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Primary cilium is a microtubule-based organelle that projects from the surfaces of most mammalian cell types and protrudes into the extracellular milieu as an antenna-like sensor to senses extracellular physical and biochemical signals, and then transmits signals into cytoplasm or nucleus to regulate numerous physical and developmental processes. Therefore, loss of primary cilia is associated to multiple cancer progression, including skin, breast, pancreas, ovarian, prostate, and kidney cancers. Our previous studies demonstrate that high prevalent loss of DAB2 Interacting Protein (DAB2IP) is associated with renal cell carcinoma, and we found a kinesin-like protein, kinesin family member 3A (KIF3a), was significantly increased in DAB2IP-interacting protein fraction. KIF3 is one of the most abundant kinesin-2 family proteins expressed in cells, and it is necessary for ciliogenesis. In this study, we observed that loss of DAB2IP in normal kidney epithelial cell significantly impair primary cilia formation. We unveiled a new mechanism of primary cilia stability via DAB2IP and KIF3a physical interaction at DAB2IP-PH domain. Furthermore, we found that KIF3a also act as a tumor suppressor in renal cell carcinoma, affect tumor development and patient survival.

Funding Information: Funding and additional information—C. G. A. has obtained funding from Maratona da Saúde 2016; CEECIND/00410/2017 financed by FCT (Portugal); ALZ AARG-19–618007 (Alzheimer’s Association); the European Union’s Horizon 2020 research and innovation program under grant agreement No 811087 (Lysocil). C. B. P. was the recipient of an FCT doctoral fellowship (PD/BD/128374/2017). T. B. is the recipient of an FCT doctoral fellowship (SFRH/BD/131513/ 2017). M. A. B. is the recipient of an FCT doctoral fellowship (2020.06758.BD). Funding Information: C. G. A. has obtained funding from Maratona da Sa?de 2016; CEECIND/00410/2017 financed by FCT (Portugal); ALZ AARG-19-618007 (Alzheimer's Association); the European Union's Horizon 2020 research and innovation program under grant agreement No 811087 (Lysocil). C. B. P. was the recipient of an FCT doctoral fellowship (PD/BD/128374/2017). T. B. is the recipient of an FCT doctoral fellowship (SFRH/BD/131513/ 2017). M. A. B. is the recipient of an FCT doctoral fellowship (2020.06758.BD). Funding Information: Acknowledgments—We thank M. Arpin (I Curie) and Z. Lenkei (ESPCI-ParisTech) for the gift of plasmids and cells, respectively. We thank Ana Cláudia Marques for her preliminary observations and lab members for helpful discussions and critical reading of the manuscript. We thank S. Marques (CEDOC Animal Facility), T. Pereira (CEDOC Microscopy Facility), and Ana Oliveira (CEDOC Cell Culture Facility) for their technical assistance. This project has received institutional funding from iNOVA4Health— UID/Multi/ 04462/2019; H2020-WIDESPREAD-01-2016-2017-TeamingPhase2 - GA739572; the research infrastructure PPBI-POCI-01-0145-FEDER-022122, co-financed by FCT (Portugal) and Lisboa2020, under the PORTUGAL2020 agreement (European Regional Development Fund). Publisher Copyright: © 2021 THE AUTHORS. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Genetic studies have identified BIN1 as the second most important risk locus associated with late-onset Alzheimer's disease (LOAD). However, it is unclear how mutation of this locus mechanistically promotes Alzheimer's disease (AD) pathology. Here we show the consequences of two coding variants in BIN1 (rs754834233 and rs138047593), both in terms of intracellular beta-amyloid (iAbeta) accumulation and early endosome enlargement, two interrelated early cytopathological AD phenotypes, supporting their association with LOAD risk. We previously found that Bin1 deficiency potentiates iAbeta production by enabling BACE1 cleavage of the amyloid precursor protein in enlarged early endosomes due to decreased BACE1 recycling. Here, we discovered that the expression of the two LOAD mutant forms of Bin1 does not rescue the iAbeta accumulation and early endosome enlargement induced by Bin1 knockdown and recovered by wild-type Bin1. Moreover, the overexpression of Bin1 mutants, but not wild-type Bin1, increased the iAbeta42 fragment by reducing the recycling of BACE1, which accumulated in early endosomes, recapitulating the phenotype of Bin1 knockdown. We showed that the mutations in Bin1 reduced its interaction with BACE1. The endocytic recycling of transferrin was similarly affected, indicating that Bin1 is a general regulator of endocytic recycling. These data demonstrate that the LOAD-coding variants in Bin1 lead to a loss of function in endocytic recycling, which may be an early causal mechanism of LOAD. publishersversion published

Human cell division is a highly regulated process that relies on the accurate capture and movement of chromosomes to the metaphase plate. Errors in the fidelity of chromosome congression and alignment can lead to improper chromosome segregation, which is correlated with aneuploidy and tumorigenesis. These processes are known to be regulated by extracellular signal-regulated kinase 2 (ERK2) in other species, but the role of ERK2 in mitosis in mammals remains unclear. Here, we have identified the dual-specificity phosphatase 7 (DUSP7), known to display selectivity for ERK2, as important in regulating chromosome alignment. During mitosis, DUSP7 bound to ERK2 and regulated the abundance of active phospho-ERK2 through its phosphatase activity. Overexpression of DUSP7, but not catalytically inactive mutants, led to a decrease in the levels of phospho-ERK2 and mitotic chromosome misalignment, while knockdown of DUSP7 also led to defective chromosome congression that resulted in a prolonged mitosis. Consistently, knockdown or chemical inhibition of ERK2 or chemical inhibition of the MEK kinase that phosphorylates ERK2 led to chromosome alignment defects. Our results support a model wherein MEK-mediated phosphorylation and DUSP7-mediated dephosphorylation regulate the levels of active phospho-ERK2 to promote proper cell division.

Burkitt's lymphoma (BL) is a highly aggressive form of B-cell non-Hodgkin's lymphoma. The clinical outcome in children with BL has improved over the last years but the prognosis for adults is still poor, highlighting the need for novel treatment strategies. Here, we report that the combinational treatment with the Smac mimetic BV6 and TRAIL triggers necroptosis in BL when caspases are blocked by zVAD.fmk (TBZ treatment). The sensitivity of BL cells to TBZ correlates with MLKL expression. We demonstrate that necroptotic signaling critically depends on MLKL, since siRNA-induced knockdown and CRISPR/Cas9-mediated knockout of MLKL profoundly protect BL cells from TBZ-induced necroptosis. Conversely, MLKL overexpression in cell lines expressing low levels of MLKL leads to necroptosis induction, which can be rescued by pharmacological inhibitors, highlighting the important role of MLKL for necroptosis execution. Importantly, the methylation status analysis of the MLKL promoter reveals a correlation between methylation and MLKL expression. Thus, MLKL is epigenetically regulated in BL and might serve as a prognostic marker for treatment success of necroptosis-based therapies. These findings have crucial implications for the development of new treatment options for BL., Graphical Abstract Image, graphical abstract

Traditionally, lipolysis has been regarded as an enzymatic activity that liberates fatty acids as metabolic fuel. However, recent work has shown that novel substrates, including a variety of lipid compounds such as fatty acids and their derivatives, release lipolysis products that act as signaling molecules and transcriptional modulators. While these studies have expanded the role of lipolysis, the mechanisms underpinning lipolysis signaling are not fully defined. Here, we uncover a new mechanism regulating glucose uptake, whereby activation of lipolysis, in response to elevated cAMP, leads to the stimulation of thioredoxin-interacting protein (TXNIP) degradation. This, in turn, selectively induces glucose transporter 1 surface localization and glucose uptake in 3T3-L1 adipocytes and increases lactate production. Interestingly, cAMP-induced glucose uptake via degradation of TXNIP is largely dependent upon adipose triglyceride lipase (ATGL) and not hormone-sensitive lipase or monoacylglycerol lipase. Pharmacological inhibition or knockdown of ATGL alone prevents cAMP-dependent TXNIP degradation and thus significantly decreases glucose uptake and lactate secretion. Conversely, overexpression of ATGL amplifies the cAMP response, yielding increased glucose uptake and lactate production. Similarly, knockdown of TXNIP elicits enhanced basal glucose uptake and lactate secretion, and increased cAMP further amplifies this phenotype. Overexpression of TXNIP reduces basal and cAMP-stimulated glucose uptake and lactate secretion. As a proof of concept, we replicated these findings in human primary adipocytes and observed TXNIP degradation and increased glucose uptake and lactate secretion upon elevated cAMP signaling. Taken together, our results suggest a crosstalk between ATGL-mediated lipolysis and glucose uptake.

The clinical efficacy of docetaxel (DTX) in prostate cancer treatment is barely satisfactory due to diverse responses of the patients, including the development of resistance. Recently, aberrant androgen receptor (AR) signaling, including expression of the constitutively active ARV7, was reported to contribute to DTX resistance. However, the underlying molecular mechanism remains largely unknown. Of note, previous studies have highlighted that ARV7, unlike its parental AR, potentially favors the expression of some genes involved in cell cycle progression. Since DTX mainly targets microtubule dynamics and mitosis, we wanted to test whether ARV7 plays a specific role in mitotic regulation and whether this activity is involved in DTX resistance. In the present study, we found that ARV7 mediates DTX sensitivity through inactivating the spindle assembly checkpoint (SAC) and promoting mitotic slippage. By shifting the balance to the slippage pathway, ARV7-expressing cells are more likely to escape from mitotic death induced by acute DTX treatment. Furthermore, we also identified E2 enzyme UBE2C as the primary downstream effector of ARV7 in promoting the SAC inactivation and premature degradation of cyclin B1. Moreover, we showed that combination treatment of DTX and an inhibitor of mitotic exit can exert synergistic effect in high ARV7-expressing prostate cancer cells. In sum, our work identified a novel role of ARV7 in promoting DTX resistance and offering a potential path to combat DTX resistance related to abnormal activation of the AR signaling and mitotic dysregulation.

The Yes-associated protein (YAP), one of the major effectors of the Hippo pathway together with its related protein WW-domain-containing transcription regulator 1 (WWTR1; also known as TAZ), mediates a range of cellular processes from proliferation and death to morphogenesis. YAP and WW-domain-containing transcription regulator 1 (WWTR1; also known as TAZ) regulate a large number of target genes, acting as coactivators of DNA-binding transcription factors or as negative regulators of transcription by interacting with the nucleosome remodeling and histone deacetylase complexes. YAP is expressed in self-renewing embryonic stem cells (ESCs), although it is still debated whether it plays any crucial roles in the control of either stemness or differentiation. Here we show that the transient downregulation of YAP in mouse ESCs perturbs cellular homeostasis, leading to the inability to differentiate properly. Bisulfite genomic sequencing revealed that this transient knockdown caused a genome-wide alteration of the DNA methylation remodeling that takes place during the early steps of differentiation, suggesting that the phenotype we observed might be due to the dysregulation of some of the mechanisms involved in regulation of ESC exit from pluripotency. By gene expression analysis, we identified two molecules that could have a role in the altered genome-wide methylation profile: the long noncoding RNA ephemeron, whose rapid upregulation is crucial for the transition of ESCs into epiblast, and the methyltransferase-like protein Dnmt3l, which, during the embryo development, cooperates with Dnmt3a and Dnmt3b to contribute to the de novo DNA methylation that governs early steps of ESC differentiation. These data suggest a new role for YAP in the governance of the epigenetic dynamics of exit from pluripotency.

ST6Gal-I, an enzyme upregulated in numerous malignancies, adds α2-6-linked sialic acids to select membrane receptors, thereby modulating receptor signaling and cell phenotype. In this study, we investigated ST6Gal-I's role in epithelial to mesenchymal transition (EMT) using the Suit2 pancreatic cancer cell line, which has low endogenous ST6Gal-I and limited metastatic potential, along with two metastatic Suit2-derived subclones, S2-013 and S2-LM7AA, which have upregulated ST6Gal-I. RNA-Seq results suggested that the metastatic subclones had greater activation of EMT-related gene networks than parental Suit2 cells, and forced overexpression of ST6Gal-I in the Suit2 line was sufficient to activate EMT pathways. Accordingly, we evaluated expression of EMT markers and cell invasiveness (a key phenotypic feature of EMT) in Suit2 cells with or without ST6Gal-I overexpression, as well as S2-013 and S2-LM7AA cells with or without ST6Gal-I knockdown. Cells with high ST6Gal-I expression displayed enrichment in mesenchymal markers (N-cadherin, slug, snail, fibronectin) and cell invasiveness, relative to ST6Gal-I-low cells. Contrarily, epithelial markers (E-cadherin, occludin) were suppressed in ST6Gal-I-high cells. To gain mechanistic insight into ST6Gal-I's role in EMT, we examined the activity of epidermal growth factor receptor (EGFR), a known EMT driver. ST6Gal-I-high cells had greater α2-6 sialylation and activation of EGFR than ST6Gal-I-low cells. The EGFR inhibitor, erlotinib, neutralized ST6Gal-I-dependent differences in EGFR activation, mesenchymal marker expression, and invasiveness in Suit2 and S2-LM7AA, but not S2-013, lines. Collectively, these results advance our understanding of ST6Gal-I's tumor-promoting function by highlighting a role for ST6Gal-I in EMT, which may be mediated, at least in part, by α2-6-sialylated EGFR.

Circular RNAs (circRNAs) are a novel class of widespread noncoding RNAs that regulate gene expression in mammals. Recent studies demonstrate that functional peptides can be encoded by short open reading frames in noncoding RNAs, including circRNAs. However, the role of circRNAs in various physiological and pathological states, such as cancer, is not well understood. In this study, through deep RNA sequencing on human endometrial cancer (EC) samples and their paired adjacent normal tissues, we uncovered that the circRNA hsa-circ-0000437 is significantly reduced in EC compared with matched paracancerous tissue. The hsa-circ-0000437 contains a short open reading frame encoding a functional peptide termed CORO1C-47aa. Overexpression of CORO1C-47aa is capable of inhibiting angiogenesis at the initiation stage by suppressing endothelial cell proliferation, migration, and differentiation through competition with transcription factor TACC3 to bind to ARNT and suppress VEGF. CORO1C-47aa directly bound to ARNT through the PAS-B domain, and blocking the association between ARNT and TACC3, which led to reduced expression of VEGF, ultimately lead to reduced angiogenesis. The antitumor effects of CORO1C-47aa on EC progression suggest that CORO1C-47aa has potential value in anticarcinoma therapies and warrants further investigation.

Metformin is the first-line antidiabetic agent for type 2 diabetes mellitus (T2DM) treatment. Although accumulated evidence has shed light on the consequences of metformin action, the precise mechanisms of its action, especially in the pancreas, are not fully understood. Aquaporin 7 (AQP7) acts as a critical regulator of intraislet glycerol content, which is necessary for insulin production and secretion. The aim of this study was to investigate the effects of different doses of metformin on AQP7 expression and explore the possible mechanism of its protective effects in the pancreatic islets. We used an in vivo model of high-fat diet in streptozocin-induced diabetic rats and an in vitro model of rat pancreatic β-cells (INS-1 cells) damaged by hyperglycemia and hyperlipidemia. Our data showed that AQP7 expression levels were decreased, whereas p38 and JNK mitogen-activated protein kinases (MAPKs) were activated in vivo and in vitro in response to hyperglycemia and hyperlipidemia. T2DM rats treated with metformin demonstrated a reduction in blood glucose levels and increased regeneration of pancreatic β-cells. In addition, metformin upregulated AQP7 expression as well as inhibited activation of p38 and JNK MAPKs both in vivo and in vitro. Overexpression of AQP7 increased glycerol influx into INS-1 cells, whereas inhibition of AQP7 reduced glycerol influx, thereby decreasing subsequent insulin secretion. Our findings demonstrate a new mechanism by which metformin suppresses the p38 and JNK pathways, thereby upregulating pancreatic AQP7 expression and promoting glycerol influx into pancreatic β-cells and subsequent insulin secretion in T2DM.

Ultraviolet B (UVB) irradiation can induce reactive oxygen species (ROS) production and apoptosis in human lens epithelial cells (HLECs), thus leading to the formation of cataracts. We studied the role of tripartite motif 69 (TRIM69) in cataract formation. The expression of TRIM69 protein was down-regulated in both human cataract capsule tissues and HLECs treated with UVB, whereas the expression of p53 protein exhibited an opposite trend. Ectopic expression of TRIM69 in HLECs significantly suppressed UVB-induced apoptosis and ROS production, whereas knockdown of TRIM69 promoted apoptosis and ROS production. TRIM69 can interact with p53 and induce its ubiquitination. The effects of TRIM69 overexpression in UVB-induced cell apoptosis and ROS production was clearly weakened by p53 overexpression, thus suggesting a role for p53 in TRIM69 functions. Furthermore, inhibition of ROS mitigated the effects of UVB irradiation on ROS production, cell apoptosis, forkhead box protein 3a (Foxo3a) phosphorylation, and TRIM69 expression. Additionally, Foxo3a overexpression significantly enhanced TRIM69 promoter activity, whereas Foxo3a knockdown had the opposite effect. In conclusion, we provide the first demonstration that Foxo3a is a potential transcription factor for TRIM69, and TRIM69 induces p53 ubiquitination. These results suggest that the Foxo3a/TRIM69/p53 regulatory network may be involved in cataract formation., Graphical abstract Image 1, Highlights • TRIM69 significantly suppressed UVB-induced apoptosis and ROS production. • TRIM69 can interact with p53 and induce its ubiquitination. • Foxo3a overexpression significantly enhanced TRIM69 promoter activity. • The Foxo3a/TRIM69/p53 regulatory network may be involved in cataract formation.

The superoxide free radical (O2•−) has been viewed as a likely major contributor to aging. If this is correct, then superoxide dismutase (SOD), which removes O2•−, should contribute to longevity assurance. In Caenorhabditis elegans, overexpression (OE) of the major cytosolic Cu/Zn-SOD, sod-1, increases life span. But is this increase caused by enhanced antioxidant defense? sod-1 OE did not reduce measures of lipid oxidation or glycation and actually increased levels of protein oxidation. The effect of sod-1 OE on life span was dependent on the DAF-16/FoxO transcription factor (TF) and, partially, on the heat shock TF HSF-1. Similarly, overexpression of sod-2 (major mitochondrial Mn-SOD) resulted in life-span extension that was daf-16 dependent. sod-1 OE increased steady-state hydrogen peroxide (H2O2) levels in vivo. However, co-overexpression of catalase did not suppress the life-span extension, arguing against H2O2 as a cause of longevity. sod-1 OE increased hsp-4 expression, suggesting increased endoplasmic reticulum (ER) stress. Moreover, longevity was partially suppressed by inactivation of ire-1 and xbp-1, mediators of the ER stress response. This suggests that high levels of SOD-1 protein may challenge protein-folding homeostasis, triggering a daf-16- and hsf-1-dependent stress response that extends life span. These findings imply that SOD overexpression increases C. elegans life span, not by removal of O2•−, but instead by activating longevity-promoting transcription factors., Highlights ► sod-1 over-expression increases levels of cellular ROS and of molecular damage. ► sod-1 over-expression effects on lifespan are dependent on DAF-16 (FoxO) and HSF-1. ► Increased lifespan is partially suppressed by inactivation of ER stress mediators. ► sod-2 (MnSOD) over-expression effects on lifespan are dependent on DAF-16.