Downregulation Of Transcription Factors Research Articles

Sildenafil Reverses the Neuropathological Alzheimer's Disease Phenotype in Cholinergic-Like Neurons Carrying the Presenilin 1 E280A Mutation.

Familial Alzheimer's disease (FAD) presenilin 1 E280A (PSEN 1 E280A) is characterized by functional impairment and the death of cholinergic neurons as a consequence of amyloid-β (Aβ) accumulation and abnormal phosphorylation of the tau protein. Currently, there are no available therapies that can cure FAD. Therefore, new therapies are urgently needed for treating this disease. To assess the effect of sildenafil (SIL) on cholinergic-like neurons (ChLNs) harboring the PSEN 1 E280A mutation. Wild-type (WT) and PSEN 1 E280A ChLNs were cultured in the presence of SIL (25μM) for 24 h. Afterward, proteinopathy, cell signaling, and apoptosis markers were evaluated via flow cytometry and fluorescence microscopy. We found that SIL was innocuous toward WT PSEN 1 ChLNs but reduced the accumulation of intracellular Aβ fragments by 87%, decreased the non-physiological phosphorylation of the protein tau at residue Ser202/Thr205 by 35%, reduced the phosphorylation of the proapoptotic transcription factor c-JUN at residue Ser63/Ser73 by 63%, decreased oxidized DJ-1 at Cys106-SO3 by 32%, and downregulated transcription factor TP53 (tumor protein p53), BH-3-only protein PUMA (p53 upregulated modulator of apoptosis), and cleaved caspase 3 (CC3) expression by 20%, 32%, and 22%, respectively, compared with untreated mutant ChLNs. Interestingly, SIL also ameliorated the dysregulation of acetylcholine-induced calcium ion (Ca2+) influx in PSEN 1 E280A ChLNs. Although SIL showed no antioxidant capacity in the oxygen radical absorbance capacity and ferric ion reducing antioxidant power assays, it might function as an anti-amyloid and antiapoptotic agent and functional neuronal enhancer in PSEN 1 E280A ChLNs. Therefore, the SIL has therapeutic potential for treating FAD.

Abstract 4775: Unveiling caveolin-1's crucial role in breast cancer progression and metastasis: A molecular perspective

Abstract Background: Breast cancer, particularly its metastatic form, constitutes a significant portion of oncological mortality. Caveolin-1 (Cav-1), a caveolae structural protein, has been implicated in cellular processes such as proliferation, apoptosis, autophagy, and cellular motility (aberrant in breast cancer). With its dual role as a tumor suppressor and promoter, Cav-1's function in breast cancer pathophysiology requires elucidation for potential therapeutic targeting. Methods: We utilized a syngeneic mouse model to engineer Cav-1 deficient 4T1 murine mammary carcinoma cells which mirror the characteristics of stage IV breast cancer to investigate Cav-1's role in tumor growth and metastasis. We assessed cellular proliferation and lung metastasis, compared with wild-type controls, and performed molecular analyses to evaluate epithelial and mesenchymal marker expression, stemness-related markers, and autophagy-related gene expression. Results: Cav-1 ablation corresponded with a marked decrease in tumor proliferation and lung metastasis, indicating a suppressive role in tumor progression. Molecular characterization revealed downregulated epithelial markers (E-cadherin, β-catenin), transcription factors (TWIST1/2), and stem cell markers (NANOG, OCT4, SOX2, c-MYC, SALL4), with concurrent upregulation of the mesenchymal marker vimentin. Enhanced expression of autophagy-related genes (Beclin-1, ATG5, ATG16) was also observed, suggesting a disruption in autophagic pathways. Conclusion: Our findings underscore the critical role of Cav-1 in breast cancer progression, specifically its function in suppressing metastasis and modulating cellular phenotypes. Targeting Cav-1 could represent a novel therapeutic strategy to mitigate breast cancer growth and dissemination. Further research is warranted to explore the therapeutic potential of modulating Cav-1 in breast cancer treatment. Citation Format: Rizwana Begum, Shilpa Thota, Dhirendra Pratap Singh, Naveen Chintala, Abhishek Pandit, Biplov Sapkota, Joseph Francis. Unveiling caveolin-1's crucial role in breast cancer progression and metastasis: A molecular perspective [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 4775.

Interleukin-4 downregulates transcription factor BCL6 to promote memory B cell selection in germinal centers

Germinal center (GC)-derived memory B cells (MBCs) are critical for humoral immunity as they differentiate into protective antibody-secreting cells during re-infection. GC formation and cellular interactions within the GC have been studied in detail, yet the exact signals that allow for the selection and exit of MBCs are not understood. Here, we showed that IL-4 cytokine signaling in GC B cells directly downregulated the transcription factor BCL6 via negative autoregulation to release cells from the GC program and to promote MBC formation. This selection event required additional survival cues and could therefore result in either GC exit or death. We demonstrate that both increasing IL-4 bioavailability or limiting IL-4 signaling disrupted MBC selection stringency. In this way, IL-4 control of BCL6 expression serves as a tunable switch within the GC to tightly regulate MBC selection and affinity maturation.

Abstract C081: The lysine demethylase KDM4C is an oncogenic driver in pancreatic ductal adenocarcinoma

Abstract Deregulation of proteins involved in chromatin regulation is common in pancreatic ductal adenocarcinoma (PDAC), occurring via both genomic and non-genomic mechanisms. Lysine demethylase 4C (KDM4C) is one of the chromatin-modifying proteins frequently overexpressed across multiple solid cancers. We observed upregulation of Kdm4c protein in a panel of human and murine PDAC cell lines, as well as in PDAC patient samples compared to non-neoplastic controls. CRISPR/Cas9-mediated deletion of KDM4C in human and murine PDAC cells resulted in reduced proliferation and clonogenicity, and increased survival of orthotopically implanted murine PDAC allografts. Notably, in multiple KDM4C-null PDAC clones analyzed by reverse phase protein arrays (RPPA), we observed downregulation of activated mitogen-activated protein kinase (MAP kinase) pathway components (including phospho-MEK and phospho-ERK), which are key effector pathways downstream of mutant KRAS. In vitro propagation of KDM4C-null PDAC lines eventually led to adaptation and restitution of MAP kinase signaling via compensatory upregulation of other KDM4 family members (KDM4A and KDM4B). In order to bypass genetic compensation by KDM4 family members, we utilized a preclinical grade pan-KDM4 inhibitor (TACH107, Tachyon Therapeutics) in PDAC cell lines, which confirmed profound reduction in proliferation and colony formation; in vivo xenograft and allograft studies with TACH107 are now ongoing. To characterize signaling pathways perturbed by KDM4C deregulation, we investigated the transcriptional changes in PDAC cells with and without KDM4C deletion by RNA-seq and integrated these with alterations in H3K9me3 and H3K36me3 marks using histone ChIP-Seq. Regulatory network analysis identified ATF4 as one of the top downregulated transcription factor networks, validating a previously published observation linking KDM4C to amino acid biosynthesis and cell proliferation. Gene set enrichment analysis showed downregulation of RAS signatures, consistent with the RPPA data on attenuation of MAP Kinase pathway. DUSP2, a member of the dual specificity protein phosphatase subfamily that inactivates ERK, is transcriptionally upregulated and has increased H3K36me3 marks near the promoter region in KDM4C-depleted cells, nominating it as a candidate of KDM4C-mediated epigenetic regulation. To further delineate the essential binding partners of KDM4C in chromatin regulation, we have created HA-tagged catalytic domain and reader domain deletion constructs that will be used for immunoprecipitation followed by mass spectrometry analysis. Overall, our data suggests that KDM4C - and other KDM4 family members – regulate MAPK signaling in KRAS-mutant cells, at least partially via attenuation of transcripts encoding specific protein phosphatases. To the best of our knowledge, this is the first demonstration linking the requirement of sustained activity of the KDM4 family of lysine demethylases to MAPK signaling in cancer and presents an opportunity to leverage this oncogenic pathway for therapeutic purposes. Citation Format: Menna-t-Allah Shaheen, Laila Barkoudeh, Sarah Dhebat, Kimal I. Rajapakshe, Bidyut Ghosh, Anirban Maitra. The lysine demethylase KDM4C is an oncogenic driver in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C081.

Integrated Transcriptomic and Metabolomic Analyses Identify Critical Genes and Metabolites Associated with Seed Vigor of Common Wheat.

Seed aging is a common physiological phenomenon during storage which has a great impact on seed quality. An in-depth analysis of the physiological and molecular mechanisms of wheat seed aging is of great significance for cultivating high-vigor wheat varieties. This study reveals the physiological mechanisms of wheat seed aging in two cultivars differing in seed vigor, combining metabolome and transcriptome analyses. Differences between cultivars were examined based on metabolomic differential analysis. Artificial aging had a significant impact on the metabolism of wheat seeds. A total of 7470 (3641 upregulated and 3829 downregulated) DEGs were detected between non-aging HT and LT seeds; however, 10,648 (4506 up and 6142 down) were detected between the two cultivars after aging treatment. Eleven, eight, and four key metabolic-related gene families were identified in the glycolysis/gluconeogenesis and TCA cycle pathways, starch and sucrose metabolism pathways, and galactose metabolism pathways, respectively. In addition, 111 up-regulated transcription factor genes and 85 down-regulated transcription factor genes were identified in the LT 48h group. A total of 548 metabolites were detected across all samples. Cultivar comparisons between the non-aged groups and aged groups revealed 46 (30 upregulated and 16 downregulated) and 62 (38 upregulated and 24 downregulated) DIMs, respectively. Network analysis of the metabolites indicated that glucarate O-phosphoric acid, L-methionine sulfoxide, isocitric acid, and Gln-Gly might be the most crucial DIMs between HT and LT. The main related metabolites were enriched in pathways such as glyoxylate and dicarboxylate metabolism, biosynthesis of secondary metabolites, fatty acid degradation, etc. However, metabolites that exhibited differences between cultivars were mainly enriched in carbon metabolism, the TCA cycle, etc. Through combined metabolome and transcriptome analyses, it was found that artificial aging significantly affected glycolysis/gluconeogenesis, pyruvate metabolism, and glyoxylate and dicarboxylate metabolism, which involved key genes such as ACS, F16P2, and PPDK1. We thus speculate that these genes may be crucial in regulating physiological changes in seeds during artificial aging. In addition, an analysis of cultivar differences identified pathways related to amino acid and polypeptide metabolism, such as cysteine and methionine metabolism, glutathione metabolism, and amino sugar and nucleotide sugar metabolism, involving key genes such as BCAT3, CHI1, GAUT1, and GAUT4, which may play pivotal roles in vigor differences between cultivars.

Three-Dimensional Chromosomal Landscape Revealing miR-146a Dysfunctional Enhancer in Lupus and Establishing a CRISPR-Mediated Approach to Inhibit the Interferon Pathway.

The diminished expression of microRNA-146a (miR-146a) in systemic lupus erythematosus (SLE) contributes to the aberrant activation of the interferon pathway. Despite its significance, the underlying mechanism driving this reduced expression remains elusive. Considering the integral role of enhancers in steering gene expression, our study seeks to pinpoint the SLE-affected enhancers responsible for modulating miR-146a expression. Additionally, we aim to elucidate the mechanisms by which these enhancers influence the contribution of miR-146a to the activation of the interferon pathway. Circular chromosome conformation capture sequencing and epigenomic profiles were applied to identify candidate enhancers of miR-146a. CRISPR activation was performed to screen functional enhancers. Differential analysis of chromatin accessibility was used to identify SLE-dysregulated enhancers, and the mechanism underlying enhancer dysfunction was investigated by analyzing transcription factor binding. The therapeutic value of a lupus-related enhancer was further evaluated by targeting it in the peripheral blood mononuclear cells (PBMCs) of patients with SLE through a CRISPR activation approach. We identified shared and cell-specific enhancers of miR-146a in distinct immune cells. An enhancer 32.5 kb downstream of miR-146a possesses less accessibility in SLE, and its chromatin openness was negatively correlated with SLE disease activity. Moreover, CCAAT/enhancer binding protein α, a down-regulated transcription factor in patients with SLE, binds to the 32.5-kb enhancer and induces the epigenomic change of this locus. Furthermore, CRISPR-based activation of this enhancer in SLE PBMCs could inhibit the activity of interferon pathway. Our work defines a promising target for SLE intervention. We adopted integrative approaches to define cell-specific and functional enhancers of the SLE critical gene and investigated the mechanism underlying its dysregulation mediated by a lupus-related enhancer.

Abstract 16746: Novel Role of Endothelial CD45 in Regulating Endothelial-to-Mesenchymal Transition (EndMT) in Atherosclerosis

Introduction: Atherosclerosis is the leading cause of mortality in the United States and most other developed countries. Endothelial cell (EC) activation is a hallmark of the initiation and progression of atherosclerosis; identification of molecular markers of dysfunctional endothelium represents a biological challenge to developing new therapeutic targets. CD45 is a marker of hematopoietic cells, which is rarely expressed in ECs. However, recent studies indicated that CD45 is indispensable in driving the endothelial-to-mesenchymal transition (EndMT) following myocardial infarction. EndMT has also been implicated in the pathogenesis of atheroma progression. Thus, we aim to investigate whether endothelial CD45 contributes to the progression of atherosclerosis by facilitating the EndMT. Methods and results: In vitro , we used CRISPR/inactive Cas9 transcriptional activation system to express CD45 in human endothelial colony-forming cells (ECFCs). Using FACS analyses, we found that CD45+ECFCs upregulate transforming growth factors (TGFβ 1-3 ) that implicate EndMT but downregulate FGFR1 expression, which exacerbates EndMT. qRT-PCR and WB showed that CD45+ECFCs downregulate transcription factors KLFs. In vivo , we generated a tamoxifen-inducible EC-specific CD45 deficient mouse strain (EC-CD45iKO) in ApoE-deficient (ApoE-/-) background and fed with a Western diet (C57BL/6) for atherosclerosis and molecular analyses. We found that ApoE-/- / EC-iCD45KO mice reduced plaque macrophages, foam cells, expression of adhesion molecule, and lesion development compared to ApoE-/- controls. Interestingly, ApoE-/- / EC-iCD45KO mice showed the reduction of α-smooth muscle actin, which is typical of EndMT, in the aortic plaque. Single-cell RNA-seq analyses have been performed to characterize that endothelial CD45 regulates the expression of pro-inflammatory, EndMT, and endothelial cell markers genes on the aortic ECs. Conclusion: These findings demonstrate that endothelial CD45 potentiates atherosclerosis by inducing TGF signaling and mesenchymal phenotype, causing FGFR1 downregulation and attenuating KLFs expression to drive EndMT, highlighting endothelial CD45 may be a new drug target for the treatment of atherosclerosis.

Protective roles of adiponectin and molecular signatures of HNF4α and PPARα as downstream targets of adiponectin in pancreatic β cells

The disease progression of the metabolic syndrome is associated with prolonged hyperlipidemia and insulin resistance, eventually giving rise to impaired insulin secretion, often concomitant with hypoadiponectinemia. As an adipose tissue derived hormone, adiponectin is beneficial for insulin secretion and β cell health and differentiation. However, the down-stream pathway of adiponectin in the pancreatic islets has not been studied extensively. Here, along with the overall reduction of endocrine pancreatic function in islets from adiponectin KO mice, we examine PPARα and HNF4α as additional down-regulated transcription factors during a prolonged metabolic challenge. To elucidate the function of β cell-specific PPARα and HNF4α expression, we developed doxycycline inducible pancreatic β cell-specific PPARα (β-PPARα) and HNF4α (β-HNF4α) overexpression mice. β-PPARα mice exhibited improved protection from lipotoxicity, but elevated β-oxidative damage in the islets, and also displayed lowered phospholipid levels and impaired glucose-stimulated insulin secretion. β-HNF4α mice showed a more severe phenotype when compared to β-PPARα mice, characterized by lower body weight, small islet mass and impaired insulin secretion. RNA-sequencing of the islets of these models highlights overlapping yet unique roles of β-PPARα and β-HNF4α. Given that β-HNF4α potently induces PPARα expression, we define a novel adiponectin-HNF4α-PPARα cascade. We further analyzed downstream genes consistently regulated by this axis. Among them, the islet amyloid polypeptide (IAPP) gene is an important target and accumulates in adiponectin KO mice. We propose a new mechanism of IAPP aggregation in type 2 diabetes through reduced adiponectin action.

Icaritin ameliorates RANKL-mediated osteoclastogenesis and ovariectomy-induced osteoporosis.

A rapidly aging society and longer life expectancy are causing osteoporosis to become a global epidemic. Over the last five decades, a number of drugs aimed at reducing bone resorption or restoring bone mass have been developed, but their efficacy and safety are limited. Icaritin (ICT) is a natural compound extracted from anti-osteoporosis herb Epimedium spp. and has been shown to inhibit osteoclast differentiation. However, the molecular mechanism by which ICT weaken RANKL-induced osteoclast differentiation has not been completely investigated. Here, we evaluated the anti-osteoclastogenic effect of ICT in vitro and the potential drug candidate for treating osteoporosis in vivo. In vitro study, ICT was found to inhibit osteoclast formation and bone resorption function via downregulating transcription factors activated T cell cytoplasm 1 (NFATc1) and c-fos, which further downregulate osteoclastogenesis-specific gene. In addition, the enhanced mitochondrial mass and function required for osteoclast differentiation was mitigated by ICT. The histomorphological results from an in vivo study showed that ICT attenuated the bone loss associated with ovariectomy (OVX). Based on these results, we propose ICT as a promising new drug strategy for osteoporosis that inhibits osteoclast differentiation.

The chaperone protein GRP78 released from MPN cells increases the expression of lysyl oxidase in a human stromal cell line

Impaired function of the endoplasmic stress (ER) response causes numerous pathological conditions, including tissue fibrosis. In the present study, we aimed to determine the pathological role of ER stress response systems in myeloproliferative neoplasms (MPNs). We found increased expression of the chaperone protein glucose-regulated protein (GRP) 78, a central regulator of ER stress, in megakaryocytes from primary myelofibrosis or postessential thrombocythemia myelofibrosis patients. GRP78 was overexpressed in JAK2V617F-harboring cell lines; however, inhibitors of ER stress did not affect the expression levels of GRP78. In contrast, ruxolitinib, a well-known inhibitor of JAK2V617F, clearly blocked GRP78 expression in these cells through downregulation of transcription factor 4 (ATF4). Interestingly, GRP78 was secreted from HEL and SET-2 cells into culture media. Coculture of these cells with HS-5 cells, a human bone marrow stroma-derived cell line, induced enhanced expression of lysyl oxidase (LOX), which mediates cross-linking of collagen fibers and induces tissue fibrosis, in HS-5 cells. An anti-GRP78 neutralizing antibody abrogated LOX elevation; in contrast, recombinant GRP78 protein induced LOX protein expression in HS-5 cells. Our observations suggest that the oncogenic protein JAK2V617F induces overexpression and release of GRP78, which may induce a fibrotic phenotype in surrounding bone marrow stromal cells.

Effect of D-pinitol on MK-801-induced schizophrenia-like behaviors in mice.

Schizophrenia is a chronic brain disorder characterized by positive symptoms (delusions or hallucinations), negative symptoms (impaired motivation or social withdrawal), and cognitive impairment. In the present study, we explored whether D-pinitol could ameliorate schizophrenia-like behaviors induced by MK-801, an N-methyl-D-aspartate receptor antagonist. Acoustic startle response test was conducted to evaluate the effects of D-pinitol on sensorimotor gating function. Social interaction and novel object recognition tests were employed to measure the impact of D-pinitol on social behavior and cognitive function, respectively. Additionally, we examined whether D-pinitol affects motor coordination. Western blotting was conducted to investigate the mechanism of action of D-pinitol. Single administration of D-pinitol at 30, 100, or 300 mg/kg improved the sensorimotor gating deficit induced by MK801 in the acoustic startle response test. D-Pinitol also reversed social behavior deficits and cognitive impairments induced by MK-801 without causing any motor coordination deficits. Furthermore, D-pinitol reversed increased expression levels of pNF-kB induced by MK-801 treatment and consequently increased expression levels of TNF-α and IL-6 in the prefrontal cortex. These results suggest that D-pinitol could be a potential candidate for treating sensorimotor gating deficits and cognitive impairment observed in schizophrenia by down-regulating transcription factor NF-κB and pro-inflammatory cytokines in the prefrontal cortex.

Epigenetically upregulating TROP2 and SLFN11 enhances therapeutic efficacy of TROP2 antibody drug conjugate sacitizumab govitecan

TROP2 antibody drug conjugates (ADCs) are under active development. We seek to determine whether we can enhance activity of TROP2 ADCs by increasing TROP2 expression. In metaplastic breast cancers (MpBC), there is limited expression of TROP2, and downregulating transcription factor ZEB1 upregulates E-cad and TROP2, thus sensitizing cancers to TROP2 ADC sacituzumab govitecan (SG). Demethylating agent decitabine decreases DNA methyltransferase expression and TROP2 promoter methylation and subsequently increases TROP2 expression. Decitabine treatment as well as overexpression of TROP2 significantly enhance SG antitumor activity. Decitabine also increases SLFN11, a biomarker of topoisomerase 1 inhibitor (TOP1) sensitivity and is synergistic with SG which has a TOP1 payload, in TROP2-expressing SLFN11-low BC cells. In conclusion, TROP2 and SLFN11 expression can be epigenetically modulated and the combination of demethylating agent decitabine with TROP2 ADCs may represent a novel therapeutic approach for tumors with low TROP2 or SLFN11 expression.

Transcription factor expression repertoire basis for epigenetic and transcriptional subtypes of colorectal cancers

Colorectal cancers (CRCs) form a heterogenous group classified into epigenetic and transcriptional subtypes. The basis for the epigenetic subtypes, exemplified by varying degrees of promoter DNA hypermethylation, and its relation to the transcriptional subtypes is not well understood. We link cancer-specific transcription factor (TF) expression alterations to methylation alterations near TF-binding sites at promoter and enhancer regions in CRCs and their premalignant precursor lesions to provide mechanistic insights into the origins and evolution of the CRC molecular subtypes. A gradient of TF expression changes forms a basis for the subtypes of abnormal DNA methylation, termed CpG-island promoter DNA methylation phenotypes (CIMPs), in CRCs and other cancers. CIMP is tightly correlated with cancer-specific hypermethylation at enhancers, which we term CpG-enhancer methylation phenotype (CEMP). Coordinated promoter and enhancer methylation appears to be driven by downregulation of TFs with common binding sites at the hypermethylated enhancers and promoters. The altered expression of TFs related to hypermethylator subtypes occurs early during CRC development, detectable in premalignant adenomas. TF-based profiling further identifies patients with worse overall survival. Importantly, altered expression of these TFs discriminates the transcriptome-based consensus molecular subtypes (CMS), thus providing a common basis for CIMP and CMS subtypes.

Natural compound So-2 suppresses triple-negative breast cancer through inducing ferroptosis via downregulating transcription factor E2F7

Triple-negative breast cancer (TNBC), accounting for about 15∼18% of all breast cancers, is notorious for its poor prognosis, high rate of relapse and short overall survival. Because of lacking effective therapeutic targets or drugs, treatment of TNBC in clinical encounters great obstacle. Siegesbeckiaorientalis L. have been used as a traditional Chinese medicine “Xi-Xian-Cao” for centuries with multiple medicinal benefits including cancerous treatment. We have reported the isolation of twenty-seven germacranolides including So-2 from the aerial parts of S. orientalis with potent cytotoxicity against breast cancer cells. The studyaims to verified the anti-TNBC function of the natural compound So-2 both in vitro and vivo and uncover the underlying mechanism. The results showed that So-2 caused cell cycle arrest and suppress TNBC cell proliferation and migration. Also, So-2 was first identified to be a bona fide ferroptosis inducer in TNBC cells. So-2 effectively suppressed tumor growth of TNBC by using an orthotopic transplantation tumor model. We also characterized the oncogenic role of the transcription factor E2F7 in TNBC. E2F7 was demonstrated to be involved in the ferroptosis-inducing and tumor suppression effect of So-2. Altogether, So-2 exhibits inhibitory effect on TNBC both in vitro and vivo by inducing TNBC ferroptosis via downregulating the expression of E2F7. These findings provide valuable insight into the pathogenesis of TNBC. The natural compound So-2, isolated from Chinese traditional medicine, might be a prospective drug candidate in TNBC therapy.

Comparative cytological and transcriptome analyses of ny2 mutant delayed degeneration of tapetal cells and promotes abnormal microspore development in neo-tetraploid rice.

We aimed to investigate the genetic defects related to pollen development and infertility in NY2, a novel tetraploid rice germplasm known as Neo-tetraploid rice. This rice variety was created through the crossbreeding and selective breeding of various autotetraploid rice lines and has previously shown high fertility. Our previous research has revealed that the NY2 gene, encoding a eukaryotic translation initiation factor 3 subunit E, regulates pollen fertility. However, the underlying mechanism behind this fertility is yet to be understood. To shed light on this matter, we performed a combined cytological and transcriptome analysis of the NY2 gene. Cytological analysis indicated that ny2 underwent abnormal tapetal cells, microspore, and middle layer development, which led to pollen abortion and ultimately to male sterility. Genetic analysis revealed that the F1 plants showed normal fertility and an obvious advantage for seed setting compared to ny2. Global gene expression analysis in ny2 revealed a total of 7545 genes were detected at the meiosis stage, and 3925 and 3620 displayed upregulation and downregulation, respectively. The genes were significantly enriched for the gene ontology (GO) term "carbohydrate metabolic process. Moreover, 9 genes related to tapetum or pollen fertility showed down-regulation, such as OsABCG26 (ATP Binding Cassette G26), TMS9-1 (Thermosensitive Male Sterility), EAT1 (Programmed cell death regulatory), KIN14M (Kinesin Motor), OsMT1a (Metallothionein), and OsSTRL2 (Atypical strictosidine synthase), which were validated by qRT-PCR. Further analyses of DEGs identified nine down-regulated transcription factor genes related to pollen development. NY2 is an important regulator of the development of tapetum and microspore. The regulatory gene network described in this study may offer important understandings into the molecular processes that underlie fertility control in tetraploid rice.

Peroxisome proliferator-activated receptor γ activation ameliorates liver fibrosis-differential action of transcription factor EB and autophagy on hepatocytes and stellate cells.

Peroxisome proliferator-activated receptor γ (PPARγ) activation suppresses HSC activation and liver fibrosis. Moreover, autophagy is implicated in hepatic lipid metabolism. Here, we determined whether PPARγ activation ameliorates HSC activation by downregulating transcription factor EB (TFEB)-mediated autophagy. Atg7 or Tfeb knockdown in human HSC line LX-2 cells downregulated the expression of fibrogenic markers including α smooth muscle actin, glial fibrillary acidic protein, and collagen type 1. Conversely, Atg7 or Tfeb overexpression upregulated fibrogenic marker expression. Rosiglitazone (RGZ)-mediated PPARγ activation and/or overexpression in LX-2 cells and primary HSCs decreased autophagy, as indicated by LC3B conversion, total and nuclear-TFEB contents, mRFP-LC3 and BODIPY 493/503 colocalization, and GFP-LC3 and LysoTracker colocalization. RGZ treatment decreased liver fat content, liver enzyme levels, and fibrogenic marker expression in high-fat high-cholesterol diet-fed mice. Electron microscopy showed that RGZ treatment restored the high-fat high-cholesterol diet-mediated lipid droplet decrease and autophagic vesicle induction in primary HSCs and liver tissues. However, TFEB overexpression in LX-2 cells offset the aforementioned effects of RGZ on autophagic flux, lipid droplets, and fibrogenic marker expression. Activation of PPARγ with RGZ ameliorated liver fibrosis and downregulation of TFEB and autophagy in HSCs may be important for the antifibrotic effects of PPARγ activation.

Downregulated mitochondrial transcription factor A enhances mycoplasma infection to promote the metastasis of hepatocellular carcinoma.

Mycoplasma is widespread in various hosts and may cause various diseases in animals. Interestingly, the occurrence of mycoplasma infection was observed in many tumor types. However, the mechanism regulating its infection is far from clear. We unexpectedly found that the knockdown of mitochondrial transcription factor A (TFAM) remarkably enhanced mycoplasma infection in hepatocellular carcinoma (HCC) cells. More importantly, we found that mycoplasma infection facilitated by TFAM knockdown significantly promoted HCC cell metastasis. Mycoplasma infection was further found to be positively correlated with poor prognosis in patients with HCC. Mechanistically, the decreased TFAM expression upregulated the transcription factor Sp1 to increase the expression level of Annexin A2 (ANXA2), which was reported to interact with membrane protein of mycoplasma. Moreover, we found that mycoplasma infection enhanced by the TFAM downregulation promoted HCC migration and invasion by activating the nuclear factor-κB signaling pathway. The downregulation of TFAM enhanced mycoplasma infection in HCC cells and promoted HCC cell metastasis. Our study contributes to the understanding of the pathological role of mycoplasma infection and provides supporting evidence that targeting TFAM could be a potential strategy for the treatment of HCC with mycoplasma infection.

Alternative splicing of POUM2 regulates embryonic cuticular formation and tanning in Bombyx mori.

Insect cuticle is an apical extracellular matrix produced by the epidermis, tracheal, hind- and foregut epithelia during embryogenesis and renewed during molting and metamorphosis. However, the underlying regulatory mechanism for embryonic cuticle formation remains largely unclear. Here, we investigate the function of the transcription factor POUM2 in the embryonic cuticular formation in Bombyx mori, a model lepidopteran insect. Clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein-9-mediated knockout of POUM2 resulted in the defect of cuticular deposition, pigmentation, and sclerotization in the embryos. Differentially expressed transcripts analysis of 7-d-old embryos identified 174 up- or downregulated cuticular protein transcripts, 8 upregulated chitin degradation transcripts, 2 downregulated chitin synthesis transcripts and 48 up- or downregulated transcription factor transcripts in the POUM2-/- embryos. The expression levels of the key factors of the tyrosine metabolic pathway, such as tyrosine hydroxylase (Th), Dopa decarboxylase (DDC), and arylalkylamine N-acetyltransferase (aaNAT), were significantly decreased in the POUM2-/- embryos. POUM2 isoform POUM2-L specifically bound the POU cis-regulatory element (CRE) in the Th promoter and increased the transcription of Th, whereas POUM2-S could not bind the POU CRE, although it also increased the transcription of Th. Heterogeneous nuclear ribonucleoprotein Squid-1 directly bound the POUM2 pre-mRNA (messenger RNA) and inhibited the alternative splicing of POUM2-L to POUM2-S mRNA. These results suggest that POUM2 participates in the cuticular formation by regulating the chitin and cuticular protein synthesis and metabolism, and the cuticular pigmentation and sclerotization by regulating tyrosine metabolism during embryogenesis. This study provides new insights into novel function of POUM2 in embryogenesis.

Abstract A012: Transcription factor expression repertoire basis for epigenetic and transcriptional subtypes of colorectal cancers

Abstract Colorectal cancers (CRCs) form a heterogenous group classified into epigenetic and transcriptional subtypes. The basis for the epigenetic subtypes, exemplified by varying degrees of promoter DNA hypermethylation, and its relation to the transcriptional subtypes is not well understood. We now link cancer-specific transcription factor (TF) expression dysregulation to methylation alterations near TF-binding sites at promoter and enhancer regions in CRCs and their pre-malignant precursor lesions to provide mechanistic insights into the origins and evolution of the CRC molecular subtypes. A gradient of TF-expression changes forms a basis for the subtypes of abnormal DNA methylation, termed CpG-island promoter DNA methylation phenotypes (CIMP), in CRCs and other cancers. CIMP is tightly correlated with cancer-specific hypermethylation at enhancers, which we term CpG-enhancer methylation phenotype (CEMP). CIMP/CEMP coordination appears to be driven by augmented downregulation of TFs with common binding sites at the hypermethylated enhancers and promoters. The dysregulated expression of TFs related to CIMP/CEMP subtypes occurs early during CRC development, detectable in pre-malignant adenomas. TF-based profiling further identifies patients with worse overall survival. Importantly, altered expression of these TFs discriminates the transcriptome-based consensus molecular subtypes (CMS), thus providing a common basis for CIMP and CMS subtypes. Citation Format: Yuba R. Bhandari, Vinod Krishna, Rachael Powers, Sehej Parmar, Sara-Jayne Thursby, Ekta Gupta, Ozlem Kulak, Prashanth Gokare, Joke Reumers, Liesbeth Van Wesenbeeck, Kurtis E. Bachman, Stephen B. Baylin, Hariharan Easwaran. Transcription factor expression repertoire basis for epigenetic and transcriptional subtypes of colorectal cancers. [abstract]. In: Proceedings of the AACR Special Conference: Cancer Epigenomics; 2022 Oct 6-8; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2022;82(23 Suppl_2):Abstract nr A012.

Comparative transcriptomics of tropical woody plants supports fast and furious strategy along the leaf economics spectrum in lianas.

ABSTRACTLianas, climbing woody plants, influence the structure and function of tropical forests. Climbing traits have evolved multiple times, including ancestral groups such as gymnosperms and pteridophytes, but the genetic basis of the liana strategy is largely unknown. Here, we use a comparative transcriptomic approach for 47 tropical plant species, including ten lianas of diverse taxonomic origins, to identify genes that are consistently expressed or downregulated only in lianas. Our comparative analysis of full-length transcripts enabled the identification of a core interactomic network common to lianas. Sets of transcripts identified from our analysis reveal features related to functional traits pertinent to leaf economics spectrum in lianas, include upregulation of genes controlling epidermal cuticular properties, cell wall remodeling, carbon concentrating mechanism, cell cycle progression, DNA repair and a large suit of downregulated transcription factors and enzymes involved in ABA-mediated stress response as well as lignin and suberin synthesis. All together, these genes are known to be significant in shaping plant morphologies through responses such as gravitropism, phyllotaxy and shade avoidance.