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Introduction: Despite radiotherapy being one of the major treatments for triple-negative breast cancer (TNBC), new molecular targets for its treatment are still required due to radioresistance. CDK2 plays a critical role in TNBC. However, the mechanism by which CDK2 promotes TNBC radioresistance remains to be clearly elucidated. Objectives: We aimed to elucidate the relationship between CDK2 and TRIM32 and the regulation mechanism in TNBC. Methods: We performed immunohistochemical staining to detect nuclear TRIM32, CDK2 and STAT3 on TNBC tissues. Western blot assays and PCR were used to detect the protein and mRNA level changes. CRISPR/Cas9 used to knock out CDK2. shRNA-knockdown and transfection assays also used to knock out target genes. GST pull-down analysis, immunoprecipitation (IP) assay and in vitro isomerization analysis also used. Tumorigenesis studies also used to verify the results in vitro. Results: Herein, tripartite motif-containing protein 32 (TRIM32) is revealed as a substrate of CDK2. Radiotherapy promotes the binding of CDK2 and TRIM32, thus leading to increased CDK2-dependent phosphorylation of TRIM32 at serines 328 and 339. This causes the recruitment of PIN1, involved in cis–trans isomerization of TRIM32, resulting in importin α3 binding to TRIM32 and contributing to its nuclear translocation. Nuclear TRIM32 inhibits TC45-dephosphorylated STAT3, Leading to increased transcription of STAT3 and radioresistance in TNBC. These results were validated by clinical prognosis confirmed by the correlative expressions of the critical components of the CDK2/TRIM32/STAT3 signaling pathway. Conclusions: Our findings demonstrate that regulating the CDK2/TRIM32/STAT3 pathway is a promising strategy for reducing radioresistance in TNBC.

Duchenne muscular dystrophy (DMD) is an X-linked recessive progressive degenerative disease of skeletal muscle, characterized by intramuscular inflammation, muscle regeneration disorder and replacement of muscle with fibroadipose tissue. DMD is caused by the absence of normal dystrophy. Impaired self-renew ability and limited differentiation capacity of satellite cells are proved as main reasons for muscle regeneration failure. The deficiency of estrogen impedes the process of muscle regeneration. However, the role of estrogen receptor β (ERβ) in muscle regeneration is still unclear. This study aims to investigate the role and the pharmacological effect of ERβ activation on muscle regeneration in mdx mice. This study showed that mRNA levels of ERβ and myogenic-related genes both witnessed increasing trends in dystrophic context. Our results revealed that treatment with selective ERβ agonist (DPN, diarylpropionitrile) significantly increased myogenic differentiation 1 (MyoD-1) level and promoted muscle regeneration in mdx mice. Similarly, in mdx mice with muscle-specific estrogen receptor α (ERα) ablation, DPN treatment still promoted muscle regeneration. Moreover, we demonstrated that myoblasts differentiation was accompanied by raised nuclear accumulation of ERβ. DPN treatment augmented the nuclear accumulation of ERβ and, thus, contributed to myotubes formation. One important finding was that forkhead box O3A (FOXO3A), as a pivotal transcription factor in Myod-1 transcription, participated in the ERβ-promoted muscle regeneration. Overall, we offered an interesting explanation about the crucial role of ERβ during myogenesis.

Pancreatic ductal adenocarcinoma (PDAC) poses a formidable challenge in oncology due to its limited treatment options and poor long-term survival rates. Our previous work identified MG53, a member of the tripartite motif family protein (TRIM72), as a key player in tissue repair with potential applications in regenerative medicine. Despite the focus on MG53’s cytosolic functions, its nuclear role in suppressing pancreatic cancer remains unknown. Through orthotopic and subcutaneous transplantation studies in mice, we observed enhanced tumor growth in MG53-deficient mice compared to wild-type counterparts. The overexpression of KIF11, a motor protein crucial for cell mitosis regulation, has been linked to the aggressive proliferation of pancreatic cancer cells. Confocal imaging confirmed MG53′s presence in the nucleus of human pancreatic cancer cells, while functional assays demonstrated its impact on KIF11 expression and subsequent cell proliferation. Mechanistically, we revealed MG53′s transcriptional control over KIF11, leading to cell cycle arrest. Our findings position MG53 as a promising tumor suppressor in PDAC, offering a novel avenue for therapeutic intervention by regulating KIF11 expression.

Long-term denervation-induced atrophy and fibrosis of skeletal muscle due to denervation leads to poor recovery of muscle function. Studies have shown that the transforming growth factor-β1 (TGF-β1)-Smad signaling pathway plays a central role in muscle atrophy and fibrosis. Recent studies demonstrate the role of microRNAs (miRs) in various pathological conditions, including muscle regeneration. miR-21 has been shown to play a dynamic role in inflammatory responses and in accelerating injury responses to fibrosis. We used both RNA sequencing and quantitative RT-PCR strategies to examine the alternations of miRNAs during denervation-induced gastrocnemius muscle atrophy and fibrosis. Our data showed that MiR-21 was upregulated in denervated gastrocnemius muscle tissue, and TGF-β1treatment increased miR-21 expression. Inhibition of miR-21 reduced gastrocnemius muscle fibrosis and significantly downregulated the expression of p-SMAD2/3 and the fibrosis-associated markers TGF-β1, connective tissue growth factor, alpha smooth muscle actin. Masson's trichrome staining revealed that atrophy and fibrosis in gastrocnemius muscle tissue were reduced in the miR-21 inhibition group compared to the control group. We confirmed that SMAD7 is a direct target of miR-21 using a dual luciferase assay. Furthermore, Immunofluorescence and Western blot analyses revealed that miR-21 inhibition reduced SMAD2/3 phosphorylation and nuclear translocation. While SMAD7-siRNA abolished the effect. Consequently, the discovery that miR-21 regulates the atrophy and fibrosis of the gastrocnemius muscle offers a possible therapeutic approach for their management.

Cardiac hypertrophy is initially an adaptive response to physiological and pathological stimuli. Although pathological myocardial hypertrophy is the main cause of morbidity and mortality, our understanding of its mechanism is still weak. NFAT3 (nuclear factor of activated T-cell-3) is a member of the nuclear factor of the activated T cells (NFAT) family. NFAT3 plays a critical role in regulating the expression of cardiac hypertrophy genes by inducing their transcription. Recently, accumulating evidence has indicated that NFAT3 is a potent regulator of the progression of cardiac hypertrophy. This review, for the first time, summarizes the current studies on NFAT3 in cardiac hypertrophy, including the pathophysiological processes and the underlying pathological mechanism, focusing on the nuclear translocation and transcriptional function of NFAT3. This review will provide deep insight into the pathogenesis of cardiac hypertrophy and a theoretical basis for identifying new therapeutic targets in the NFAT3 network.

Summary: The dynamic subcellular localization of ERK1/2 plays an important role in regulating cell fate. Differentiation of mouse embryonic stem cells (mESCs) involves inductive stimulation of ERK1/2, and therefore, inhibitors of the ERK cascade are used to maintain pluripotency. Interestingly, we found that in pluripotent mESCs, ERK1/2 do not translocate to the nucleus either before or after stimulation. This inhibition of nuclear translocation may be dependent on a lack of stimulated ERK1/2 interaction with importin7 rather than a lack of ERK1/2 phosphorylation activating translocation. At late stages of naive-to-primed transition, the action of the translocating machinery is restored, leading to elevation in ERK1/2-importin7 interaction and their nuclear translocation. Importantly, forcing ERK2 into the naive cells’ nuclei accelerates their early differentiation, while prevention of the translocation restores stem cells’ pluripotency. These results indicate that prevention of nuclear ERK1/2 translocation serves as a safety mechanism for keeping pluripotency of mESCs.

Duchenne muscular dystrophy (DMD) is an X-linked recessive progressive degenerative disease of skeletal muscle, characterized by intramuscular inflammation, muscle regeneration disorder and replacement of muscle with fibroadipose tissue. DMD is caused by the absence of normal dystrophy. Impaired self-renew ability and limited differentiation capacity of satellite cells are proved as main reasons for muscle regeneration failure. The deficiency of estrogen impedes the process of muscle regeneration. However, the role of estrogen receptor β (ERβ) in muscle regeneration is still unclear. This study aims to investigate the role and the pharmacological effect of ERβ activation on muscle regeneration in mdx mice. This study showed that mRNA levels of ERβ and myogenic-related genes both witnessed increasing trends in dystrophic context. Our results revealed that treatment with selective ERβ agonist (DPN, diarylpropionitrile) significantly increased myogenic differentiation 1 (MyoD-1) level and promoted muscle regeneration in mdx mice. Similarly, in mdx mice with muscle-specific estrogen receptor α (ERα) ablation, DPN treatment still promoted muscle regeneration. Moreover, we demonstrated that myoblasts differentiation was accompanied by raised nuclear accumulation of ERβ. DPN treatment augmented the nuclear accumulation of ERβ and, thus, contributed to myotubes formation. One important finding was that forkhead box O3A (FOXO3A), as a pivotal transcription factor in Myod-1 transcription, participated in the ERβ-promoted muscle regeneration. Overall, we offered an interesting explanation about the crucial role of ERβ during myogenesis., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Peptidylarginine deiminase 4 (PAD4), a Ca2+-dependent enzyme, catalyzes the conversion of arginine to citrulline and has been strongly associated with many malignant tumors. However, the molecular mechanisms of PAD4 in the development and progression of colorectal cancer (CRC) remain unclearly defined. In our study, PAD4 expression was increased in CRC tissues and cells, and was closely related to tumor size, lymph node metastasis. Moreover, the transcription factor KLF9 directly bound to PADI4 gene promoter, leading to overexpression of PAD4 in CRC cells, which augmented cell growth and migration. We revealed that PAD4 interacted with and citrullinated glycogen synthase kinase-3β (GSK3β) in CRC cells, and GSK3β Arg-344 was the dominating PAD4-citrullination site. Furthermore, IgL2 and catalytic domains of PAD4 directly bound to the kinase domain of GSK3β in CRC cells. Mechanistically, PAD4 promoted the transport of GSK3β from the cytoplasm to the nucleus, thereby increasing the ubiquitin-dependent proteasome degradation of nuclear cyclin-dependent kinase inhibitor 1 (CDKN1A). Our study is the first to reveal the details of a critical PAD4/GSK3β/CDKN1A signaling axis for CRC progression, and provides evidence that PAD4 is a potential diagnosis biomarker and therapeutic target in CRC.

Airway mucus hypersecretion is a vital pathophysiologic feature in chronic obstructive pulmonary disease (COPD) patients in which airflow limitation result, and it is key to strategizing in the management of COPD. To investigate the mechanisms underlying the action of interleukin-6 neutralizing antibody (IL-6 Ab) in attenuating airway mucus hypersecretion in COPD, human and mouse primary bronchial epithelial cells from COPD patients and mice were isolated, human organoid model of trachea was established and all treated with IL-6 and/or IL-6 Ab. The differential expression of Muc5ac and Nrf2 were determined in pDHBE compared to pNHBE cells via high-throughput sequencing of transcriptome. The serum concentration of Muc5ac was significantly elevated and positively correlated with IL-6 in COPD patients using ELISA, and the excessive mucus secretion was observed in the trachea of COPD patients using HE, AB-PAS and IHC staining. The levels of Muc5ac were significantly elevated in the IL-6-treated group, and diminished with IL-6 Ab treatment, both in vitro and in the organoid model using qRT-PCR, WB and IF. The expression levels of protein Muc5ac were significantly reduced in cells transfected with the IL-6 small interfering RNA (siRNA-IL-6), which was in contrast to the levels of protein Nrf2, and the protective effects of IL-6 Ab were inhibited in cells transfected with Nrf2 short hairpin RNA (shRNA-Nrf2). IL-6 Ab significantly attenuated hypersecretion of airway mucus by inducing nuclear translocation of Nrf2 in COPD. These findings indicated that IL-6 Ab may constitute a novel therapeutic agent for IL-6-induced airway mucus hypersecretion by improving airflow limitation in COPD patients.

Hexavalent chromium [Cr(VI)] is a pervasive environmental pollutant that can enter the body through a variety of routes and cause organ toxicity, genetic damage, and cancer. Cr(VI)-induced apoptosis is a toxicant mechanism of Cr(VI). Studies have shown that Cr(VI) can induce p53-independent apoptosis, but the mechanisms are not fully understood. The intracellular calcium concentration affects cellular life. Apoptosis-inducing factor (AIF), a caspase-independent apoptotic effector, can induce DNA degradation. Using p53-null Hep3B cells, we investigated the effects of cytoplasmic calcium homeostasis and AIF on Cr(VI)-induced apoptosis. We found that 20 µM of Cr(VI) induced DNA damage and mitochondrial permeability transition pore (MPTP) openings, causing calcium overload that was accompanied by decreased Ca2+-Mg2+-ATPase and Na+-K+-ATP activities, downregulation of calmodulin (CaM) and Ca2+/CaM-dependent protein kinase II (CAMKII) mRNA, and increased expression of p-CaMKII/CaMKII protein. After treatment with calcium chelating agent BAPTA-AM, Cr(VI)-induced DNA damage, calcium overload, and apoptosis were reduced. AIF was released from the mitochondria and translocated into the nuclei. As the Cr(VI) treatment time progressed, the mRNA and protein expression of B cell lymphoma 2 (Bcl-2) and heat-shock protein 70 (HSP70) decreased, whereas the mRNA and protein expression of Bcl-2-associated X (Bax), cyclophilin A (CypA), and endonuclease G (EndoG) were upregulated. These results indicated that Cr(VI)-induced apoptosis of Hep3B cells (p53-null) was closely associated with calcium overload, and was accompanied by the activation of Ca2+/CaM/CaMKII signaling pathway. Besides, Cr(VI) triggered AIF nuclear translocation in Hep3B cells, accompanied by the changes in the levels of apoptosis-associated factors. These results provide additional experimental evidence of the molecular mechanisms involved in Cr(VI)-induced p53-independent apoptosis.

The imbalance in the phosphorylation and the dephosphorylation of proteins leads to various diseases. Therefore, in vivo, the functions of protein kinases and protein phosphatases are strictly regulated. Mg 2+ /Mn 2+ -dependent protein phosphatase PPM1M has been implicated in immunity and cancer; however, the regulation mechanism remains unknown. In this study, we show that PPM1M is regulated in different ways by multiple phosphorylation. PPM1M has four Ser/Thr-Pro motifs (Ser27, Ser43, Ser60, and Thr254) that are recognized by proline-directed kinases, and Ser60 was found to be phosphorylated by cyclin-dependent kinase 5 (CDK5) in the cell. The phospho-mimetic mutation of Ser27 and Ser43 in the N-terminal domain suppresses the nuclear localization of PPM1M and promotes its accumulation in the cytoplasm. The phospho-mimetic mutation of Ser60 decreases PPM1M activity; conversely, the phospho-mimetic mutation of Thr254 increases PPM1M activity. These results suggest that the subcellular localization and phosphatase activity of PPM1M are regulated by protein kinases, including CDK5, via phosphorylation at multiple sites. Thus, PPM1M is differentially regulated by proline-directed kinases, including CDK5., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Innate immunity is the first line of defense against viral pathogens. Retinoic Acid-Inducible Gene 1 (RIG-I) is a pattern recognition receptor that recognizes virus-associated double-stranded RNA and initiates the interferon responses. Besides signal transduction, RIG-I exerts direct antiviral functions to displace viral proteins on dsRNA via its Helicase activity. Nevertheless, this effector-like activity of RIG-I against herpesviruses remains largely unexplored. It has been previously reported that herpesviruses deamidate RIG-I, resulting in the abolishment of its Helicase activity and signal transduction. In this study, we discovered that RIG-I possessed signaling-independent antiviral activities against murine gamma herpesviruses 68 (γHV68, murid herpesvirus 4). Importantly, a Helicase-dead mutant of RIG-I (K270A) demonstrated comparable inhibition on herpesviruses lytic replication, indicating that this antiviral activity is Helicase-independent. Mechanistically, RIG-I bound the Replication and Transcription Activator (RTA) and diminished its nuclear localization to repress viral transcription. We further demonstrated that RIG-I blocked the nuclear translocation of ORF21 (Thymidine Kinase), ORF75c (vGAT), both of which form a nuclear complex with RTA and RNA polymerase II (Pol II) to facilitate viral transcription. Moreover, RIG-I retained ORF59 (DNA processivity factor) in the cytoplasm to repress viral DNA replication. Altogether, we illuminated a previously unidentified, Helicase-independent effector-like function of RIG-I against γHV68, representing an exquisite host strategy to counteract viral manipulations on innate immune signaling. IMPORTANCE: Retinoic acid-inducible gene I (RIG-I), a member of DExD/H box RNA helicase family, functions as a key pattern recognition receptor (PRR) responsible for the detection of intracellular double-stranded RNA (dsRNA) from virus-infected cells and induction of type I interferon (IFN) responses. Nevertheless, our understanding of the helicase-independent effector-like activity of RIG-I against virus infection, especially herpesvirus infection, remains largely unknown. Herein, by deploying murine gamma herpesviruses 68 (γHV68) as a model system, we demonstrated that RIG-I possessed an interferon and helicase-independent antiviral activity against γHV68 via blocking the nuclear trafficking of viral proteins, which concomitantly repressed the viral early transcription and genome replication thereof. Our work illuminates a previously unidentified antiviral strategy of RIG-I against herpesvirus infection., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Skin damage caused by exposure to ultraviolet (UV) light has been well documented clinically and histologically. Dopamine receptor D2 (DRD2) possesses various biological functions. However, no study has reported the possible association of DRD2 with UV-induced skin damage. We established DRD2 conditional knockout and UV damage models in this work. The results showed that DRD2 played an important role in the treatment of UV-induced skin damage. The findings of the molecular mechanism study revealed that the internalization of DRD2 after activation can stabilize nuclear factor erythroid 2-related factor 2 (Nrf2). However, the entry of Nrf2 into the nucleus did not increase. We prepared and characterized hyaluronic acid (HA)-coated mesoporous polydopamine (MPDA) nanoparticles (H@P@M). HA facilitated skin epidermal penetration of the nanoparticles to reach the site of inflammation smoothly. Meanwhile, MPDA activated DRD2 internalization to stabilize Nrf2. The release of prunetin inhibited the interaction of Kelch-like ECH-associated protein 1 with Nrf2 and promoted the nuclear translocation of Nrf2. In summary, this study unveiled that in skin inflammation, H@P@M activated and internalized DRD2, which subsequently formed a protein complex with arrestin beta 1-ubiquitin specific protease 8 (USP8)-Nrf2. Deubiquitination was performed to stabilize Nrf2 while promoting the nuclear translocation of Nrf2 to exert anti-inflammatory and antioxidant functions. STATEMENT OF SIGNIFICANCE: Skin is the body's largest physical barrier, always protecting the body from the interference of the external environment. However, excessive exposure to ultraviolet rays in the sun can cause skin inflammation, leading to skin erythema, itching, edema and pain, which can be troublesome in our daily lives. The complex mechanism of skin inflammation caused by ultraviolet radiation has not been fully clarified. In this study, the role of DRD2 in UV-induced skin inflammation was explored, and nano-composite particles HA@Prunetin@MPDA, which act on multiple targets in the anti-inflammatory pathway of DRD2, were developed to maximize the effect of the drug. It provides a new way to treat skin inflammation caused by UV., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2023 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Fat-specific protein 27 (FSP27), a member of the cell death-inducing DNA fragmentation factor α-like effector (Cide) family, is highly expressed in adipose tissues and is a lipid droplet (LD)-associated protein that induces the accumulation of LDs. Using a yeast two-hybrid system to examine potential interactions of FSP27 with other proteins, a direct interaction with the N-terminal region of nuclear factor of activated T cells 5 (NFAT5) was identified. NFAT5 is a transcription factor that induces osmoprotective and inflammatory genes after its translocation to the nucleus. The interaction between FSP27 and NFAT5 was confirmed by bimolecular fluorescence complementation and coimmunoprecipitation. Using immunocytochemistry, NFAT5 is detected in the cytoplasm and in the nucleus under isotonic conditions; however, overexpression of FSP27 inhibited the hypertonic-induced nuclear translocation of NFAT5. Consistent with the suppression of NFAT5 nuclear translocation, in cells transfected with a reporter construct containing the NFAT5 response element from the monocyte chemoattractant protein 1 (MCP1) promoter, FSP27 overexpression repressed hypertonic-induced luciferase activity and the expression of NFAT5 target genes. Knockdown of FSP27 in differentiated 3T3-L1 adipocytes increased the NFAT5-mediated rise in MCP1. These results suggest that FSP27 not only modulates LD homeostasis but also modulates the response to osmotic stress via a physical interaction with NFAT5 at the LD surface.

The non-β-oxidisable tetradecylthioacetic acid (TTA) is incorporated into cellular membranes when C3H/10T1/2 cells are cultured in TTA-containing medium. We here demonstrate that this alteration in cellular membranes affect the nuclear translocation of proteins involved in signal transduction. Analysis of cellular fatty acid composition shows that TTA and TTA:1n-8 constitute approximately 40 mol% of total fatty acids in cellular/nuclear membranes. Activation of c-fos expression is significantly inhibited in TTA-treated cells but the enzymatic activation of mitogen activated protein kinase (ERK) is not affected. Immunofluorescence and confocal microscopy studies demonstrate that in mitogene-stimulated TTA-treated cells, the translocation of phosphorylated ERK1/2, protein kinase Cα (PKCα), and PKCβ1 from the cytoplasm into the nucleus is considerably decreased and delayed. Concomitant with a decreased nuclear import, ERK1/2 dephosphorylation is decreased in TTA-treated cells. There is no TTA-induced inhibition of nuclear import of proteins with a classical nuclear localization signal (NLS), as seen by in vitro nuclear import experiments of BSA fused to the NLS from SV40 large T, or in vivo studies of hnRNP A1 nuclear import. The expression levels of Importin α, Importin β, Importin 7, and NTF2 are not altered in the TTA-treated cells.Taken together, our data indicate that TTA treatment causes changes in cellular fatty acid composition that negatively affect NLS-independent mechanisms of protein translocation through the nuclear pore complex.

Ischemic preconditioning (IPC) is a potential intervention known to protect the heart against ischemia/reperfusion injury, but its role in the no-reflow phenomenon that follows reperfusion is unclear. Dihydrotanshinone I (DT) is a natural compound and this study illustrates its role in cardiac ischemic injury from the aspect of IPC. Pretreatment with DT induced modest ROS production and protected cardiomyocytes against oxygen and glucose deprivation (OGD), but the protection was prevented by a ROS scavenger. In addition, DT administration protected the heart against isoprenaline challenge. Mechanistically, PKM2 reacted to transient ROS via oxidization at Cys423/Cys424, leading to glutathionylation and nuclear translocation in dimer form. In the nucleus, PKM2 served as a co-factor to promote HIF-1 α -dependent gene induction, contributing to adaptive responses. In mice subjected to permanent coronary ligation, cardiac-specific knockdown of Pkm2 blocked DT-mediated preconditioning protection, which was rescued by overexpression of wild-type Pkm2 , rather than Cys423/424-mutated Pkm2 . In conclusion, PKM2 is sensitive to oxidation, and subsequent glutathionylation promotes its nuclear translocation. Although IPC has been viewed as a protective means against reperfusion injury, our study reveals its potential role in protection of the heart from no-reflow ischemia., Competing Interests: The authors declare no conflicts of interest., (© 2022 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.)

Wnt family proteins are secreted glycolipoproteins that signal through multitude of signal transduction pathways. The Wnt-pathways are conserved and critical in all metazoans. They are essential for embryonic development, organogenesis and homeostasis, and associated with many diseases when defective or deregulated. Wnt signaling pathways comprise the canonical Wnt pathway, best known for its stabilization of β-catenin and associated nuclear β-catenin activity in gene regulation, and several non-canonical signaling branches. Wnt-Planar Cell Polarity (PCP) signaling has received the most attention among the non-canonical Wnt pathways. The relationship of cilia to Wnt-signaling is complex. While it was suggested that canonical Wnt signaling requires cilia this notion was always challenged by results suggesting the opposite. Recent developments provide insight and clarification to the relationship of Wnt signaling pathways and cilia. First, it has been now demonstrated that while ciliary proteins, in particular the IFT-A complex, are required for canonical Wnt/β-catenin signaling, the cilium as a structure is not. In contrast, recent work has defined a diverged canonical signaling branch (not affecting β-catenin) to be required for ciliary biogenesis and cilia function. Furthermore, the non-canonical Wnt-PCP pathway does not affect cilia biogenesis per se, but it regulates the position of cilia within cells in many cell types, possibly in all cells where it is active, with cilia being placed near the side of the cell that has the Frizzled-Dishevelled complex. This Wnt/PCP feature is conserved with both centrioles and basal bodies/cilia being positioned accordingly, and it is also used to align mitotic spindles within the Wnt-PCP polarization axis. It also coordinates the alignment of cilia in multiciliated cells. This article addresses these new insights and different links and relationships between cilia and Wnt signaling., (Copyright © 2023. Published by Elsevier Inc.)

Peptidylarginine deiminase 4 (PAD4), a Ca 2+ -dependent enzyme, catalyzes the conversion of arginine to citrulline and has been strongly associated with many malignant tumors. However, the molecular mechanisms of PAD4 in the development and progression of colorectal cancer (CRC) remain unclearly defined. In our study, PAD4 expression was increased in CRC tissues and cells, and was closely related to tumor size, lymph node metastasis. Moreover, the transcription factor KLF9 directly bound to PADI4 gene promoter, leading to overexpression of PAD4 in CRC cells, which augmented cell growth and migration. We revealed that PAD4 interacted with and citrullinated glycogen synthase kinase-3β (GSK3β) in CRC cells, and GSK3β Arg-344 was the dominating PAD4-citrullination site. Furthermore, IgL2 and catalytic domains of PAD4 directly bound to the kinase domain of GSK3β in CRC cells. Mechanistically, PAD4 promoted the transport of GSK3β from the cytoplasm to the nucleus, thereby increasing the ubiquitin-dependent proteasome degradation of nuclear cyclin-dependent kinase inhibitor 1 (CDKN1A). Our study is the first to reveal the details of a critical PAD4/GSK3β/CDKN1A signaling axis for CRC progression, and provides evidence that PAD4 is a potential diagnosis biomarker and therapeutic target in CRC., Competing Interests: Declaration of Competing Interest The authors declare no potential conflict of interest., (Copyright © 2022. Published by Elsevier Inc.)

The Dishevelled gene was first identified in Drosophila mutants with disoriented hair and bristle polarity and subsequent work has now demonstrated its importance in critical and diverse aspects of biology. Since those early discoveries, Dishevelled has been shown to coordinate a plethora of developmental and cellular processes that range from controlling cell polarity during gastrulation to partnering with chromatin modifying enzymes to regulate histone methylation at genomic loci. While the role of DVL in development is well-respected and the cytosolic function of DVL has been studied more extensively, its nuclear role continues to remain murky. In this review we highlight some of the seminal discoveries that have contributed to the field, but the primary focus is to discuss recent advances with respect to the nuclear role of Dishevelled. This nuclear function of Dishevelled is a dimension which is proving to be increasingly important yet remains enigmatic., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The Wnt family of secreted glycolipo-proteins signals through multiple signal transduction pathways and is essential for embryonic development and organ development and homeostasis. The Wnt-pathways are conserved and critical in all metazoans. Wnt signaling pathways comprise the canonical Wnt/β-catenin pathway and several non-canonical signaling branches, of which Wnt-Planar Cell Polarity (PCP) signaling and the Wnt/Calcium pathway have received the most attention and are best understood. nterestingly, all Wnt-pathways have a nuclear signaling branch and also can affect many cellular processes independent of its nuclear transcriptional regulation. Canonical Wnt/β-catenin signaling is the most critical for a nuclear transcriptional response, in both development and disease, yet the mechanism(s) on how the "business end" of the pathway, β-catenin, translocates to the nucleus to act as co-activator to the TCF/Lef transcription factor family still remains obscure. Here we discuss and compare the very different strategies on how the respective Wnt signaling pathways activate a nuclear transcriptional response. We also highlight some recent new insights into how β-catenin is translocated to the nucleus via an IFT-A, Kinesin-2, and microtubule dependent mechanism and how this aspect of canonical Wnt-signaling uses ciliary proteins in a cilium independent manner, conserved between Drosophila and mammalian cells., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Rescuing cells from stress damage emerges a potential therapeutic strategy to combat myocardial infarction. Protocatechuic aldehyde (PCA) is a major phenolic acid in Chinese herb Danshen ( Salvia miltiorrhiza root). This study investigated whether PCA regulated nuclear pyruvate kinase isoform M2 (PKM2) function to protect cardiomyocytes. In rats subjected to isoprenaline, PCA attenuated heart injury and protected cardiomyocytes from apoptosis. Through DARTS and CETSA assays, we identified that PCA bound and promoted PKM2 nuclear translocation in cardiomyocytes exposed to oxygen/glucose deprivation (OGD). In the nucleus, PCA increased the binding of PKM2 to β -catenin via preserving PKM2 acetylation, and the complex, in cooperation with T-cell factor 4 (TCF4), was required for transcriptional induction of genes encoding anti-apoptotic proteins, contributing to rescuing cardiomyocyte survival. In addition, PCA ameliorated mitochondrial dysfunction and prevented mitochondrial apoptosis dependent on PKM2. Consistently, PCA increased the binding of PKM2 to β -catenin, improved heart contractive function, normalized heart structure and attenuated oxidative damage in mice subjected to artery ligation, but the protective effects were lost in Pkm2 -deficient heart. Together, we showed that PCA regulated nuclear PKM2 function to rescue cardiomyocyte survival via β -catenin/TCF4 signaling cascade, suggesting the potential of pharmacological intervention of PKM2 shuttle to protect the heart., (© 2021 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.)

Cell migration is a key step of cancer metastasis, immune-cell navigation, homing of stem cells and development. What adds complexity to it is the heterogeneity of the tissue environment that gives rise to a vast diversity of migratory mechanisms utilized by cells. A majority of cell motility mechanisms reported elsewhere largely converge in depicting the importance of the activity and complexity of actomyosin networks in the cell. In this review, we highlight the less discussed functional diversity of these actomyosin complexes and describe in detail how the major cellular actin-binding molecular motor proteins, nonmuscle myosin IIs are regulated and how they participate and mechanically reciprocate to changes in the microenvironment during cancer cell migration and tumor progression. Understanding the role of nonmuscle myosin IIs in the cancer cell is important for designing efficient therapeutic strategies to prevent cancer metastasis., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

PAC1-R is neuropeptide PACAP (pituitary adenylate cyclase activating polypeptide) preferring receptor mediates the antioxidant and cytoprotective effects of PACAP. It was found in this research that in both PAC1R-CHO cells with high expression of PAC1R-eGFP and retinal ganglion cells (RGC-5) with natural expression of PAC1-R, blue light and hydrogen peroxide (H 2 O 2 ) trigger the significant nuclear translocation of PAC1-R, and the nuclear translocation of PAC1-R was positive correlation with the up-regulation of expression level and promoter activity of PAC1-R its own, while red light worked much less efficiently than blue light. Reactive oxygen species (ROS) scavenger NAC (N-acetyl-L-cysteine) and palmitoylation inhibitor 2-bromopalmitate (2-BP) disturbed the nuclear shifting associated with the correlative up-regulation of PAC1 significantly, and PAC1-R mutant (M-PAC1-R) on Cys25/Ala25 displayed the significant decreased nuclear trafficking efficiency. Furthermore, the Western Blot results with the antibody raised against the C-terminal of PAC1-R showing PAC1-R in the nucleus was fragmentation hinting that C-terminal of PAC1-R with theoretical nuclear location signal (NLS) may be involved in activation of PAC1-R promoter in the nucleus. All above results indicated that PAC1-R makes the nuclear translocation to trigger the activation of PAC1-R itself promoter resulting into the up-regulation of of PAC1-R in response to the oxidative stress induced by blue light and ROS such as H 2 O 2 ., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The glucocorticoid receptor (GR) plays an important role in steroid-dependent regulation of metabolism, development, and the immune response in humans. Although GR is known to be activated by the binding of glucocorticoid, the mechanism of action is poorly understood. We investigated dimerization of GR in the cytoplasm and nuclear trans-localization in response to treatment with the ligand dexamethasone. GFP-tagged GR and FLAG-tagged GR were co-expressed in COS-1 cells, and cell lysates were subjected to co-immunoprecipitation assay with anti-GFP antibody to determine their dimerization. FLAG-GR was co-precipitated with GFP-GR in the cytoplasmic fraction of COS-1 cells. Treatment with the GR agonist dexamethasone significantly decreased the cytoplasmic interaction between FLAG- and GFP-GR, and significantly increased interaction of the GRs in the nuclear fraction. The two amino acids, Pro625 and Ile628 known to be located in GR-GR dimer interface, were mutated to alanine and the influence of the mutation on dimerization, ligand-dependent nuclear localization, and transcriptional activities were determined. Mutant GR showed a dramatic decrease in interaction in the cytoplasmic fraction and no detectable nuclear translocation in the presence or absence of dexamethasone. Furthermore, luciferase assays showed that mutant GR showed no detectable transcriptional activation via the GR-responsive DNA element (GRE) compared to the wild-type. Our results suggest that GR exists as a dimer in the cytoplasm and this dimerization may be essential for GRE-mediated transcriptional activation following ligand binding., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Hepatocellular carcinoma (HCC) is one of the most common human malignant tumors. It is known that in the cells of many cancers, including HCC, nuclear translocation and accumulation of YB-1 often indicates a poor prognosis. This nuclear translocation is induced by genotoxic stress resulting from administration of anticancer agents. Accumulation of YB-1 in the nucleus induces the expression of many genes related to cancer aggressiveness. Therefore, compounds capable of inhibiting anticancer drug-induced YB-1 nuclear translocation without cytotoxicity will be a powerful tool for cancer chemotherapy. In the present study, we found that indirubin derivative, 7-hydroxyindirubin strongly inhibited the actinomycin D-induced nuclear translocation of YB-1 more efficiently without showing cytotoxicity in HepG2, a human HCC cells. The compound successfully suppressed the nuclear YB-1-mediated expression of genes such as MDR1, MVP, EGFR, and CXCR4, which are known to disturb cancer treatment. 7-Hydroxyindirubin also increased the susceptibility of drug-resistant HepG2 cells to ActD. It was also demonstrated that 7-hydroxyindirubin inhibits the nuclear translocation of YB-1 with or without phosphorylation at the Ser102 residue., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Pyruvate kinase M2 isoform (PKM2) plays a key role in cancer progression through both metabolic and non-metabolic functions, thus it is recognized as a potential target for cancer diagnosis and treatment. In this study, we discovered a sulfonamide-dithiocarbamate compound 8a as a novel PKM2 activator from a random screening of an in-house compound library. Then, a series of lead compound 8a analogs were designed and synthesized for screening as potent PKM2 activators. Among them, compound 8b (AC 50  = 0.136 µM) and 8k (AC 50  = 0.056 µM) showed higher PKM2 activation activities than positive control NZT (AC 50  = 0.228 µM), and they (IC 50  < 1 µM) exhibited more significant anti-proliferative activities against human tumor cell lines than NZT (IC 50  > 10 µM). Especially, compound 8k inhibited the proliferation of multiple cancer cells, but showed little toxicity on normal cells. In addition, we found that compound 8k inhibit the colony formation of MCF7 cells. Western blot analysis demonstrated that 8k could reduce PKM2 nuclear localization and block the downstream signaling pathway of PKM2, resulting in suppression of tumor cell proliferation. Overall, compound 8k may be a promising candidate for further mechanistic investigation of PKM2 and cancer therapy., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Hexavalent chromium [Cr(VI)] is a pervasive environmental pollutant that can enter the body through a variety of routes and cause organ toxicity, genetic damage, and cancer. Cr(VI)-induced apoptosis is a toxicant mechanism of Cr(VI). Studies have shown that Cr(VI) can induce p53-independent apoptosis, but the mechanisms are not fully understood. The intracellular calcium concentration affects cellular life. Apoptosis-inducing factor (AIF), a caspase-independent apoptotic effector, can induce DNA degradation. Using p53-null Hep3B cells, we investigated the effects of cytoplasmic calcium homeostasis and AIF on Cr(VI)-induced apoptosis. We found that 20 µM of Cr(VI) induced DNA damage and mitochondrial permeability transition pore (MPTP) openings, causing calcium overload that was accompanied by decreased Ca 2+ -Mg 2+ -ATPase and Na + -K + -ATP activities, downregulation of calmodulin (CaM) and Ca 2+ /CaM-dependent protein kinase II (CAMKII) mRNA, and increased expression of p-CaMKII/CaMKII protein. After treatment with calcium chelating agent BAPTA-AM, Cr(VI)-induced DNA damage, calcium overload, and apoptosis were reduced. AIF was released from the mitochondria and translocated into the nuclei. As the Cr(VI) treatment time progressed, the mRNA and protein expression of B cell lymphoma 2 (Bcl-2) and heat-shock protein 70 (HSP70) decreased, whereas the mRNA and protein expression of Bcl-2-associated X (Bax), cyclophilin A (CypA), and endonuclease G (EndoG) were upregulated. These results indicated that Cr(VI)-induced apoptosis of Hep3B cells (p53-null) was closely associated with calcium overload, and was accompanied by the activation of Ca 2+ /CaM/CaMKII signaling pathway. Besides, Cr(VI) triggered AIF nuclear translocation in Hep3B cells, accompanied by the changes in the levels of apoptosis-associated factors. These results provide additional experimental evidence of the molecular mechanisms involved in Cr(VI)-induced p53-independent apoptosis., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Dysregulation of pseudogenes is involved in the progression of various types of cancer, including glioblastoma (GBM). Proliferation associated-2G4 (PA2G4) pseudogene 4 (PA2G4P4) has been shown to play an oncogenic role in bladder cancer development. Our study aimed to explore the role and mechanism of PA2G4P4 in GBM progression. PA2G4P4 and PA2G4 expression in GBM tissues was analyzed using the GEPIA database. Cell viability, apoptosis, and activities of caspase-3 and caspase-9 in GBM cells were explored by CCK-8, flow cytometry analysis, and colorimetric activity assay kits, respectively. GEPIA database showed that PA2G4P4 and PA2G4 were both upregulated in GBM tissues. PA2G4P4 expression was also boosted in GBM cells. Knockdown of PA2G4P4 or PA2G4 inhibited cell viability, induced apoptosis, and increased caspase-3 and caspase-9 activities in GBM cells. Data from UALCAN database showed that among top 15 genes correlated with PA2G4P4, PA2G4 had the highest correlation coefficient. Additionally, knockdown of PA2G4P4 inhibited PA2G4 expression and nuclear translocation in GBM cells. Overexpression of PA2G4 abolished the functions of PA2G4P4 knockdown on viability and apoptosis in GBM cells. Summarily, pseudogene PA2G4P4 promotes oncogene PA2G4 expression and nuclear translocation to affect cell viability and apoptosis in GBM cells., (Copyright © 2020. Published by Elsevier Inc.)

Optical manipulations are widely used to analyze neuronal functions in vivo. Blue light is frequently used to activate channelrhodopsins or LOV domains, although the degrees of its absorption and scattering are higher than those of longer wavelength light. High spatial resolution of optical manipulation is easily achieved in vitro, while the light is unevenly scattered and absorbed in tissues due to many factors. It is difficult to spatially measure a blue light transmission area in vivo. Here, we propose a genetic method to visualize blue light transmission in the brain and other organs using light-induced nuclear translocation of fluorescent proteins with a LOV domain. A light-inducible nuclear localization signal (LINuS) consists of a LOV2 domain fused with a nuclear localization signal (NLS). We confirmed that blue light illumination induced reversible translocation of NES-tdTomato-LINuS from the cytosol to the nucleus within 30 min in HEK293 cells. By employing a PHP.eb capsid that can penetrate the blood-brain barrier, retro-orbital sinus injection of adeno-associated virus (AAV) vectors induced scattered expression of nuclear export signal (NES)-tdTomato-LINuS in the brain. We confirmed that 30-min transcranial blue light illumination induced nuclear translocation of NES-tdTomato-LINuS in the cortex, the hippocampus, and even the paraventricular nucleus of the thalamus. We also found that mice exposed to blue light in a shaved abdominal area exhibited a substantial increase in nuclear translocation in the ventral surface lobe of the liver. These results provide a simple way to obtain useful information on light transmission in tissues without any transgenic animals or skillful procedures., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Mutations of PTEN-induced kinase 1 (PINK1) cause recessive familial Parkinson's disease. Cells lacking PINK1 display mitochondrial deficits and increased sensitivity to oxidative and proteasomal stress. It has been shown that the 52-kDa (mature) form of PINK1 in the cytoplasm mitigates proteasomal stress-induced cell death by enhancing aggresomes formation and autophagy. Here we newly demonstrate that proteasome dysfunction triggers mono-ubiquitination and nuclear translocation of mature PINK1. Enhancing PINK1 mono-ubiquitination by two different means increased nuclear accumulation of PINK1 independent of proteasome inhibition. Moreover, we show that PINK1 harbors a hitherto unknown nuclear export sequence (NES) in its C-terminus. Blocking CRM1-dependent nuclear export with leptomycin B augmented PINK1 levels in the nucleus of MG132-treated cells but not in normal cells. Overall, these results show that proteasomal stress-induced mono-ubiquitination of PINK1 mediates PINK1 nuclear translocation, while PINK1 is excluded from the nucleus of healthy cells via its NES. Therefore, mature PINK1 may have a nuclear function in cells under proteasomal stress., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Rewiring glucose metabolism, termed as Warburg effect or aerobic glycolysis, is a common signature of cancer cells to meet their high energetic and biosynthetic demands of rapid growth and proliferation. Pyruvate kinase M2 isoform (PKM2) is a key player in such metabolic reshuffle, which functions as a rate-limiting glycolytic enzyme in the cytosol of highly-proliferative cancer cells. During the recent decades, PKM2 has been extensively studied in non-canonical localizations such as nucleus, mitochondria, and extracellular secretion, and pertained to novel biological functions in tumor progression. Such functions of PKM2 open a new avenue for cancer researchers. This review summarizes up-to-date functions of PKM2 at various subcellular localizations of cancer cells and draws attention to the translocation of PKM2 from cytosol into the nucleus induced by posttranslational modifications. Moreover, PKM2 in tumor cells could have an important role in resistance acquisition processes against various chemotherapeutic drugs, which have raised a concern on PKM2 as a potential therapeutic target. Finally, we summarize the current status and future perspectives to improve the potential of PKM2 as a therapeutic target for the development of anticancer therapeutic strategies., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Aims: To investigate whether mucin 1 (MUC1) downregulation reduced the sensitivity of tumor necrosis factor-alpha (TNF-α)-induced bronchial epithelial cells to glucocorticoid-mediated necroptosis and explore the underlying mechanisms., Main Methods: The human lung bronchial epithelial cell line (16HBE) was transfected with small interfering RNA (siRNA) against MUC1 and then stimulated by TNF-α, where some cells were pretreated with dexamethasone. Flow cytometry was performed to analyze necroptosis in 16HBE cells, and western blot analysis was used to detect protein expression levels of MUC1, glucocorticoid receptor (GR)α, GRβ, NF-κB p65, phospho-p65 (p-p65), and histone deacetylase-2 (HDAC2). Additionally, nuclear translocation of MUC1 and GRα was assessed by immunofluorescence., Key Findings: We observed that MUC1 downregulation by siRNA significantly augmented TNF-α-induced necroptosis in 16HBE cells, and that dexamethasone showed impaired anti-necroptotic effects of MUC1 downregulation. Furthermore, we found that GRα nuclear translocation was inhibited in 16HBE cells with MUC1 downregulation, and that dexamethasone-mediated inhibition of p65 phosphorylation was lower in cells transfected with MUC1-siRNA compared to those transfected with negative control siRNA., Significance: Impaired GRα nuclear translocation and inhibited p-p65 expression might contribute to glucocorticoid resistance caused by MUC1 deficiency in TNF-α-induced necroptosis in 16HBE cells, and should be considered as a potential target for the development of novel therapeutics for asthma., (Copyright © 2019 Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology. Published by Elsevier Inc. All rights reserved.)

Glucocorticoid binding to the intracellular glucocorticoid receptor (GR) stimulates the translocation of the GR from the cytosol to the nucleus, which leads to the transactivation or transrepression of gene transcription. However, multiple lines of evidence suggest that glucocorticoid signaling can also be initiated from the plasma membrane. Here, we provide evidence for membrane-initiated glucocorticoid signaling by a membrane-impermeant dexamethasone-bovine serum albumin (Dex-BSA) conjugate, which induced GR nuclear trafficking in hypothalamic neurons in vitro and in vivo. The GR nuclear translocation induced by a membrane-impermeant glucocorticoid suggests trafficking of an unliganded GR. The membrane-initiated GR trafficking was not blocked by inhibiting ERK MAPK, p38 MAPK, PKA, Akt, Src kinase, or calcium signaling, but was inhibited by Akt activation. Short-term exposure of hypothalamic neurons to dexamethasone (Dex) activated the glucocorticoid response element (GRE), suggesting transcriptional transactivation, whereas exposure to the Dex-BSA conjugate failed to activate the GRE, suggesting differential transcriptional activity of the liganded compared to the unliganded GR. Microarray analysis revealed divergent transcriptional regulation by Dex-BSA compared to Dex. Together, our data suggest that signaling from a putative membrane glucocorticoid receptor induces the trafficking of unliganded GR to the nucleus, which elicits a pattern of gene transcription that differs from that of the liganded receptor. The differential transcriptional signaling by liganded and unliganded receptors may contribute to the broad range of genetic regulation by glucocorticoids, and may help explain some of the different off-target actions of glucocorticoid drugs., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: staphylococcal enterotoxin plays an important role in patients with glucocorticoid (GC)-resistant atopic dermatitis (AD), but the exact mechanism is not fully understood., Objective: The aim of this study was to investigate the mechanisms underlying the ability of staphylococcal enterotoxin B (SEB) to induce steroid insensitivity through impaired nuclear translocation of GRα in keratinocytes., Methods: The steroid-resistant AD induced by SEB was assessed by analyzing dermatitis score, dermal thickness, scratching behavior, infiltrating cells/HPF, levels of SEB-specific IgE and IgG2a antibody. In addition, dexamethasone (DEX)-induced GRα nuclear translocation and keratinocyte-derived cytokines and chemokines were analyzed in the lesional keratinocytes of AD and in HaCaT cells. Furthermore, the expressions of immunophilins FKBP51, FKBP52 and HSP90 responsive to GC in HaCaT cells were determined in the presence of SEB., Results: SEB dose-dependently diminished the inhibitory effect of DEX on dermatitis score, dermal thickness, scratching behavior, infiltrating cells/HPF, keratinocyte-derived cytokines and chemokines such as RANTES, MCP-1, TSLP and GM-CSF. In vivo and in vitro data showed that in the presence of DEX, SEB dose-dependently caused a marked decrease of GRα nuclear translocation in lesional keratinocytes of AD and in HaCaT cells. Importantly, in the presence of DEX, SEB increased the expression of FKBP51 and the product of keratinocyte-derived cytokines and chemokines in HaCaT cells., Conclusion: These results demonstrate that GC insensitivity by SEB in keratinocytes of AD is associated with impaired nuclear translocation of the GRα. Increased DEX-induced FKBP51 by SEB may contribute to accumulation of the GRα in cytoplasm of keratinocytes., (Copyright © 2018 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)

Insulin binding to its cell surface receptor (IR) activates a cascade of events leading to its biological effects. The Insulin-IR complex is rapidly internalized and then is either recycled back to the plasma membrane or sent to lysosomes for degradation. Although most of the receptor is recycled or degraded, a small amount may escape this pathway and migrate to the nucleus of the cell where it might be important in promulgation of receptor signals. In this study we explored the mechanism by which insulin induces IR translocation to the cell nucleus. Experiments were performed cultured L6 myoblasts, AML liver cells and 3T3-L1 adipocytes. Insulin treatment induced a rapid increase in nuclear IR protein levels within 2 to 5 min. Treatment with WGA, an inhibitor of nuclear import, reduced insulin-induced increases nuclear IR protein; IR was, however, translocated to a perinuclear location. Bioinformatics tools predicted a potential nuclear localization sequence (NLS) on IR. Immunofluorescence staining showed that a point mutation on the predicted NLS blocked insulin-induced IR nuclear translocation. In addition, blockade of nuclear IR activation in isolated nuclei by an IR blocking antibody abrogated insulin-induced increases in IR tyrosine phosphorylation and nuclear PKCδ levels. Furthermore, over expression of mutated IR reduced insulin-induced glucose uptake and PKB phosphorylation. When added to isolated nuclei, insulin induced IR phosphorylation but had no effect on nuclear IR protein levels. These results raise questions regarding the possible role of nuclear IR in IR signaling and insulin resistance., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Major depressive disorder (MDD) is a common, severe and recurrent psychiatric disorder worldwide; however, the underlying neuropathological mechanisms remain elusive. Histone deacetylases (HDACs) appear to play an essential role in depression. As the class III HDACs, Sirt1 and Sirt2 have attracted the most interest in the nervous system. Indeed, chronic stress decreased Sirt1 activity and down-regulated Sirt1 gene expression in MDD. Nevertheless, there is a paucity of literature on the role of Sirt2. To study the role of Sirt2 we established a MDD mouse model in wild type and Sirt2 knockout C57BL/6 mice using social defeat stress (SDS). We found that a lack of Sirt2 blocked the development of SDS-induced depressive-like behavior. Moreover, SDS led to Sirt2 phosphorylation in the amygdala without changing total Sirt2 levels, and blocking the phosphorylation of Sirt2 by CDK5 at serine residues 368 and 372 prevented SDS-induced depressive-like behavior and Sirt2 nuclear import. We also discovered that SDS-induced Sirt2 phosphorylation was involved in VTA-amygdala modulation using TetTag-pharmacogenetic method. These results suggest that CDK5 mediates phosphorylation of Sirt2 in the amygdala and contributes to the depressive-like behavior induced by SDS. This study highlights that inhibiting CDK5-dependent phosphorylation of Sirt2 at serine residues 368 and 372 by myristoylated membrane-permeabilising peptide (Sirt2-p), rather than using non-specific sirtuin inhibitors, may be a novel strategy for treating depression., (Copyright © 2017. Published by Elsevier B.V.)

Hepatocellular carcinoma (HCC) is a disease with poor prognosis. Nuclear accumulation of YB-1 is closely related to the malignancy of HCC. Treatment with anticancer agents often induces translocation of YB-1 from cytoplasm to nucleus and activates the expression of multidrug resistance gene 1 (MDR1). Therefore, any effective inhibitor of this phenomenon would be useful for cancer treatment. Here we examined various indirubin derivatives and found that indirubin 3'-oxime inhibits actinomycin D-induced nuclear transport of YB-1 and suppresses the activation of MDR1 gene expression in the human hepatocellular carcinoma cell line HepG2. Furthermore, use of both indirubin 3'-oxime and actinomycin D in combination increased the anticancer effect on HepG2 cells. Indirubin 3'-oxime is a novel and efficient inhibitor of anticancer agent-induced YB-1 nuclear translocation., (Copyright © 2017 Elsevier Inc. All rights reserved.)