele invitro and invivo. Furthermore, we found polymorphisms in CDKN2A in the vicinity of a polymorphism in mouse Cdkn2a associated with the risk of human cancers in the Japanese population. Genetic polymorphisms in Cdkn2a and CDKN2A may affect the cancer risk in both mice and humans.

Analysis of Activating GCM2 Sequence Variants in Sporadic Parathyroid Adenomas.

Sporadic, solitary parathyroid adenoma is the most common cause of primary hyperparathyroidism (PHPT). Apart from germline variants in certain cyclin-dependent kinase inhibitor genes and occasionally in MEN1, CASR, or CDC73, little is known about possible genetic variants in the population that may confer increased risk for development of typical sporadic adenoma. Transcriptionally activating germline variants, especially within in the C-terminal conserved inhibitory domain (CCID) of glial cells missing 2 (GCM2), encoding a transcription factor required for parathyroid gland development, have recently been reported in association with familial and sporadic PHPT. To evaluate the potential role of specific GCM2 activating variants in sporadic parathyroid adenoma. Regions encoding hyperparathyroidism-associated, activating GCM2 variants were PCR amplified and sequenced in genomic DNA from 396, otherwise unselected, cases of sporadic parathyroid adenoma. Activating GCM2 CCID variants (p.V382M and p.Y394S) were identified in six of 396 adenomas (1.52%), and a hyperparathyroidism-associated GCM2 non-CCID activating variant (p.Y282D) was found in 20 adenomas (5.05%). The overall frequency of tested activating GCM2 variants in this study was 6.57%, approximately threefold greater than their frequency in the general population. The examined, rare CCID variants in GCM2 were enriched in our cohort of patients and appear to confer a moderately increased risk of developing sporadic solitary parathyroid adenoma compared with the general population. However, penetrance of these variants is low, suggesting that the large majority of individuals with such variants will not develop a sporadic parathyroid adenoma.

Adipose Tissue Senescence and Inflammation in Aging is Reversed by the Young Milieu.

Visceral adipose tissue (VAT) inflammation plays a central role in longevity and multiple age-related disorders. Cellular senescence (SEN) is a fundamental aging mechanism that contributes to age-related chronic inflammation and organ dysfunction, including VAT. Recent studies using heterochronic parabiosis models strongly suggested that circulating factors in young plasma alter the aging phenotypes of old animals. Our study investigated if young plasma rescued SEN phenotypes in the VAT of aging mice. With heterochronic parabiosis model using young (3 months) and old (18 months) mice, we found significant reduction in the levels of pro-inflammatory cytokines and altered adipokine profile that are protective of SEN in the VAT of old mice. These data are indicative of protection from SEN of aging VAT by young blood circulation. Old parabionts also exhibited diminished expression of cyclin-dependent kinase inhibitors (CDKi) genes p16 (Cdkn2a) and p21 (Cdkn1a/Cip1) in the VAT. In addition, when exposed to young serum condition in an ex vivo culture system, aging adipose tissue-derived stromovascular fraction cells produced significantly lower amounts of pro-inflammatory cytokines (MCP-1 and IL-6) compared to old condition. Expressions of p16 and p21 genes were also diminished in the old stromovascular fraction cells under young serum condition. Finally, in 3T3-preadipocytes culture system, we found reduced pro-inflammatory cytokines (Mcp-1 and Il-6) and diminished expression of cyclin-dependent kinase inhibitor genes in the presence of young serum compared to old serum. In summary, this study demonstrates that young milieu is capable of protecting aging adipose tissue from SEN and thereby inflammation.