Iridium(III) carbene complexes as potent girdin inhibitors against metastatic cancers.

Many cancer-driving protein targets remain undruggable due to a lack of binding molecular scaffolds. In this regard, octahedral metal complexes with unique and versatile three-dimensional structures have rarely been explored as inhibitors of undruggable protein targets. Here, we describe antitumor iridium(III) pyridinium-N-heterocyclic carbene complex 1a, which profoundly reduces the viability of lung and breast cancer cells as well as cancer patient-derived organoids at low micromolar concentrations. Compound 1a effectively inhibits the growth of non-small-cell lung cancer and triple-negative breast cancer xenograft tumors, impedes the metastatic spread of breast cancer cells, and can be modified into an antibody-drug conjugate payload to achieve precise tumor delivery in mice. Identified by thermal proteome profiling, an important molecular target of 1a in cellulo is Girdin, a multifunctional adaptor protein that is overexpressed in cancer cells and unequivocally serves as a signaling hub for multiple pivotal oncogenic pathways. However, specific small-molecule inhibitors of Girdin have not yet been developed. Notably, 1a exhibits high binding affinity to Girdin with a Kd of 1.3 µM and targets the Girdin-linked EGFR/AKT/mTOR/STAT3 cancer-driving pathway, inhibiting cancer cell proliferation and metastatic activity. Our study reveals a potent Girdin-targeting anticancer compound and demonstrates that octahedral metal complexes constitute an untapped library of small-molecule inhibitors that can fit into the ligand-binding pockets of key oncoproteins.

Conditional knockout of Shank3 in the ventral CA1 by quantitative in vivo genome-editing impairs social memory in mice.

Individuals with autism spectrum disorder (ASD) have a higher prevalence of social memory impairment. A series of our previous studies revealed that hippocampal ventral CA1 (vCA1) neurons possess social memory engram and that the neurophysiological representation of social memory in the vCA1 neurons is disrupted in ASD-associated Shank3 knockout mice. However, whether the dysfunction of Shank3 in vCA1 causes the social memory impairment observed in ASD remains unclear. In this study, we found that vCA1-specific Shank3 conditional knockout (cKO) by the adeno-associated virus (AAV)- or specialized extracellular vesicle (EV)- mediated in vivo gene editing was sufficient to recapitulate the social memory impairment in male mice. Furthermore, the utilization of EV-mediated Shank3-cKO allowed us to quantitatively examine the role of Shank3 in social memory. Our results suggested that there is a certain threshold for the proportion of Shank3-cKO neurons required for social memory disruption. Thus, our study provides insight into the population coding of social memory in vCA1, as well as the pathological mechanisms underlying social memory impairment in ASD.

Tumor promoting effect of PDLIM2 downregulation involves mitochondrial ROS, oncometabolite accumulations and HIF-1α activation.

BACKGROUND: Cancer is characterized by dysregulated cellular metabolism. Thus, understanding the mechanisms underlying these metabolic alterations is important for developing targeted therapies. In this study, we investigated the pro-tumoral effect of PDZ and LIM domain 2 (PDLIM2) downregulation in lung cancer growth and its association with the accumulation of mitochondrial ROS, oncometabolites and the activation of hypoxia-inducible factor-1 (HIF-1) α in the process. METHODS: Databases and human cancer tissue samples were analyzed to investigate the roles of PDLIM2 and HIF-1α in cancer growth. DNA microarray and gene ontology enrichment analyses were performed to determine the cellular functions of PDLIM2. Seahorse assay, flow cytometric analysis, and confocal microscopic analysis were employed to study mitochondrial functions. Oncometabolites were analyzed using liquid chromatography-mass spectrometry (LC-MS). A Lewis lung carcinoma (LLC) mouse model was established to assess the in vivo function of PDLIM2 and HIF-1α. RESULTS: The expression of PDLIM2 was downregulated in lung cancer, and this downregulation correlated with poor prognosis in patients. PDLIM2 highly regulated genes associated with mitochondrial functions. Mechanistically, PDLIM2 downregulation resulted in NF-κB activation, impaired expression of tricarboxylic acid (TCA) cycle genes particularly the succinate dehydrogenase (SDH) genes, and mitochondrial dysfunction. This disturbance contributed to the accumulation of succinate and other oncometabolites, as well as the buildup of mitochondrial reactive oxygen species (mtROS), leading to the activation of hypoxia-inducible factor 1α (HIF-1α). Furthermore, the expression of HIF-1α was increased in all stages of lung cancer. The expression of PDLIM2 and HIF-1α was reversely correlated in lung cancer patients. In the animal study, the orally administered HIF-1α inhibitor, PX-478, significantly reduces PDLIM2 knockdown-promoted tumor growth. CONCLUSION: These findings shed light on the complex action of PDLIM2 on mitochondria and HIF-1α activities in lung cancer, emphasizing the role of HIF-1α in the tumor-promoting effect of PDLIM2 downregulation. Additionally, they provide new insights into a strategy for precise targeted treatment by suggesting that HIF-1α inhibitors may serve as therapy for lung cancer patients with PDLIM2 downregulation.

Spa2 remodels ADP-actin via molecular condensation under glucose starvation.

Actin nucleotide-dependent actin remodeling is essential to orchestrate signal transduction and cell adaptation. Rapid energy starvation requires accurate and timely reorganization of the actin network. Despite distinct treadmilling mechanisms of ADP- and ATP-actin filaments, their filament structures are nearly identical. How other actin-binding proteins regulate ADP-actin filament assembly is unclear. Here, we show that Spa2 which is the polarisome scaffold protein specifically remodels ADP-actin upon energy starvation in budding yeast. Spa2 triggers ADP-actin monomer nucleation rapidly through a dimeric core of Spa2 (aa 281-535). Concurrently, the intrinsically disordered region (IDR, aa 1-281) guides Spa2 undergoing phase separation and wetting on the surface of ADP-G-actin-derived F-actin and bundles the filaments. Both ADP-actin-specific nucleation and bundling activities of Spa2 are actin D-loop dependent. The IDR and nucleation core of Spa2 are evolutionarily conserved by coexistence in the fungus kingdom, suggesting a universal adaptation mechanism in the fungal kingdom in response to glucose starvation, regulating ADP-G-actin and ADP-F-actin with high nucleotide homogeneity.

Expression of Fascin-1 and its diagnostic value in liver cancer.

Although some studies have reported on the expression and clinical significance of Fascin-1 (FSCN1) in liver cancer, the clinical application and differential diagnosis value of FSCN1 in liver cancer are still unclear. The aim of this study was to analyze the expression level of FSCN1 protein in liver cancer tissues and explore its diagnostic and application value in differentiating between hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC). The immunehistochemical analysis was used to detect the expression of FSCN1 in 108 cases of HCC, 26 cases of ICC, 23 cases of liver cirrhosis, and 11 cases of normal liver tissues. The differences in the positive expression rate and strong positive expression rate of FSCN1 among different groups were analyzed. The positive rate of FSCN1 in normal liver tissues, liver cirrhosis, HCC, and ICC tissues was 0.0% (0/11), 0.0% (0/23), 13.9% (15/108), and 92.3% (24/26), respectively, while the strong positive rate was 0.0% (0/11), 0.0% (0/23), 0.9% (1/108), and 69.2% (18/26), respectively. Both the positive rate and strong positive rate of FSCN1 in ICC tissues were significantly higher than those in HCC, liver cirrhosis, and normal liver tissues. Additionally, the positive rate of FSCN1 in moderately to poorly differentiated HCC tissues was 18.8% (15/80), significantly higher than in well-differentiated HCC (0.0%, 0/28) (P = 0.031). In liver cancer, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of FSCN1 positive prediction for ICC were 92.3%, 86.1%, 61.5%, and 97.9%, respectively, whereas the sensitivity, specificity, PPV, and NPV of FSCN1 strong positive prediction for ICC were 69.2%, 99.1%, 94.7%, and 93.0%, respectively. These results suggest that FSCN1 may play an important role in the occurrence and progression of liver cancer, and it can be used as a novel diagnostic marker for ICC.

The dysadherin/FAK axis promotes individual cell migration in colon cancer.

Dysregulation of cancer cell motility is a key driver of invasion and metastasis. High dysadherin expression in cancer cells is correlated with invasion and metastasis. Here, we found the molecular mechanism by which dysadherin regulates the migration and invasion of colon cancer (CC). Comprehensive analysis using single-cell RNA sequencing data from CC patients revealed that high dysadherin expression in cells is linked to cell migration-related gene signatures. We confirmed that the deletion of dysadherin in tumor cells hindered local invasion and distant migration using in vivo tumor models. In this context, by performing cell morphological analysis, we found that aberrant cell migration resulted from impaired actin dynamics, focal adhesion turnover and protrusive structure formation upon dysadherin expression. Mechanistically, the activation of focal adhesion kinase (FAK) was observed in dysadherin-enriched cells. The dysadherin/FAK axis enhanced cell migration and invasion by activating the FAK downstream cascade, which includes the Rho family of small GTPases. Overall, this study illuminates the role of dysadherin in modulating cancer cell migration by forcing actin dynamics and protrusive structure formation via FAK signaling, indicating that targeting dysadherin may be a potential therapeutic strategy for CC patients.

TAGLN2 targeted control of ARPC5-mediated activation of the MEK/ERK signaling pathway influences the proliferation, invasion, and metastasis of pancreatic cancer cells.

PURPOSE: Pcancreatic cancer (PC) is a common tumor of the digestive tract with an insidious onset and high malignancy potential. Currently, surgery is the only effective treatment modality. Therefore, it is crucial to discover new targeted therapeutic modalities. We studied whether transgelin 2 (TAGLN2) targeted control of actin-related protein 2/3 complex subunit 5 (ARPC5)-mediated activation of the MEK/ERK signaling pathway to Influences the proliferation, invasion, and metastasis of pancreatic cancer cells. METHODS: The effects of TAGLN2 overexpression and knockdown on the proliferative viability and invasive metastatic ability of pancreatic cancer cells were verified through in vitro and in vivo assays via constructing a stable lentiviral transfection of human pancreatic cancer cell lines PANC-1 and SW1990. Bioinformatics analysis was used to predict the relationship between TAGLN2 and ARPC5. These findings were subsequently verified through protein profiling, immunofluorescence (IF), and coimmunoprecipitation (CO-IP) assays. In vitro experiments were also conducted to confirm the effect of TAGLN2 modulation on ARPC5 expression, which subsequently affects the proliferation and invasive metastatic ability of pancreatic cancer cells. The study analyzed the relationship between TAGLN2 and the MEK/ERK signaling pathway through bioinformatics and in vitro experiments with the MEK signaling pathway inhibitor U0126. RESULTS: TAGLN2 is expressed at high levels in pancreatic cancer cell lines, and its expression is positively correlated with poor prognosis of pancreatic cancer. ARPC5 is a direct target of TAGLN2 and is associated with the MEK/ERK signaling pathway. In vivo and ex vivo experiments confirmed that overexpression of TAGLN2 promoted the proliferation, invasion, and metastasis of pancreatic cancer cells, and silencing ARPC5 reversed these effect. CONCLUSION: Our research revealed that TAGLN2 protein binds to ARPC5 protein and contributes to increased ARPC5 expression and activation of the MEK/ERK signaling pathway. This activation promotes pancreatic cancer cell growth, infiltration, and spread. Hence, TAGLN2 is a potential viable therapeutic target in pancreatic cancer and represents a novel therapeutic approach.

CARMIL1 regulates liver cancer cell proliferation by activating the ERK/mTOR pathway through the TRIM27/p53 axis.

Capping protein regulatory factor and myosin 1 linker 1 is termed CARMIL1. CARMIL1 is involved in several physiological processes; it forms an actin filament network and plasma membrane-bound cellular projection tissues and positively regulates the cellular components and tissues. CARMIL1 exhibits important biological functions in cancer; nonetheless, these functions have not been completely explored. We aimed to investigate the novel functions of CARMIL1 in liver cancer, particularly in cell proliferation. The cell counting kit-8, 5-ethynyl-2'-deoxyuridine, Component A experiments, and subcutaneous tumor formation model suggest that CARMIL1 is central to the proliferation of liver cancer cells both in vivo and in vitro. We extracted CARMIL1 samples from The Cancer Genome Atlas Program and analyzed its enrichment. CARMIL1 regulated the pathway activity by affecting the expression of star molecular proteins of the extracellular signal-regulated kinase (ERK) and mammalian target of rapamycin (mTOR). Moreover, it influenced the proliferation ability of liver cancer cells. Western blotting suggested that CARMIL1 downregulation could affect ERK and mTOR phosphorylation. Results of the co-immunoprecipitation demonstrated that CARMIL1 binds to tripartite motif (TRIM)27, which in turn binds to p53. Subsequently, CARMIL1 can regulate p53 stability and promote its degradation through TRIM27. Additionally, CARMIL1 inhibition enhanced the sensitivity of liver cancer cells to sorafenib. Tumor growth was significantly inhibited in the group treated with sorafenib and CARMIL1, compared with the group treated with CARMIL1 alone. Sorafenib is a first-line targeted chemotherapeutic drug for hepatocellular carcinoma treatment. It increases the long-term survival of hepatocellular carcinoma by 44%. In this study, downregulated CARMIL1 combined with sorafenib significantly reduced the tumor volume and weight of the mouse subcutaneous tumor model, indicating the potential possibility of combining CARMIL1 with sorafenib in hepatocellular carcinoma treatment. In summary, CARMIL1 promotes liver cancer cell proliferation by regulating the TRIM27/p53 axis and activating the ERK/mTOR pathway.

[The effect of the AIM2 inflammasome in noise-induced cognitive dysfunction in rats].

Objective: To explore the effect of the absent in melanoma 2 (AIM2) -mediated neuroinflammation in noise-induced cognitive dysfunction in rats. Methods: In April 2023, sixteen male Wistar rats were randomly divided into control group and noise group, with 8 rats in each group. The rats in the noise group were placed in 50 cm×50 cm×40 cm transparent boxes and exposed to 100 dB (A) white noise with a sound pressure level of 100 dB (A) (4 h/d for 30 d) . At the same time, rats in the control group were kept in similar boxes with environmental noise less than 60 dB (A) . After 30 days of noise exposure, the Morris water maze experiment was applied to test the learning and memory abilities of the rats; the pathological morphology of hippocampal tissues was observed by Hematoxylin-Eosin (HE) staining. Western blot was used to detect the protein expression levels of AIM2, cysteinyl aspartate specific proteinase-1 (caspase-1) , apoptosis-associated speck-like protein (ASC) , interleukin-1ß (IL-1ß) , IL-18, ionic calcium-binding articulation molecule-1 (Iba-1) , and glial fibrillary acidic protein (GFAP) . The expression of both Iba-1 and GFAP in hippocampal tissue was assessed by immunohistochemical staining. The co-localization of AIM2 with Iba-1 or GFAP was determined by immunofluorescence double staining. Results: Compared with the control group, the escape latency of rats in the noise group was increased by 16.29 s, 17.71 s, and 20.26 s on days 3, 4, and 5, respectively. On day 6, the noise-exposed rats spent shorter time in the target quadrant and had fewer times in crossing the platform[ (7.25±2.27) s and (1.13±0.64) times] than the control group[ (15.64±3.99) s and (4.25±2.12) times] (P<0.05) . After noise exposure, hippocampal neurons of rats displayed marked nuclear hyperchromatic and pyknosis phenomenon. The noise-exposed rats had higher numbers of both microglia and astrocytes (27.00±2.65 and 43.33±5.51) in the DG area of the hippocampus relative to the control group (14.67±3.06 and 20.00±4.58) (P<0.05) . Moreover, the glial cells in the noise group had larger cell cytosol with more and thicker branches. The protein expression levels of inflammatory cytokines Cleaved-IL-1ß and Cleaved-IL-18 in the hippocampus of rats in the noise group (1.55±0.19 and 1.74±0.12) were significantly higher than the control group (1.00±0.11 and 1.00±0.13) (P<0.05) . After noise exposure, the protein expression levels of AIM2, Cleaved-Caspase-1 and ASC (1.19±0.09, 1.34±0.07 and 1.14±0.01) were higher than the control group (1.00±0.07, 1.00±0.14 and 1.00±0.06) and differences between the two groups were statistically significant (P<0.05) . A significant increase in the number of cells co-localizing AIM2 with Iba-1 or GFAP in the noise group (28.67±4.04 and 40.67±5.13) compared with the control group (15.67±4.04 and 17.67±3.79) , and statistically significant differences were observed between the two groups (P<0.05) . Conclusion: Noise exposure may activate the AIM2 inflammasome in hippocampal glial cells of rats, releasing excessive inflammatory cytokines and causing neuroinflammation that damages neurons.

Monastrol suppresses invasion and metastasis in human colorectal cancer cells by targeting fascin independent of kinesin-Eg5 pathway.

Rearrangement of the actin cytoskeleton is a prerequisite for carcinoma cells to develop cellular protrusions, which are required for migration, invasion, and metastasis. Fascin is a key protein involved in actin bundling and is expressed in aggressive and invasive carcinomas. Additionally, fascin appears to be involved in tubulin-binding and microtubule rearrangement. Pharmacophoric-based in silico screening was performed to identify compounds with better fascin inhibitory properties than migrastatin, a gold-standard fascin inhibitor. We hypothesized that monastrol displays anti-migratory and anti-invasive properties via fascin blocking in colorectal cancer cell lines. Biophysical (thermofluor and ligand titration followed by fluorescence spectroscopy), biochemical (NMR), and cellular assays (MTT, invasion of human tissue), as well as animal model studies (zebrafish invasion) were performed to characterize the inhibitory effect of monastrol on fascin activity. In silico analysis revealed that monastrol is a potential fascin-binding compound. Biophysical and biochemical assays demonstrated that monastrol binds to fascin and interferes with its actin-bundling activity. Cell culture studies, including a 3D human myoma disc model, showed that monastrol inhibited fascin-driven cytoplasmic protrusions as well as invasion. In silico, confocal microscopy, and immunoprecipitation assays demonstrated that monastrol disrupted fascin-tubulin interactions. These anti-invasive effects were confirmed in vivo. In silico confocal microscopy and immunoprecipitation assays were carried out to test whether monastrol disrupted the fascin-tubulin interaction. This study reports, for the first time, the in vitro and in vivo anti-invasive properties of monastrol in colorectal tumor cells. The number and types of interactions suggest potential binding of monastrol across actin and tubulin sites on fascin, which could be valuable for the development of antitumor therapies.

A Coronin 1-Derived Peptide Inhibits Membrane Fusion by Modulating Membrane Organization and Dynamics.

Membrane fusion is considered the first step in the entry of enveloped viruses into the host cell. Several targeted strategies have been implemented to block viral entry by limiting the fusion protein to form a six-helix bundle, which is a prerequisite for fusion. Nonetheless, the development of broad-spectrum fusion inhibitors is essential to combat emerging and re-emerging viral infections. TG-23, a coronin 1, a tryptophan-aspartate-rich phagosomal protein-derived peptide, demonstrated inhibition of fusion between small unilamellar vesicles (SUVs) by modulating the membrane's physical properties. However, its inhibitory efficacy reduces with an increasing concentration of membrane cholesterol. The present work aims to develop a fusion inhibitor whose efficacy would be unaltered in the presence of membrane cholesterol. A stretch of the tryptophan-aspartic acid-containing peptide with a similar secondary structure and hydrophobicity profile of TG-23 from coronin 1 was synthesized, and its ability to inhibit SUV-SUV fusion with varying concentrations of membrane cholesterol was evaluated. Our results demonstrate that the GG-21 peptide inhibits fusion irrespective of the cholesterol content of the membrane. We have further evaluated the peptide-induced change in the membrane organization and dynamics utilizing arrays of steady-state and time-resolved fluorescence measurements and correlated these results with their effect on fusion. Interestingly, GG-21 displays inhibitory efficacy in a wide variety of lipid compositions despite having a secondary structure and physical properties similar to those of TG-23. Overall, our results advocate that the secondary structure and physical properties of the peptide may not be sufficient to predict its inhibitory efficacy.

Mesenchymal stem cell-derived exosomes improve neurogenesis and cognitive function of mice with methamphetamine addiction: A novel treatment approach.

BACKGROUND: Methamphetamine (METH) is a psychostimulant substance with highly addictive and neurotoxic effects, but no ideal treatment option exists to improve METH-induced neurocognitive deficits. Recently, mesenchymal stem cells (MSCs)-derived exosomes have raised many hopes for treating neurodegenerative sequela of brain disorders. This study aimed to determine the therapeutic potential of MSCs-derived exosomes on cognitive function and neurogenesis of METH-addicted rodents. METHODS: Male BALB/c mice were subjected to chronic METH addiction, followed by intravenous administration of bone marrow MSCs-derived exosomes. Then, the spatial memory and recognition memory of animals were assessed by the Barnes maze and the novel object recognition test (NORT). The neurogenesis-related factors, including NeuN and DCX, and the expression of Iba-1, a microglial activation marker, were assessed in the hippocampus by immunofluorescence staining. Also, the expression of inflammatory cytokines, including TNF-α and NF-κB, were evaluated by western blotting. RESULTS: The results showed that BMSCs-exosomes improved the time spent in the target quadrant and correct-to-wrong relative time in the Barnes maze. Also, NORT's discrimination index (DI) and recognition index (RI) were improved following exosome therapy. Additionally, exosome therapy significantly increased the expression of NeuN and DCX in the hippocampus while decreasing the expression of inflammatory cytokines, including TNF-α and NF-κB. Besides, BMSC-exosomes down-regulated the expression of Iba-1. CONCLUSION: Our findings indicate that BMSC-exosomes mitigated METH-caused cognitive dysfunction by improving neurogenesis and inhibiting neuroinflammation in the hippocampus.

Vasorin promotes endothelial differentiation of glioma stem cells via stimulating the transcription of VEGFR2.

Gliomas are highly vascularized malignancies, but current anti-angiogenic treatments have not demonstrated practical improvements in patient survival. Studies have suggested that glioma-derived endothelial cell (GdEC) formed by glioma stem cell (GSC) differentiation may contribute to the failure of this treatment. However, the molecular mechanisms involved in GSC endothelial differentiation remain poorly understood. We previously reported that vasorin (VASN) is highly expressed in glioma and promotes angiogenesis. Here, we show that VASN expression positively correlates with GdEC signatures in glioma patients. VASN promotes the endothelial differentiation capacity of GSC in vitro and participates in the formation of GSC-derived vessels in vivo. Mechanistically, vascular endothelial growth factor receptor 2 (VEGFR2) is a critical factor that mediates the regulation of VASN on GSC endothelial differentiation. Separation of cell chromatin fractionation and chromatin immunoprecipitation-sequencing analysis show that VASN interacts with Notch1 and co-translocates into the cell nuclei, where VASN binds to the VEGFR2 gene promoter to stimulate its transcription during the progression of GSC differentiation into GdEC. Together, these findings elucidate the role and mechanisms of VASN in promoting the endothelial differentiation of GSC and suggest VASN as a potential target for anti-angiogenic therapy based on intervention in GdEC formation in gliomas.

The F-actin bundler SWAP-70 promotes tumor metastasis.

Dynamic rearrangements of the F-actin cytoskeleton are a hallmark of tumor metastasis. Thus, proteins that govern F-actin rearrangements are of major interest for understanding metastasis and potential therapies. We hypothesized that the unique F-actin binding and bundling protein SWAP-70 contributes importantly to metastasis. Orthotopic, ectopic, and short-term tail vein injection mouse breast and lung cancer models revealed a strong positive dependence of lung and bone metastasis on SWAP-70. Breast cancer cell growth, migration, adhesion, and invasion assays revealed SWAP-70's key role in these metastasis-related cell features and the requirement for SWAP-70 to bind F-actin. Biophysical experiments showed that tumor cell stiffness and deformability are negatively modulated by SWAP-70. Together, we present a hitherto undescribed, unique F-actin modulator as an important contributor to tumor metastasis.

Nuclear actin structure regulates chromatin accessibility.

Polymerized ß-actin may provide a structural basis for chromatin accessibility and actin transport into the nucleus can guide mesenchymal stem cell (MSC) differentiation. Using MSC, we show that using CK666 to inhibit Arp2/3 directed secondary actin branching results in decreased nuclear actin structure, and significantly alters chromatin access measured with ATACseq at 24 h. The ATAC-seq results due to CK666 are distinct from those caused by cytochalasin D (CytoD), which enhances nuclear actin structure. In addition, nuclear visualization shows Arp2/3 inhibition decreases pericentric H3K9me3 marks. CytoD, alternatively, induces redistribution of H3K27me3 marks centrally. Such alterations in chromatin landscape are consistent with differential gene expression associated with distinctive differentiation patterns. Further, knockdown of the non-enzymatic monomeric actin binding protein, Arp4, leads to extensive chromatin unpacking, but only a modest increase in transcription, indicating an active role for actin-Arp4 in transcription. These data indicate that dynamic actin remodeling can regulate chromatin interactions.

Mitotic spindle positioning protein (MISP) preferentially binds to aged F-actin.

Actin bundling proteins crosslink filaments into polarized structures that shape and support membrane protrusions including filopodia, microvilli, and stereocilia. In the case of epithelial microvilli, mitotic spindle positioning protein (MISP) is an actin bundler that localizes specifically to the basal rootlets, where the pointed ends of core bundle filaments converge. Previous studies established that MISP is prevented from binding more distal segments of the core bundle by competition with other actin-binding proteins. Yet whether MISP holds a preference for binding directly to rootlet actin remains an open question. By immunostaining native intestinal tissue sections, we found that microvillar rootlets are decorated with the severing protein, cofilin, suggesting high levels of ADP-actin in these structures. Using total internal reflection fluorescence microscopy assays, we also found that purified MISP exhibits a binding preference for ADP- versus ADP-Pi-actin-containing filaments. Consistent with this, assays with actively growing actin filaments revealed that MISP binds at or near their pointed ends. Moreover, although substrate attached MISP assembles filament bundles in parallel and antiparallel configurations, in solution MISP assembles parallel bundles consisting of multiple filaments exhibiting uniform polarity. These discoveries highlight nucleotide state sensing as a mechanism for sorting actin bundlers along filaments and driving their accumulation near filament ends. Such localized binding might drive parallel bundle formation and/or locally modulate bundle mechanical properties in microvilli and related protrusions.

Human umbilical cord blood mesenchymal stem cells mediate microglia activation and improve anxiety-like behavior in MIA-induced offspring of schizophrenic rats.

Current treatments for schizophrenia (SCZ) remain largely ineffective in one-third of patients. Recent studies using stem cell therapy show a close relationship between stem cell immunomodulatory function and neuroinflammation in SCZ. To better investigate the efficacy of stem cell therapy for SCZ, human umbilical cord blood mesenchymal stem cells (hUC-MSC) with powerful immunomodulatory effects were administered to rats via the tail vein (once a week for 5 consecutive weeks starting from the weaning period) using a maternal immune activation (MIA) rodent model. Open field, PPI, Western blotting, Q-PCR, and immunofluorescence were used to assess the biological effects of repeated tail vein injections of hUC-MSC in offspring rats following the MIA model of SCZ. The results indicated that offspring of MIA rats exhibited schizophrenia-like (SCZ-like) anxiety behavior, with observed microglial activation triggering neuroinflammation. Furthermore, levels of IBA1, HMGB1, and PSD95 were significantly up-regulated, while SYP was significantly down-regulated. It is suggested that hUCB-MSCs may act through HMGB1, Iba1, PSD95, and related pathway molecules to alleviate neuroinflammation and repair synaptic damage by regulating the activity state of microglia. Consequently, this could improve the abnormal behavior observed in MIA offspring rats.

Afadin-nectin forces its way to the front.

Force transmission at cell-cell junctions critically regulates embryogenesis, tissue homeostasis, and diseases including cancer. The cadherin-catenin linkage has been considered the keystone of junctional force transmission, but new findings challenge this paradigm, arguing instead that the nectin-afadin linkage plays the more important role in mature junctions in the intestinal epithelium.

Shank3 deficiency elicits autistic-like behaviors by activating p38α in hypothalamic AgRP neurons.

BACKGROUND: SH3 and multiple ankyrin repeat domains protein 3 (SHANK3) monogenic mutations or deficiency leads to excessive stereotypic behavior and impaired sociability, which frequently occur in autism cases. To date, the underlying mechanisms by which Shank3 mutation or deletion causes autism and the part of the brain in which Shank3 mutation leads to the autistic phenotypes are understudied. The hypothalamus is associated with stereotypic behavior and sociability. p38α, a mediator of inflammatory responses in the brain, has been postulated as a potential gene for certain cases of autism occurrence. However, it is unclear whether hypothalamus and p38α are involved in the development of autism caused by Shank3 mutations or deficiency. METHODS: Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and immunoblotting were used to assess alternated signaling pathways in the hypothalamus of Shank3 knockout (Shank3-/-) mice. Home-Cage real-time monitoring test was performed to record stereotypic behavior and three-chamber test was used to monitor the sociability of mice. Adeno-associated viruses 9 (AAV9) were used to express p38α in the arcuate nucleus (ARC) or agouti-related peptide (AgRP) neurons. D176A and F327S mutations expressed constitutively active p38α. T180A and Y182F mutations expressed inactive p38α. RESULTS: We found that Shank3 controls stereotypic behavior and sociability by regulating p38α activity in AgRP neurons. Phosphorylated p38 level in hypothalamus is significantly enhanced in Shank3-/- mice. Consistently, overexpression of p38α in ARC or AgRP neurons elicits excessive stereotypic behavior and impairs sociability in wild-type (WT) mice. Notably, activated p38α in AgRP neurons increases stereotypic behavior and impairs sociability. Conversely, inactivated p38α in AgRP neurons significantly ameliorates autistic behaviors of Shank3-/- mice. In contrast, activated p38α in pro-opiomelanocortin (POMC) neurons does not affect stereotypic behavior and sociability in mice. LIMITATIONS: We demonstrated that SHANK3 regulates the phosphorylated p38 level in the hypothalamus and inactivated p38α in AgRP neurons significantly ameliorates autistic behaviors of Shank3-/- mice. However, we did not clarify the biochemical mechanism of SHANK3 inhibiting p38α in AgRP neurons. CONCLUSIONS: These results demonstrate that the Shank3 deficiency caused autistic-like behaviors by activating p38α signaling in AgRP neurons, suggesting that p38α signaling in AgRP neurons is a potential therapeutic target for Shank3 mutant-related autism.

N6-methyladenosine methylation on FSCN1 mediated by METTL14/IGF2BP3 contributes to human papillomavirus type 16-infected cervical squamous cell carcinoma.

Human papillomavirus (HPV) infection has been reported to be associated with N6-methyladenosine (m6A) modification in cancers. However, the underlying mechanism by which m6A methylation participates in HPV-related cervical squamous cell carcinoma (CSCC) remains largely unclear. In this study, we observed that m6A regulators methyltransferase like protein (METTL14) and insulin like growth factor 2 mRNA binding protein 3 (IGF2BP3) were upregulated in HPV-positive CSCC tissues and cell lines, and their high expression predicted poor prognosis for HPV-infected CSCC patients. Cellular functional experiments verified that HPV16 oncogenes E6/E7 upregulated the expression of METTL14 and IGF2BP3 to promote cell proliferation and epithelial mesenchymal transition of CSCC cells. Next, we found that E6/E7 stabilized fascin actin-bundling protein 1 (FSCN1) mRNA and elevated FSCN1 expression in CSCC cells through upregulating METTL14/IGF2BP3-mediated m6A modification, and FSCN1 expression was also validated to be positively associated with worse outcomes of HPV-positive CSCC patients. Finally, HPV16-positive CSCC cell lines SiHa and CaSki were transfected with knockdown vector for E6/E7 or METTL14/IGF2BP3 and overexpressing vector for FSCN1, and functional verification experiments were performed through using MTT assay, flow cytometry, wound healing assay and tumour formation assay. Results indicated that knockdown of E6/E7 or METTL14/IGF2BP3 suppressed cell proliferation, migration and tumorigenesis, and accelerated cell apoptosis of HPV-positive CSCC cells. Their tumour-suppressive effects were abolished through overexpressing FSCN1. Overall, HPV E6/E7 advanced CSCC development through upregulating METTL14/IGF2BP3-mediated FSCN1 m6A modification.