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Upstream regulator of genomic imprinting in rice endosperm is a small RNA-associated chromatin remodeler
No full textGenomic imprinting is observed in endosperm, a placenta-like seed tissue, where transposable elements (TEs) and repeat-derived small RNAs (sRNAs) mediate epigenetic changes in plants. In imprinting, uniparental gene expression arises due to parent-specific epigenetic marks on one allele but not on the other. The importance of sRNAs and their regulation in endosperm development or in imprinting is poorly understood in crops. Here we show that a previously uncharacterized CLASSY (CLSY)-family chromatin remodeler named OsCLSY3 is essential for rice endosperm development and imprinting, acting as an upstream player in the sRNA pathway. Comparative transcriptome and genetic analysis indicated its endosperm-preferred expression and its likely paternal imprinted nature. These important features are modulated by RNA-directed DNA methylation (RdDM) of tandemly arranged TEs in its promoter. Upon perturbation of OsCLSY3 in transgenic lines, we observe defects in endosperm development and a loss of around 70% of all sRNAs. Interestingly, well-conserved endosperm-specific sRNAs (siren) that are vital for reproductive fitness in angiosperms are also dependent on OsCLSY3. We observed that many imprinted genes and seed development-associated genes are under the control of OsCLSY3. These results support an essential role of OsCLSY3 in rice endosperm development and imprinting, and propose similar regulatory strategies involving CLSY3 homologs among other cereals
REPRESSOR OF UV-B PHOTOMORPHOGENESIS proteins target ABSCISIC ACID INSENSITIVE 5 for degradation to promote early plant development
No full textREPRESSOR OF UV-B PHOTOMORPHOGENESIS 1 (RUP1) and REPRESSOR OF UV-B PHOTOMORPHOGENESIS 2 (RUP2) are WD-40 domain-containing proteins that have been extensively characterized for their role in UV-B signaling. However, the roles of the RUP proteins outside the canonical UV-signaling pathway are less known. Here, we identify that RUP1 and RUP2 play important roles in ABA signaling to regulate seed germination and early seedling development in Arabidopsis thaliana. Our protein interaction studies confirmed that RUP1 and RUP2 physically interact with ABA INSENSITIVE 5 (ABI5). In the presence of abscisic acid, rup1, rup2, and rup1rup2 exhibited reduced germination and seedling establishment compared with the wild type. Germination and seedling establishment in rup1rup2abi5-8 were similar to abi5-8, suggesting that RUP1 and RUP2 suppress ABA-mediated inhibition of germination and early seedling development in an ABI5-dependent manner. The DDB1-binding WD40 protein RUP2 promoted the ubiquitination of ABI5 to regulate its degradation. ABI5, in turn, establishes a negative feedback loop to inhibit the expression of RUP1/RUP2. ABI5 also inhibited the direct binding of ELONGATED HYPOCOTYL 5 (HY5) to the promoters of RUP1 and RUP2 under ABA. This study highlights the coordinated action of RUP1, RUP2, ABI5, and HY5 in regulating early plant development
Curcumin-based nanoformulation for the pyroptotic death of mda-mb-231 cells
No full textCurcumin (CU), a diphenolic natural flavonoid, has been considered as a next-generation anticancer agent; however, its efficacy is hindered by poor selectivity and low bioavailability (<3 × 10–8 μM). Nanoformulation becomes a promising strategy for the target-specific delivery of flavonoid, wherein nanoformulation itself exhibits multiple therapeutic and diagnostic modalities. Presently, we have developed CU and Prussian blue nanocube (PBNC)-based nanoformulation which shows chemo-phototoxicity against triple-negative breast cancer (TNBC, ca. MDA-MB-231 cell line) but possesses compatibility against normal HEK-293 cells up to a concentration of ∼42 ± 2.1 μg/mL. We have validated the dual-mode T1–T2 weighted magnetic resonance imaging (MRI) capability of the nanoformulation having relaxivity parameters r1 ∼ 6.01 mM–1 s–1 and r2 ∼ 15.89 mM–1 s–1 under a clinical magnetic field of 3 T, higher than relaxivity parameters of commercially available MRI contrast agents [MAGNEVIST (r1 ∼ 3.1 mM–1 s–1, r2 ∼ 3.7 mM–1 s–1), GADOVIST (r1 ∼ 3.2 mM–1 s–1, r2 ∼ 3.9 mM–1 s–1), and PROHANCE (r1 ∼ 2.8 mM–1 s–1, r2 ∼ 3.2 mM–1 s–1 etc.]. Meanwhile, the size dependence of relaxivity parameters is dependent on crystal inhomogeneity and magnetization. In the therapeutic part, we have noticed oxidative stress-induced pyroptotic cell death, indexed by the expression of series of proteins including P2RX7-CASPASE1 and IL-1β-NLRP3. Smaller-sized particles show better theranostic activity due to easy internalization. In summary, our study suggests that PBNC (∼60 nm) is highly beneficial to deliver CU, while our conceptual study clearly demonstrates the role of size and functionalization on its theranostic properties
Cholesterol-driven modulation of membrane-membrane interactions by an antimicrobial peptide, NK-2, in phospholipid vesicles
No full textAntimicrobial peptides (AMPs) are essential components of the innate immune system, demonstrating their antimicrobial effects primarily through the creation of transmembrane pores that result in membrane disruption. Cholesterol within the membrane can significantly affect the interaction between AMPs and the membrane, as it is known to alter both the permeability and elastic properties of the membrane. In this study, we have investigated the influence of cholesterol on the interaction of the AMP, NK-2 with phospholipid vesicles. We prepared giant unilamellar vesicles (GUVs) composed of DOPC-DOPG and Egg PC, varying the cholesterol concentrations, and analyzed them using phase contrast microscopy. The aggregation of vesicles is evident in the phase contrast microscopy observations of GUVs. The aggregation of GUVs with cholesterol ultimately leads to a collapse state, a condition not typically seen in GUVs lacking cholesterol. Furthermore, the aggregation kinetics were determined from the analysis of phase contrast micrographs. This biophysical investigation offers valuable insights into how cholesterol affects the interactions between membranes induced by antimicrobial peptides
Multifunctional suture coating for combating surgical site infections and mitigating associated complications
No full textDespite advancements in preventive measures and hospital protocols, surgical site infections (SSIs) remain a significant concern following surgeries. Sutures, commonly used for wound closure, can serve as a platform for microbial adherence and contamination, leading to extensive debridement and recurrent antibiotic therapy. The emergence of drug resistance and the formation of biofilms on sutures have further complicated the management of SSIs. Drug-eluting sutures incorporating biocides like triclosan have limitations due to uncontrolled release and associated toxicity. Therefore, there is a need for alternative approaches to impart antimicrobial properties to sutures. In this study, we present a one-step covalent cross-linking method to coat surgical sutures with an antimicrobial small molecule, quaternary benzophenone-based antimicrobial (QSM). Additionally, the sutures are dip-coated with ibuprofen, a nonsteroidal anti-inflammatory drug with analgesic properties. The coated sutures maintained their morphological and tensile properties after in vivo implantation. The antimicrobial coating demonstrated efficacy against a broad-spectrum pathogens, including drug-resistant bacteria and fungi. The optimized formulation retained its biodegradability in vivo. Furthermore, the coated sutures exhibited ∼3 log reduction in methicillin-resistant Staphylococcus aureus (MRSA) burden in a subcutaneous implantation mouse model. Overall, this multifunctional coating provides antimicrobial properties to surgical sutures while preserving their mechanical integrity and biodegradability. These coated sutures have the potential to address the challenge of SSIs and contribute to improved surgical outcomes
The behaviour of pile group and combined piled-raft foundation in liquefiable soil under seismic conditions.
No full textThis paper highlights the beneficial usage of Combined Pile-Raft Foundation (CPRF) over conventional pile group foundation subjected to seismic loading in liquefiable soil. Firstly, a single pile resting on a liquefiable soil is numerically modelled and subsequently validated with available dynamic centrifuge test result by using finite difference based computer programme, FLAC3D. Thereafter, the model is extended for simulating CPRF and pile group.Further parametric studies are performed to understand the effect of pile spacing (s), pile length (l) and different seismic motions on the behaviour of CPRF and pile group. Results are presented in terms of normalised bending moment (M/Mmax), shear forces and pore water pressure (PWP) ratio. Increase in shear resistances in the range of (35 – 60)% and (40 – 70)% are observed for the piles in CPRF over the conventional pile group foundation, having a pile spacing of 2 to 5 times of its diameter (d) and the (l/d) of 14 to 20, respectively. These outcomes portray the advantages of employing CPRF over pile group founded in liquefiable area under seismic loading
Deciphering the intricate relationship between macrophages, pigmentation, and prognosis in uveal melanoma
No full textHigh pigmentation and the abundance of M2 macrophages have been identified as negative predictors in uveal melanoma (UM). Risk factors associated with UM that are prevalent in high-risk White populations are still present, although less common, in relatively low-risk Asian populations. Research indicates that proangiogenic M2 macrophages and monosomy 3 play significant roles in UM progression. Our aim was to investigate the impact of tumor-associated macrophages in UM and examine their correlation with monosomy 3 and pigmentation. Transmission electron microscopy was used to analyze the morphology of macrophages in UM. Forty UM samples underwent fluorescent in situ hybridization for monosomy 3 identification. Immunohistochemistry was done to assess M2/M1 macrophages on 82 UM tissue samples. IL-10 and IL-12 expressions were quantified in UM serum samples by enzyme-linked immunosorbent assay. The expression of all markers was correlated with pigmentation markers (tyrosinase-related protein 1, tyrosinase-related protein 2, silver protein, and microphthalmia-associated transcription factor). Prognostic outcomes were determined using the Cox proportional hazard model and log-rank tests. Increased expression of M2/M1 macrophages was observed in 31 UM cases, which correlated with the high expression of pigmentation markers. IL-10 concentration was high in UM cases. Monosomy 3 was evident in 50% of UM cases and significantly associated with increased immunoexpression of M2/M1 macrophages and pigmentation markers. Reduced metastasis-free survival was observed in patients with UM with high M2/M1 macrophage expression (P = .001). High pigmentation and increased M2 macrophage density could impact the tumor microenvironment in UM. This could contribute to ineffective antitumor immune responses in patients with UM. Our findings suggest avenues for developing novel therapeutic approaches to counteract these immunosuppressive effects in UM
Aml-013 a prospective study to evaluate the prognostic implications and molecular mechanism of slc40a1 gene in primary acute myeloid leukemia
No full textContext
Acute myeloid leukemia (AML) is a genetically heterogeneous hematologic disorder characterized by abnormal myeloid progenitor cell proliferation and differentiation, often accompanied by genetic and epigenetic alterations. Iron metabolism is implicated in various cancers, yet its role in AML remains poorly understood. Ferroportin, encoded by the SLC40A1 gene, is vital for cellular iron export, but its significance and interactions in AML are unclear.
Objective
This study aimed to characterize the molecular functions, clinical relevance, and prognostic value of the SLC40A1 gene in AML.\ud
Methods
We investigated SLC40A1 gene expression in AML (n=173) and control (n=70) cases, followed by correlation analysis to identify associated genes using the Linked Omics database. Prognostic significance was evaluated using Kaplan–Meier survival estimation. DNA methylation status was assessed using the MEXPRESS database. Molecular mechanisms were explored via gene set enrichment analysis (GSEA), and the relationship with immune checkpoints was studied using the SANGER Box 3.0 database.
Results
Laboratory Oncology Unit, Dr. B.R.A. Institute Rotary Cancer Hospital, All India Institute of Medical Sciences New Delhi, India SLC40A1 mRNA was significantly overexpressed in AML cohorts and correlated with poor overall survival (P<0.05). Positive correlations were found with DNAJC6, CD59, PLS1, GCLM, and CAPRIN2 genes, while negative correlations were observed with MSLN, MYH11, DAGLB, ST18, and PLCD3 (PCC <0.80). Lower methylation levels of SLC40A1 were observed in AML, inversely associated with gene expression. Gene enrichment analysis revealed enrichment in biologic processes such as lymphocyte homeostasis, endothelium development, spleen development, cell communication, and cell differentiation, while in molecular function SLC40A1 gene was enriched in iron ion transmembrane transporter, metal cluster binding, growth factor binding, and catalytic activity. KEGG pathway analysis implicated SLC40A1 in critical pathways like hematopoietic stem cell differentiation, ferroptosis, mRNA surveillance pathway, autophagy, TGF-beta signaling pathway, and signaling pathways regulating pluripotency of stem cells. Additionally, SLC40A1 positively correlated with immune checkpoints including CD160, CD274, CD40, CD44, and CD80.
Conclusions
SLC40A1 plays a significant role in AML progression and may serve as a potential prognostic biomarker and therapeutic target. This study provides insights into the molecular mechanisms underlying SLC40A1 involvement in AML pathogenesis, offering avenues for novel therapeutic interventions
AML-018 Exploring Potential Molecular Dependency of t(8;21)(q22;q22.1) Positive Acute Myeloid Leukemia on the Non-Coding RNA HOTAIRM1-miR222 Axis
No full textIntroduction
Acute myeloid leukemia (AML) is a heterogenous disease categorized into subtypes based on characteristic chromosomal translocations, each having distinct biology. Patients with translocation t(8;21)(q22;q22.1) undergoing conventional chemotherapy show an overall survival of around 50%. Studying subtype-specific biology is crucial to developing targeted therapies and improving patient outcomes. HOTAIRM1, a long non-coding RNA regulates key genes during hematopoiesis, and its expression increases as myeloid cells differentiate. In AML, HOTAIRM1 is deregulated, and we aimed to study the significance of this aberrant expression.
Materials and Methods
We performed quantitative polymerase chain reaction (qPCR) to study HOTAIRM1 expression in the bone marrow of patients with AML (n=48), and six AML cell lines (Kasumi-1, THP1, HL60, KG1, MOLM13, and MOLM14). Overexpression was achieved by cloning HOTAIRM1 into pcDNA3.1+ vector followed by transfection. Cell cycle and apoptosis assays were performed using flow cytometry. MTT assay was used to calculate inhibitory concentration 50 (IC50) values.
Results
We found that HOTAIRM1 was significantly downregulated in patients with AML (P=.0005) and in AML cell lines (P<.01), especially in the t(8;21) positive subgroup. Overexpressing HOTAIRM1 led to cell cycle arrest (P<.01) and increased apoptosis (P<.01) in t(8;21) positive Kasumi-1 cells, indicating specific regulation of HOTAIRM1 in this subtype. We identified over-expressed microRNA miR-222 as a potential regulator of HOTAIRM1 expression in Kasumi-1. High miR-222 expression was associated with significantly lower overall survival (P=.02). Inhibiting miR-222 in Kasumi-1 mirrored the effect of HOTAIRM1 overexpression, resulting in cell cycle arrest (P<.001) mediated by p27 protein upregulation and increased apoptosis (P<.001) via BCL2, BCLXL, and MCL1 proteins. Inhibiting miR-222 however, did not increase HOTAIRM1 expression. Interestingly, HOTAIRM1 overexpression in Kasumi-1 led to reduced miR-222 (P<.05) indicating HOTAIRM1’s role in regulating miR-222, not vice versa. Treating Kasumi-1 cells with the hypomethylating drug azacytidine increased HOTAIRM1 expression (P<.0001) suggesting that aberrant promoter hypermethylation contributes to low HOTAIRM1 expression in the t(8;21) positive subtype.
Conclusion
Our data indicate that rescuing HOTAIRM1 expression in Kasumi-1 cells with drugs such as azacytidine, leads to inhibition of leukemic phenotypes driven by oncogenic miR-222. This suggests that targeting the non-coding RNA HOTAIRM1-miR-222 pathway could be a novel therapeutic approach in t(8;21) AML
Refinement of risk-stratification of cytogenetically normal acute myeloid leukemia adult patients by mn1 expression
No full textIntroduction: Acute myeloid leukemia with normal cytogenetics (CN-AML) represents a heterogeneous group having diverse genetic mutations. Understanding the significance of each of these mutations is necessary. In this study, we evaluated the prognostic role of MN1 expression in adult CN-AML patients. Method: One hundred and sixty-three de-novo adult AML patients were evaluated for MN1 expression by real-time PCR. MN1 expression was correlated with the clinical characteristics of the patients and their outcomes. Results: Higher MN1 expression was associated with NPM1 wild-type (p<0.0001), CD34 positivity (p=0.006), and lower clinical remission rate (p=0.027). FLT3-ITD and CEBPA mutations had no association with MN1 expression. On survival analysis, a high MN1 expression was associated with poor event-free survival (Hazard Ratio 2.47, 95% Confidence Interval: 1.42-4.3; p<0.0001) and overall survival (Hazard Ratio 4.18, 95% Confidence Interval: 2.17-8.08; p<0.0001). On multivariate analysis, the MN1 copy number emerged as an independent predictor of EFS (p<0.0001) and OS (p<0.0001). Conclusion: MN1 expression is an independent predictor of outcome in CN-AML