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Aging is a major risk factor for common neurodegenerative diseases. Although multiple molecular, cellular, structural, and functional changes occur in the brain during aging, the involvement of caveolin-2 (Cav-2) in brain ageing remains unknown. We investigated Cav-2 expression in brains of aged mice and its effects on endothelial cells. The human umbilical vein endothelial cells (HUVECs) showed decreased THP-1 adhesion and infiltration when treated with Cav-2 siRNA compared to control siRNA. In contrast, Cav-2 overexpression increased THP-1 adhesion and infiltration in HUVECs. Increased expression of Cav-2 and iba-1 was observed in brains of old mice. Moreover, there were fewer iba-1-positive cells in the brains of aged Cav-2 knockout (KO) mice than of wild-type aged mice. The levels of several chemokines were higher in brains of aged wild-type mice than in young wild-type mice; moreover, chemokine levels were significantly lower in brains of young mice as well as aged Cav-2 KO mice than in their wild-type counterparts. Expression of PECAM1 and VE-cadherin proteins increased in brains of old wild-type mice but was barely detected in brains of young wild-type and Cav-2 KO mice. Collectively, our results suggest that Cav-2 expression increases in the endothelial cells of aged brain, and promotes leukocyte infiltration and age-associated neuroinflammation.

Objectives Caveolin family proteins, including caveolin-1 (Cav-1), caveolin-2 (Cav-2), and caveolin-3 (Cav-3), are identified as the principal protein components of caveolae in mammalian cells. Circulating form of caveolin family proteins can be used as a good potential biomarker for predicting disease. Methods To investigate the clinical significance of the serological levels of caveolin family proteins in patients with systemic lupus erythematosus (SLE), we evaluated the soluble serum levels of caveolin family proteins in patients with SLE by enzyme-linked immunosorbent assay (ELISA) and assessed their associations with various known clinical variables. Results The major findings of our study are as follows: Cav-2 was not detected in serum of SLE patients and normal controls (NCs). Serum Cav-1 and Cav-3 levels were higher in SLE patients compared with NCs. There were no significant correlations between serum Cav-1 and Cav-3 levels and SLE disease activity. Further analysis showed that serum Cav-3 may be more valuable as a marker than serum Cav-1 in SLE patients. Conclusion Serum levels of Cav-1 and Cav-3 might have a diagnostic role in patients with SLE. However, their predictive and prognostic value was not determined. Further studies are necessary to determine the potential clinical significance of these assays in SLE.

Background Pancreatic cancer is the fourth leading cause of cancer deaths in the United States both in females and in males, and is projected to become the second deadliest cancer by 2030. The overall 5-year survival rate remains at around 10%. Cancer metabolism and specifically lipid metabolism plays an important role in pancreatic cancer progression and metastasis. Lipid droplets can not only store and transfer lipids, but also act as molecular messengers, and signaling factors. As lipid droplets are implicated in reprogramming tumor cell metabolism and in invasion and migration of pancreatic cancer cells, we aimed to identify lipid droplet-associated genes as prognostic markers in pancreatic cancer. Methods We performed a literature search on review articles related to lipid droplet-associated proteins. To select relevant lipid droplet-associated factors, bioinformatics analysis on the GEPIA platform (data are publicly available) was carried out for selected genes to identify differential expression in pancreatic cancer versus healthy pancreatic tissues. Differentially expressed genes were further analyzed regarding overall survival of pancreatic cancer patients. Results 65 factors were identified as lipid droplet-associated factors. Bioinformatics analysis of 179 pancreatic cancer samples and 171 normal pancreatic tissue samples on the GEPIA platform identified 39 deferentially expressed genes in pancreatic cancer with 36 up-regulated genes (ACSL3, ACSL4, AGPAT2, BSCL2, CAV1, CAV2, CAVIN1, CES1, CIDEC, DGAT1, DGAT2, FAF2, G0S2, HILPDA, HSD17B11, ICE2, LDAH, LIPE, LPCAT1, LPCAT2, LPIN1, MGLL, NAPA, NCEH1, PCYT1A, PLIN2, PLIN3, RAB5A, RAB7A, RAB8A, RAB18, SNAP23, SQLE, VAPA, VCP, VMP1) and 3 down-regulated genes (FITM1, PLIN4, PLIN5). Among 39 differentially expressed factors, seven up-regulated genes (CAV2, CIDEC, HILPDA, HSD17B11, NCEH1, RAB5A, and SQLE) and two down-regulation genes (BSCL2 and FITM1) were significantly associated with overall survival of pancreatic cancer patients. Multivariate Cox regression analysis identified CAV2 as the only independent prognostic factor. Conclusions Through bioinformatics analysis, we identified nine prognostic relevant differentially expressed genes highlighting the role of lipid droplet-associated factors in pancreatic cancer.

Objective The emphasis of this study focused on the possible implication and the mechanism of miR-144−3p in septic acute lung injury (ALI) condition. Methods Mice were pre-injected with miR-144−3p agomir, miR-144−3p antagomir, sh-Caveolin-2 or PBS before 10 mg/kg LPS induced sepsis model establishment. The ratio of wet weight of lung tissues and body weight (W/W) was calculated. The pathological changes on lung tissues were observed by H&E staining. Secretions of inflammatory cytokines (TNF-α, IL-1β and IL-6) in both mouse serum and lung tissues were determined by ELISA. Cell apoptosis and cell morphology were measured by TUNEL staining and H&E staining. The expressions of miR-144−3p, Caveolin-2, apoptotic related proteins and JAK/STAT pathway related proteins were measured by qRT-PCR or/and Western blot. Dual luciferase reporter assay was applied to detect the binding of miR-144−3p with Caveolin-2. Results LPS resulted in increased W/W, disrupted lung tissue, enhanced inflammatory response and cell apoptosis. miR-144−3p was upregulated while Caveolin-2 was downregulated in response to LPS treatment. Inflammation and cell apoptosis induced by LPS can be alleviated by miR-144−3p antagomir injection, but enhanced by miR-144−3p agomir or sh-Caveolin-2 treatment. miR-144−3p can negatively target Caveolin-2. miR-144−3p can activate the JAK/STAT signal pathway through Caveolin-2 in septic ALI mouse. Conclusion miR-144−3 can promote LPS induced septic ALI through downregulating Caveolin-2 to activate the JAK/STAT signal pathway.

We aimed to analyze the expression of caveolin-2 (CAV2) in patients with oral cancer and its correlations with clinicopathological parameters.The expression of CAV2 in oral cancer and its influence on the survival curves of oral cancer patients were inquired through the Human Protein Atlas Database. The cancer tissue specimens and normal paracancerous tissue specimens (?2 cm away from cancer tissues) were collected from 173 patients with oral cancer confirmed by pathology. Moreover, real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry were performed to detect the messenger ribonucleic acid (mRNA) and protein expressions of CAV2 in oral cancer tissues and corresponding paracancerous tissues, respectively, and their associations with the clinicopathological characteristics and survival conditions of oral cancer patients were analyzed.It was shown in the Human Protein Atlas Database that the expression of CAV2 was increased significantly in oral cancer tissues compared with that in normal tissues (P

Here, we show that Caveolin-2 (Cav-2) is an epigenetic regulator for adipogenesis. Upon adipogenic stimulation, inner nuclear membrane (INM)-targeted pY19-Cav-2 interacted with lamin A/C to disengage the repressed Cebpb promoter from lamin A/C, which facilitated the Cebpb promoter association with lamin B1. Consequently, pY19-Cav-2 recruited lysine demethylase 4b (KDM4b) for demethylation of histone H3 lysine 9 trimethylation (H3K9me3) and histone acetyltransferase GCN5 for acetylation of H3K27, and subsequently RNA polymerase II (Pol II) on Cebpb promoter for epigenetic activation of Cebpb, to initiate adipogenesis. Cav-2 knock-down abrogated the Cebpb activation and blocked the Pparg2 and Cebpa activation. Re-expression of Cav-2 restored Cebpb activation and adipogenesis in Cav-2-deficient preadipocytes. Our data identify a new mechanism by which the epigenetic activation of Cebpb is controlled at the nuclear periphery to promote adipogenesis.

N-myristoylation is a ubiquitous protein lipidation in eukaryotes, but regulatory roles for myristoylation on proteins still remain to be explored. Here, we show that N-myristoylation of Caveolin-2 (Cav-2) controls insulin signaling. Alternative translation initiation (ATI)-yielded truncated form of non-N-myristoylable Cav-2β and various conditional Cav-2 mutants were compared to full-length form of N-myristoylable Cav-2α. Insulin induced insulin receptor (IR) tyrosine kinase-catalyzed Tyr-19 phosphorylation of N-myristoylable M14A Cav-2 and triggered activation of IR signaling cascade. In contrast, insulin induced ubiquitination of non-N-myristoylable M1A and G2A Cav-2 to facilitate protein-tyrosine phosphatase 1B interaction with IR which desensitized IR signaling through internalization. Metabolic labeling and click chemistry showed palmitoylation of M14A but not M1A and G2A Cav-2. Insulin did not induce phosphorylation of M1A and G2A Cav-2 and Cav-2β. Like Cav-2α, G2A Cav-2 and Cav-2β formed large homo-oligomers localized in lipid rafts. These findings show Cav-2 N-myristoylation plays a crucial role to coordinate its phosphorylation, palmitoylation, and ubiquitination to control insulin signaling.

Background The CAV1-CAV2 locus has been associated with primary open-angle glaucoma (POAG) and intraocular pressure. However, its association with normal-tension glaucoma (NTG) was inconclusive. Therefore, we evaluated this association in Chinese and Japanese. Methods Two single-nucleotide polymorphisms (SNPs, rs4236601 and rs1052990) from previous genome-wide association studies of POAG were genotyped in a total of 2220 study subjects: a Hong Kong Chinese cohort of 537 NTG patients and 490 controls, a Shantou Chinese cohort of 102 NTG and 731 controls and an Osaka Japanese cohort of 153 NTG and 207 controls. Subgroup analysis by gender was conducted. Outcomes from different cohorts were combined using meta-analysis. Results SNP rs4236601 was significantly associated with NTG in the two Chinese cohorts (Pmeta = .0019, OR = 4.55, I2 = 0). In contrast, rs4236601 was monomorphic in the Osaka cohort. The association of rs1052990 was insignificant in a meta-analysis combining Chinese and Japanese cohorts (Pmeta = .81, OR = 1.05; I2 = 64%), and the OR tended towards opposite directions between Chinese (OR = 1.26) and Japanese (OR = 0.69). Gender-specific effects of the SNPs were not statistically significant in the logistic regression or Breslow-day tests of ORs (P > .05), although rs4236601 was significant in males (P = .0068; OR = 10.30) but not in females (P = .14; OR = 2.65) in the meta-analysis of Chinese subjects. Conclusions In this study, we confirmed the association of rs4236601 at the CAV1-CAV2 locus with NTG in Chinese. SNP rs4236601 is monomorphic, and rs1052990 tends towards a different direction in the Japanese cohort. Further studies are warranted to verify the ethnic difference and gender-specific effects of this locus.

Immunosuppression is critical for tumor growth and metastasis as well as obstacle to effective immunotherapy. Here, we demonstrate that host deficiency in caveolin-2, a member of caveolin protein family, increases M1-polarized tumor-associated macrophage (TAM) and CD8 T cell infiltration into subcutaneously implanted murine lung carcinoma tumors. Importantly, increase in M1 TAM-specific markers and cytokines occurs prior to increased numbers of tumor-infiltrating CD8 T cells and tumor regression in caveolin-2 deficient mice, suggesting that an early increase in M1 TAMs is a novel mechanism, via which host deficiency in caveolin-2 inhibits tumor growth. Consistent with the latter, transfer and co-injection of caveolin-2 deficient bone marrow (origin of TAMs) suppresses tumor growth and increases numbers of M1-polarized TAMs in wild type mice. Collectively, our data suggest that lung cancer cells use caveolin-2 expressed in bone marrow-derived cell types including TAMs to promote tumor growth via suppressing the anti-tumor immune response and that caveolin-2 could be a potential target for cancer immunotherapy.

Objective CAV1 encodes caveolin-1, a major protein of plasma membrane microdomains called caveolae, involved in several signaling pathways. Caveolin-1 is also located at the adipocyte lipid droplet. Heterozygous pathogenic variants of CAV1 induce rare heterogeneous disorders including pulmonary arterial hypertension and neonatal progeroid syndrome. Only one patient was previously reported with a CAV1 homozygous pathogenic variant, associated with congenital generalized lipodystrophy (CGL3). We aimed to further delineate genetic transmission, clinical, metabolic, and cellular characteristics of CGL3. Design/Methods In a large consanguineous kindred referred for CGL, we performed next-generation sequencing, as well as clinical, imagery, and metabolic investigations. We studied skin fibroblasts from the index case and the previously reported patient with CGL3. Results Four patients, aged 8 months to 18 years, carried a new homozygous p.(His79Glnfs*3) CAV1 variant. They all displayed generalized lipodystrophy since infancy, insulin resistance, low HDL-cholesterol, and/or high triglycerides, but no pulmonary hypertension. Two patients also presented at the age of 15 and 18 years with dysphagia due to achalasia, and one patient had retinitis pigmentosa. Heterozygous parents and relatives (n = 9) were asymptomatic, without any metabolic abnormality. Patients’ fibroblasts showed a complete loss of caveolae and no protein expression of caveolin-1 and its caveolin-2 and cavin-1 partners. Patients’ fibroblasts also displayed insulin resistance, increased oxidative stress, and premature senescence. Conclusions The CAV1 null variant investigated herein leads to an autosomal recessive congenital lipodystrophy syndrome. Loss of caveolin-1 and/or caveolae induces specific manifestations including achalasia which requires specific management. Overlapping phenotypic traits between the different CAV1-related diseases require further studies.

Endophthalmitis is a sight-threatening infection and a serious consequence of complications during intraocular surgery or penetrating injury of which Pseudomonas aeruginosa is an important etiology. Extracellular vesicles (EVs) have evolved as a promising entity for developing diagnostic and therapeutic biomarkers due to their involvement in intracellular communication and pathogenesis of diseases. We aimed to characterise the protein cargo of extracellular vesicles, isolated from a murine (C57BL/6) model of P. aeruginosa endophthalmitis by LC-MS/MS at 24 h post infection (p.i). EVs were extracted by ultracentrifugation, characterized by Dynamic Light Scattering (DLS) and western blotting with tetraspannin markers, CD9 and CD81 and quantified by the ExoCet quantification kit. Multiplex ELISA was performed to estimate the levels of TNF-α, IL-6, IFN-γ and IL-1β. Proteomic analysis identified 2010 proteins (FDR ≤0.01) in EVs from infected mice eyes, of which 137 were differentially expressed (P-value ≤ 0.05). A total of 101 proteins were upregulated and 36 were downregulated. Additionally, 43 proteins were exclusive to infection set. KEGG and Gene Ontology revealed, Focal adhesion, Phagosome pathway, Complement cascade and IL-17 signalling pathway are crucial upregulated pathways involving proteins such as Tenascin, caveolin 1, caveolin 2, glutamine synthetase, microtubule-associated protein, C1, C8 and IL-17. Tenascin and caveolins are known to suppress anti-inflammatory cytokines further exacerbating the disease. The result of this study provides insight into the global extracellular vesicle proteome of P. aeruginosa endophthalmitis with their functional correlation and distinctive pattern of expression and tenascin, caveolin 1 and caveolin 2 are attractive biomarkers for P. aeruginosa endophthalmitis.

Ischemia/reperfusion (I/R)-induced rapid inflammation involving activation of leukocyte-endothelial adhesive interactions and leukocyte infiltration into tissues is a major contributor to postischemic tissue injury. However, the molecular mediators involved in this pathological process are not fully known. We have previously reported that caveolin-2 (Cav-2), a protein component of plasma membrane caveolae, regulated leukocyte infiltration in mouse lung carcinoma tumors. The goal of the current study was to examine if Cav-2 plays a role in I/R injury and associated acute leukocyte-mediated inflammation. Using a mouse small intestinal I/R model, we demonstrated that I/R downregulates Cav-2 protein levels in the small bowel. Further study using Cav-2-deficient mice revealed aggravated postischemic tissue injury determined by scoring of villi length in H&E-stained tissue sections, which correlated with increased numbers of MPO-positive tissue-infiltrating leukocytes determined by IHC staining. Intravital microscopic analysis of upstream events relative to leukocyte transmigration and tissue infiltration revealed that leukocyte-endothelial cell adhesive interactions in postcapillary venules, namely leukocyte rolling and adhesion were also enhanced in Cav-2-deficient mice. Mechanistically, Cav-2 deficiency increased plasminogen activator inhibitor-1 (PAI-1) protein levels in the intestinal tissue and a pharmacological inhibition of PAI-1 had overall greater inhibitory effect on both aggravated I/R tissue injury and enhanced leukocyte-endothelial interactions in postcapillary venules in Cav-2-deficient mice. In conclusion, our data suggest that Cav-2 protein alleviates tissue injury in response to I/R by dampening PAI-1 protein levels and thereby reducing leukocyte-endothelial adhesive interactions. NEW & NOTEWORTHY The role of caveolin-2 in regulating ischemia/reperfusion (I/R) tissue injury and the mechanisms underlying its effects are unknown. This study uses caveolin-2-deficient mouse and small intestinal I/R injury models to examine the role of caveolin-2 in the leukocyte-dependent reperfusion injury. We demonstrate for the first time that caveolin-2 plays a protective role from the I/R-induced leukocyte-dependent reperfusion injury by reducing PAI-1 protein levels in intestinal tissue and leukocyte-endothelial adhesive interactions in postcapillary venules.

Aim This study aimed to identify the functional genes and genetic variants associated with the prognosis of pancreatic ductal adenocarcinoma (PDAC) and reveal the mechanism underlying their prognostic roles. Methods First, we implement a two-stage exome-wide association study in a total of 1070 patients to identify the genetic variant correlated with PDAC prognosis. Then we performed fine mapping through bioinformatics analysis and dual-luciferase reporter assays to reveal the causal functional variant and prognostic gene. Next, we established the gene knockdown, knockout, and overexpression cell lines with small interfering RNA, CRISPR/Cas9, and lentivirus, respectively, and investigated the gene function on cell proliferation and migration in vivo and in vitro. Finally, we performed the RNA-seq to elucidate downstream genes and mechanisms altering PDAC prognosis. Results We identified the CAV1-CAV2 locus tagged by rs8940 was significantly associated with PDAC prognosis, and rs10249656 in the 3′untranslated region of CAV2 was the real functional variant, which upregulated CAV2 expression through abolishing miR-548s binding. We observed upregulated CAV2 in PDAC and the higher expression correlated with worse prognosis. Transient knockdown of CAV2 inhibited PDAC migration without affecting proliferation rate. Knockout of CAV2 suppressed PDAC progression and metastasis, whereas stable overexpression of CAV2 promoted. Overexpressed CAV2 promoted the PDAC progression and metastasis via perturbing genes in the focal adhesion (CCND1, IGTA1, and ZYX) and extracellular matrix organisation (PLOD2, CAST, and ITGA1) pathways mechanically. Conclusion These findings shed light on an important role of CAV2 on PDAC progression and the prognostic impact of its genetic variation.

The axon initial segment (AIS) is a specialized neuronal compartment in which synaptic input is converted into action potential (AP) output. This process is supported by a diverse complement of sodium, potassium, and calcium channels (Ca(V)). Different classes of sodium and potassium channels are scaffolded at specific sites within the AIS, conferring unique functions, but how calcium channels are functionally distributed within the AIS is unclear. Here, we use conventional two-photon laser scanning and diffraction-limited, high-speed spot two-photon imaging to resolve AP-evoked calcium dynamics in the AIS with high spatiotemporal resolution. In mouse layer 5 prefrontal pyramidal neurons, calcium influx was mediated by a mix of Ca(V)2 and Ca(V)3 channels that differentially localized to discrete regions. Ca(V)3 functionally localized to produce nanodomain hotspots of calcium influx that coupled to ryanodine-sensitive stores, whereas Ca(V)2 localized to non-hotspot regions. Thus, different pools of Ca(V)s appear to play distinct roles in AIS function. SIGNIFICANCE STATEMENT The axon initial segment (AIS) is the site where synaptic input is transformed into action potential (AP) output. It achieves this function through a diverse complement of sodium, potassium, and calcium channels (Ca(V)). While the localization and function of sodium channels and potassium channels at the AIS is well described, less is known about the functional distribution of Ca(V)s. We used high-speed two-photon imaging to understand activity-dependent calcium dynamics in the AIS of mouse neocortical pyramidal neurons. Surprisingly, we found that calcium influx occurred in two distinct domains: Ca(V)3 generates hotspot regions of calcium influx coupled to calcium stores, whereas Ca(V)2 channels underlie diffuse calcium influx between hotspots. Therefore, different Ca(V) classes localize to distinct AIS subdomains, possibly regulating distinct cellular processes.

Malignant gliomas are the most common tumor in central nervous system with poor prognosis. Due to the limitation of histological classification in earlier diagnosis and individualized medicine, it is necessary to combine the molecular signatures and the pathological characteristics of gliomas. Lots of microRNAs presented abnormal expression in gliomas and modulated gliomas development. Exploration the miRNAs profile is helpful for the diagnosis, therapy and prognosis of gliomas. It has been demonstrated that miR-144 plays important roles in solid tumors. However, the detail mechanisms remained unrevealed. In this study, we have demonstrated the level of miR-144 decreased in glioma tissues from patients, especially in gliomas with higher grades. MiR-144 was also validated have lower expression in glioma cell lines compared with cortical neuron cell by using qRT-PCR. The in vitro functional experiment indicated miR-144 improved gliomas progression through repressing proliferation, sensitizing to chemotherapeutics and inhibiting metastasis. We further identified fibroblast growth factor 7 (FGF7) and Caveolin 2 (CAV2) were target genes of miR-144 by luciferase reporter assay and western blotting. The mechanisms study suggested forced FGF7 expression elevated Akt activation and decreased reactive oxygen species (ROS) generation. The MTT and cell cycle assay indicated miR-144 suppressed glioma cells proliferation through modulating FGF mediated Akt signaling pathway. Meanwhile, miR-144 promoted Temozolomide (TMZ) induced apoptosis in glioma cells via increasing ROS production by using FACS. On the other hand, CAV2, as another target of miR-144, accelerated glioma cells migration and invasion via promoting glioma cells EMT progress. Retrieved expression of FGF7 or CAV2 rescued the proliferation and migration function mediated by miR-144. Furthermore, the in vivo experiments in PDX models displayed the anti-tumor function of miR-144, which could be retrieved by overexpression of FGF7 and CAV2. Taken together, these findings indicated miR-144 acted as a potential target against gliomas progression and uncovered a novel regulatory mechanism, which may provide a new therapeutic strategy and prognostic indicator for gliomas.

The primary aim of this study was to determine the anti-neuropathic activity of (±)-18-methoxycoronaridine [(±)-18-MC] and (+)-catharanthine in mice by using the oxaliplatin-induced neuropathic pain paradigm and cold plate test. The results showed that both coronaridine congeners induce anti-neuropathic pain activity at a dose of 72 mg/kg (per os), whereas a lower dose (36 mg/kg) of (+)-catharanthine decreased the progress of oxaliplatin-induced neuropathic pain. To determine the underlying molecular mechanism, electrophysiological recordings were performed on α9α10, α3β4, and α4β2 nAChRs as well as voltage-gated calcium (Ca

Voltage gated calcium channels (VGCCs) regulate neuronal excitability and translate activity into calcium dependent signaling. The α1 subunit of high voltage activated (HVA) VGCCs associates with α2δ accessory subunits, which may affect calcium channel biophysical properties, cell surface expression, localization and transport and are thus important players in calcium-dependent signaling. In vertebrates, the functions of the different combinations of the four α2δ and the seven HVA α1 subunits are incompletely understood, in particular with respect to partially redundant or separate functions in neurons. This study capitalizes on the relatively simpler situation in the Drosophila genetic model containing two neuronal putative α2δ subunits, straightjacket and CG4587, and one Cav1 and Cav2 homolog each, both with well-described functions in different compartments of identified motoneurons. Straightjacket is required for normal Cav1 and Cav2 current amplitudes and correct Cav2 channel function in all neuronal compartments. By contrast, CG4587 does not affect Cav1 or Cav2 current amplitudes or presynaptic function, but is required for correct Cav2 channel allocation to the axonal versus the dendritic domain. We suggest that the two different putative α2δ subunits are required in the same neurons to regulate different functions of VGCCs.

The neuromodulatory effects of GABA on pyramidal neurons are mediated by GABABreceptors (GABABRs) that signal via a conserved G-protein-coupled pathway. Two prominent effectors regulated by GABABRs include G-protein inwardly rectifying K+(GIRK) and P/Q/N type voltage-gated Ca2+(CaV2) ion channels that control excitability and synaptic output of these neurons, respectively. Regulator of G-protein signaling 7 (RGS7) has been shown to control GABABeffects, yet the specificity of its impacts on effector channels and underlying molecular mechanisms is poorly understood. In this study, we show that hippocampal RGS7 forms two distinct complexes with alternative subunit configuration bound to either membrane protein R7BP (RGS7 binding protein) or orphan receptor GPR158. Quantitative biochemical experiments show that both complexes account for targeting nearly the entire pool of RGS7 to the plasma membrane. We analyzed the effect of genetic elimination in mice of both sexes and overexpression of various components of RGS7 complex by patch-clamp electrophysiology in cultured neurons and brain slices. We report that RGS7 prominently regulates GABABR signaling to CaV2, in addition to its known involvement in modulating GIRK. Strikingly, only complexes containing R7BP, but not GPR158, accelerated the kinetics of both GIRK and CaV2 modulation by GABABRs. In contrast, GPR158 overexpression exerted the opposite effect and inhibited RGS7-assisted temporal modulation of GIRK and CaV2 by GABA. Collectively, our data reveal mechanisms by which distinctly composed macromolecular complexes modulate the activity of key ion channels that mediate the inhibitory effects of GABA on hippocampal CA1 pyramidal neurons.SIGNIFICANCE STATEMENTThis study identifies the contributions of distinct macromolecular complexes containing a major G-protein regulator to controlling key ion channel function in hippocampal neurons with implications for understanding molecular mechanisms underlying synaptic plasticity, learning, and memory.

Toyoharu Kamibeppu, Koji Yamasaki, Kozue Nakahara, Takahiro Nagai, Naoki Terada, Hiromasa Tsukino, Shoichiro Mukai, Toshiyuki Kamoto Department of Urology, Faculty of Medicine, University of Miyazaki, Kiyotake, Japan Background: Caveolin (Cav)-1 and Cav-2 are cell membrane proteins, which are structural proteins of caveolae and are reported to be positive regulators of cell survival and metastasis in prostate cancer (PC). In a previous study, we reported that elevated levels of Cav-1 and Cav-2 were significantly associated with PC progression. However, their functions in PC have not yet been clarified. In this study, we examined the function of Cav-1 and Cav-2 in PC cell invasiveness and motility.Materials and methods: We introduced Cav-1- and Cav-2-specific small interfering into PC3 cells to knock-down (KD) both molecules. We also performed cell proliferation assay, wound healing assay, migration assay, and invasion assay using PC3 cells and compared the results between Cav-1-KD, Cav-2-KD, and negative control PC3 cells. In addition, we performed real-time quantitative PCR (RT-qPCR) and RT2 Profiler PCR Array analysis to identify factors influencing migration.Results: We observed no significant difference in the proliferative and invasive activities of Cav-1-KD and Cav-2-KD PC3 cells; however, the cell motility was significantly decreased compared with negative control PC3 cells. RT-qPCR revealed that the expression of vimentin and N-cadherin was downregulated in Cav-1-KD PC3 cells. In addition, PCR array revealed a decreased expression of MGAT5, MMP13, and MYCL in Cav-1-KD PC3 and ETV4, FGFR4, and SRC in Cav-2-KD PC3.Conclusion: Cav-1 and Cav-2 may positively contribute to the upregulation of castration-resistant PC cell migration. Cav-induced regulation of several molecules including vimentin, N-cadherin, MGAT5, MMP13, MYCL, ETV4, FGFR4, and SRC may have an important role in PC3 cell motility. However, further examination will be required. Keywords: caveolin-1, caveolin-2, CRPC, cell motility

Insulin resistance, defined as attenuated sensitivity responding to insulin, impairs insulin action. Direct causes and molecular mechanisms of insulin resistance have thus far remained elusive. Here we show that alternative translation initiation (ATI) of Caveolin-2 (Cav-2) regulates insulin sensitivity. Cav-2β isoform yielded by ATI desensitizes insulin receptor (IR) via dephosphorylation by protein-tyrosine phosphatase 1B (PTP1B), and subsequent endocytosis and lysosomal degradation of IR, causing insulin resistance. Blockage of Cav-2 ATI protects against insulin resistance by preventing Cav-2β-PTP1B-directed IR desensitization, thereby normalizing insulin sensitivity and glucose uptake. Our findings show that Cav-2β is a negative regulator of IR signaling, and identify a mechanism causing insulin resistance through control of insulin sensitivity via Cav-2 ATI.

This study aimed to investigate the effects of transient receptor potential vanilloid 4 (TRPV4) inhibition on blood-brain barrier (BBB) integrity and the expressions of caveolae structural proteins caveolin-1 and caveolin-2 in rats with focal cerebral ischemia and reperfusion. BBB permeability was assessed by Evans blue extravasation. The mRNA and protein expressions of caveolin-1 and caveolin-2 were determined by RT-PCR, Western blot and immunohistochemistry assays. We found that BBB permeability significantly increased and reaches its peak at 72 h of reperfusion in cerebral ischemia-reperfusion rats and is able to be ameliorated by administration of HC-067047, an antagonist of TRPV4. Additionally, it shows a significant upregulation of caveolin-1 and caveolin-2 expression in cerebral microvessels of ischemic tissue. However, treatment with HC-067047 was shown to downregulate caveolin-1 and caveolin-2 expression during cerebral ischemia-reperfusion. This study demonstrates that inhibition of TRPV4 ameliorates BBB leakage induced by ischemia-reperfusion injury through the downregulation of caveolin-1 and caveolin-2.

Large-scale genome-wide association studies have identified several susceptibility variants associated with the risk of primary open-angle glaucoma (POAG), among which rs4236601 (CAV1/CAV2) at chromosome 7q31 and rs2157719 at chromosome 9p21 (CDKN2B-AS1). The purpose of this study was to investigate whether these variants contribute to the incidence of POAG in a sample of the Brazilian Southeastern population and to determine the best-fitted genetic model for these single nucleotide polymorphisms (SNPs). A case-control study with 557 individuals, 310 with POAG, and 247 controls was conducted through PCR and direct sequencing. We observed a significant effect of the heterozygous genotype (G/A) of rs2157719 that occurred more frequently in the control group (p = 0.0004; OR: 0.517, CI 95%: 0.357-0.745). Allele frequencies also differed between cases and controls (p = 0.006; OR: 0.694, CI 95%: 0.522-0.922) with the best-fitted genetic model for rs2157719 being the codominant model. No differences were observed for genotype and allele distributions in relation to rs4236601 in the CAV1/CAV2 region. The association of rs2157719 (CDKN2B-AS1) with the POAG phenotype corroborates previously published results, reinforcing the importance of this variant in POAG etiology.

Most studies have revealed the effects of caveolins in cancer inhibition. However, due to a lack of reports about their new transcripts, their presence and their effects on different cancers are unclear. This study was conducted to evaluate the cavolin-2 (cav-2) transcripts expression changes in tumoral and corresponding tissues and in contralateral breast, to investigate their variation associated with the variation of caveolin-1 (cav-1) expression in breast cancer. There were 40 breast-derived tumoral, corresponding, and contralateral tissues obtained from the patients with breast cancer. The RNA and proteins were extracted from these samples. So, cav-1 and cav-2 transcripts' variation were assessed in whole tumoral, corresponding, and contralateral breast. Also, their expression modifications were evaluated via the Western blotting technique. The results derived from this study verified the presence of transcript III of cav-2 for the first time, which was reported only in the gene bank, but we could not detect and validate any protein associated with these transcripts. Also, the decreasing trend of cav-1 and the cav-2 (transcripts I and II) were observed in tumoral tissues compared to unaffected tissues especially in stages I and II. It seems that the descending expression levels of cav-1 and cav-2 (transcript I, II) besides the lasting expression of cav-2 (transcript III) are associated with the incidence and promotion of breast cancer, especially in the initial stages of breast cancer. So, this may show a potential in determining the patients who can undergo the prophylactic mastectomy. Moreover, the results of the study demonstrated that transcript III may be a candidate as a non-coding RNA.

Prostate cancer is a worldwide problem. While the role of caveolin-1 has been extensively studied, little is known about the role of caveolin-2 (CAV2) in prostate cancer. Up-regulation of CAV2 in androgen independent PC3 cells compared to normal prostate cell line and androgen dependent prostate cancer cell lines has been observed. Recent studies suggest that up-regulation of CAV2 plays an important role in androgen independent prostate cancer. This study investigates whether CAV2 is important in mediating the aggressive phenotypes seen in androgen independent prostate cancer cells. The androgen independent prostate cancer cell line, PC3 was used that has been shown to express CAV2, and CAV2 knock down was performed using siRNA system. Changes to cell number, migration and invasion were assessed after knocking down CAV2. Our results showed that down-regulating CAV2 resulted in reduced cell numbers, migration and invasion in PC3 cells. This preliminary study suggests that CAV2 may act to promote malignant behavior in an androgen independent prostate cancer cell line. Further studies are required to fully elucidate the role of CAV2 in androgen independent prostate cancer.

We recently demonstrated that the heart of late pregnant (LP) rodents is more prone to ischemia/reperfusion (I/R) injury compared to non-pregnant rodents. Lipids, particularly polyunsaturated fatty acids, have received special attention in the field of cardiovascular research. Here, we explored whether Intralipid (ITLD) protects the heart against I/R injury in LP rodents and investigated the mechanisms underlying this protection.In-vivo female LP rat hearts or ex-vivo isolated Langendorff-perfused LP mouse hearts were subjected to ischemia followed by reperfusion with PBS or ITLD (one bolus of 5mg/kg of 20% in in-vivo and 1% in ex-vivo). Myocardial infarct size, mitochondrial calcium retention capacity, genome-wide expression profiling, pharmacological inhibition and co-immunoprecipitation were performed. One bolus of ITLD at reperfusion significantly reduced the in-vivo myocardial infarct size in LP rats (23.3±2% vs. 55.5±3.4% in CTRL, p0.01). Postischemic administration of ITLD also protected the LP hearts against I/R injury ex-vivo. ITLD significantly increased the threshold for the opening of the mitochondrial permeability transition pore in response to calcium overload (nmol-calcium/mg-mitochondrial protein: 290±17 vs. 167±10 in CTRL, p0.01) and significantly increased phosphorylation of STAT3 (1.8±0.08 vs. 1±0.16 in CTRL, p0.05) and GSK-3β (2.63±0.55 vs. 1±0.0.34 in CTRL, p0.05). The ITLD-induced cardioprotection was fully abolished by Stattic, a specific inhibitor of STAT3. Transcriptome analysis revealed caveolin 2 (Cav2) was significantly upregulated by ITLD in hearts of LP rats under I/R injury. Co-immunoprecipitation experiments showed that Cav2 interacts with STAT3.ITLD protects the heart in late pregnancy against I/R injury by inhibiting the mPTP opening through Cav2/STAT3/GSK-3β pathway.

Ad libitum-fed diets high in fat and carbohydrate (especially fructose) induce weight gain, obesity, and nonalcoholic fatty liver disease (NAFLD) in humans and animal models. However, interpretation is complicated since ad libitum feeding of such diets induces hyperphagia and upregulates expression of liver fatty acid binding protein (L-FABP)-a protein intimately involved in fatty acid and glucose regulation of lipid metabolism. Wild-type (WT) and L-fabp gene ablated (LKO) mice were pair-fed either high-fat diet (HFD) or high-fat/high-glucose diet (HFGD) wherein total carbohydrate was maintained constant but the proportion of glucose was increased at the expense of fructose. In LKO mice, the pair-fed HFD increased body weight and lean tissue mass (LTM) but had no effect on fat tissue mass (FTM) or hepatic fatty vacuolation as compared to pair-fed WT counterparts. These LKO mice exhibited upregulation of hepatic proteins in fatty acid uptake and cytosolic transport (caveolin and sterol carrier protein-2), but lower hepatic fatty acid oxidation (decreased serum β-hydroxybutyrate). LKO mice pair-fed HFGD also exhibited increased body weight; however, these mice had increased FTM, not LTM, and increased hepatic fatty vacuolation as compared to pair-fed WT counterparts. These LKO mice also exhibited upregulation of hepatic proteins in fatty acid uptake and cytosolic transport (caveolin and acyl-CoA binding protein, but not sterol carrier protein-2), but there was no change in hepatic fatty acid oxidation (serum β-hydroxybutyrate) as compared to pair-fed WT counterparts.

Caveolae consist in lipid raft domains composed of caveolin proteins, cholesterol, glycosphingolipids, and GPI-anchored proteins. Caveolin proteins present three different types, caveolin 1 (CAV-1), caveolin 2 (CAV-2) and caveolin 3 (CAV-3), with a very similar structure and amino acid composition. The native caveolin proteins oxidation mechanism was investigated for the first time, at a glassy carbon electrode, using cyclic, square wave and differential pulse voltammetry. The three native caveolin proteins oxidation mechanism presented only one tyrosine and tryptophan amino acid residues oxidation peak. Denatured caveolin proteins presented also the tyrosine, tryptophan and cysteine amino acid residues oxidation peaks. The reverse cholesterol transport is related to caveolae and caveolin proteins, and CAV-1 is directly connected to cholesterol transport. The influence of cholesterol on the three caveolin proteins electrochemical behaviour was evaluated. In the absence and in the presence of cholesterol, significant differences in the CAV-1 oxidation peak current were observed.

This study describes the interaction between Cav2 calcium channels and Rabconnectin-3, a di-subunit protein that is associated with synaptic vesicles. Immunostaining reveals that both Rabconnectin-3α (RB-3α) and Rabconnectin-3β (RB-3β) are colocalized in mouse hippocampal neurons. Co-immunoprecipitations from brain tissue is consistent with the formation of a protein complex between RB-3α and RB-3β and both Cav2.2 and the related Cav2.1 calcium channel. The coexpression of either RB-3α or RB-3β with Cav2.2 calcium channels in tsA-201 cells led to a reduction in Cav2.2 current density without any effects on the voltage-dependence of activation or inactivation. Coexpression of both Rabconnectin-3 subunits did not cause an additive effect on current densities. Finally, the presence of Rabconnectin-3 did not interfere with μ-opioid receptor mediated Gβγ modulation of Cav2.2 channels. Altogether, our findings show that Rabconnectin-3 has the propensity to regulate calcium entry mediated by Cav2.2 channels.

Acute kidney injury (AKI) is a heterogeneous clinical syndrome with diverse outcomes. The recovery from AKI has prognostic importance. Little research has been done in order to find biomarkers that can predict recovery from AKI. Cav-2 is one of the main constituents of caveolae and is expressed in kidney. This study analyzed the time course of Cav-2 urinary excretion and renal expression in rats treated with cisplatin. Male Wistar rats were injected with cisplatin (5 mg/kg b.w., i.p.), and the studies were performed after 2, 4 and 14 days. Cav-2 abundance was evaluated in urine, in renal homogenates and in apical membranes by Western blotting. Cav-2 in urine was increased only 14 days after treatment, in the recovery phase of cisplatin-induced AKI. These results show that Cav-2 in urine could be useful as a biomarker of renal recovery, but not as an early biomarker of cisplatin-induced AKI. Cav-2 expression in total renal homogenates was not modified with treatment, but a down-regulation of Cav-2 in apical membranes was observed in treated animals. We hypothesize that Cav-2 internalizes into renal cells from their apical membrane in response to cisplatin, and regulates in this manner different signaling proteins involved in the physiopathology of renal damage.

Listeria monocytogenes is a human bacterial pathogen that disseminates through host tissues using a process called cell-to-cell spread. This critical yet understudied virulence strategy resembles a vesicular form of intercellular trafficking that allows L. monocytogenes to move between host cells without escaping the cell. Interestingly, eukaryotic cells can also directly exchange cellular components via intercellular communication pathways (e.g. trans-endocytosis) using cell-cell adhesion, membrane trafficking, and membrane remodeling proteins. Therefore, we hypothesized that L. monocytogenes would hijack these types of host proteins during spread. Using a focused RNAi screen, we identified 22 host genes that are important for L. monocytogenes spread. We then found that caveolins (CAV1 and CAV2) and the membrane sculpting F-BAR protein PACSIN2 promote L. monocytogenes protrusion engulfment during spread, and that PACSIN2 specifically localized to protrusions. Overall, our study demonstrates that host intercellular communication pathways may be co-opted during bacterial spread and that specific trafficking and membrane remodeling proteins promote bacterial protrusion resolution.SummaryThe human bacterial pathogen Listeria monocytogenes disseminates through host tissues using a process called cell-to-cell spread. In this study, Sanderlin et al., discover that host proteins that normally regulate membrane trafficking and membrane remodeling in uninfected settings are also co-opted by Listeria to promote spread.

Endothelial dysfunction plays a central role in the pathophysiology of pulmonary arterial hypertension (PAH). MicroRNAs (miRNAs) are small single-strand and non-coding RNAs that negatively regulate gene function by binding to the 3′-untranslated region (3′-UTR) of specific mRNAs. microRNA-1 (miR-1) is upregulated in plasma from idiopathic PAH patients and in lungs from an experimental model of PAH. However, the role of miRNA-1 on endothelial dysfunction is unknown. The aim of this study was to analyze the effects of miR-1 on endothelial function in rat pulmonary arteries (PA). Endothelial function was studied in PA from PAH or healthy animals and mounted in a wire myograph. Some PA from control animals were transfected with miR-1 or scramble miR. Superoxide anion production by miR-1 was quantified by dihydroethidium (DHE) fluorescence in rat PA smooth muscle cells (PASMC). Bioinformatic analysis identified superoxide dismutase-1 (SOD1), connexin-43 (Cx43), caveolin 2 (CAV2) and Kruppel-like factor 4 (KLF4) as potential targets of miR-1. The expression of SOD1, Cx43, CAV2, and KLF4 was determined by qRT-PCR and western blot in PASMC. PA incubated with miR-1 presented decreased endothelium-dependent relaxation to acetylcholine. We also found an increase in the production of O2− and decreased expression of SOD1, Cx43, CAV2, and KLF4 in PASMC induced by miR-1, which may contribute to endothelial dysfunction. In conclusion, these data indicate that miR-1 induces endothelial dysfunction, suggesting a pathophysiological role in PAH.

Determining the mechanism of action (MOA) of novel or naturally occurring compounds mostly relies on assays tailored for individual target proteins. Here we explore an alternative approach based on pattern matching response profiles obtained using cultured neuronal networks. Conolidine and cannabidiol are plant-derivatives with known antinociceptive activity but unknown MOA. Application of conolidine/cannabidiol to cultured neuronal networks altered network firing in a highly reproducible manner and created similar impact on network properties suggesting engagement with a common biological target. We used principal component analysis (PCA) and multi-dimensional scaling (MDS) to compare network activity profiles of conolidine/cannabidiol to a series of well-studied compounds with known MOA. Network activity profiles evoked by conolidine and cannabidiol closely matched that of ω-conotoxin CVIE, a potent and selective Cav2.2 calcium channel blocker with proposed antinociceptive action suggesting that they too would block this channel. To verify this, Cav2.2 channels were heterologously expressed, recorded with whole-cell patch clamp and conolidine/cannabidiol was applied. Remarkably, conolidine and cannabidiol both inhibited Cav2.2, providing a glimpse into the MOA that could underlie their antinociceptive action. These data highlight the utility of cultured neuronal network-based workflows to efficiently identify MOA of drugs in a highly scalable assay.

Drug resistance is a major obstacle in the treatment of breast cancer. Surviving cells lead to tumor recurrence and metastasis, which remains the main cause of cancer-related mortality. Breast cancer is also highly heterogeneous, which hinders the identification of individual cells with the capacity to survive anticancer treatment. To address this, we performed extensive single-cell gene-expression profiling of the luminal-type breast cancer cell line MCF7 and its derivatives, including docetaxel-resistant cells. Upregulation of epithelial-to-mesenchymal transition and stemness-related genes and downregulation of cell-cycle–related genes, which were mainly regulated by LEF1, were observed in the drug-resistant cells. Interestingly, a small number of cells in the parental population exhibited a gene-expression profile similar to that of the drug-resistant cells, indicating that the untreated parental cells already contained a rare subpopulation of stem-like cells with an inherent predisposition toward docetaxel resistance. Our data suggest that during chemotherapy, this population may be positively selected, leading to treatment failure. Significance: This study highlights the role of breast cancer intratumor heterogeneity in drug resistance at a single-cell level.

Background: The activation of hepatic stellate cells (HSCs) participates in liver fibrosis, and emerging evidences indicate that microRNAs (miRNAs) are abnormally expressed during HSC activation. However, the potential roles of miRNAs in liver fibrosis still remain elusive. Methods: The differential expression of miRNAs between the quiescent and activated HSCs was measured by miRNA microarray analysis, which was further validated by RT-qPCR in animal models and patients. The effect of miR-199a-3p on CAV2 expression was examined by Western blotting using miRNA mimics or inhibitor. Induction of miR-199a-3p expression by TWIST1 was explored by the luciferase assay and RT-qPCR. The role of miR-199a-3p in liver fibrosis was systematically investigated in vitro and in vivo. Findings: miR-199a-3p expression was dramatically upregulated during HSC activation in vitro, and during liver fibrogenesis in CCl4-treated rats, and was increased in livers from high-fat-diet-induced mice or the patients with cirrhosis. The expression of miR-199a-3p was driven by Twist1 which could be further induced by TGF-β treatment. Functional studies showed that inhibition of miR-199a-3p in both human LX2 cells and rat HSCs significantly decreased the expression of fibrotic markers, whereas the forced expression of miR-199a-3p exhibited opposite effects, demonstrating the role of miR-199a-3p in promoting HSCs activation. Mechanistically, miR-199a-3p plays an important role in TGF-β signalling pathway activation through targeting CAV2 that negatively regulates the expression of transforming growth factor-beta receptor type I (TGFβRI). Importantly, administration of antagomiR-199a-3p in the CCl4 treated mice significantly ameliorated hepatic fibrosis. Interpretation: Twist1-induced miR-199a-3p mediates the activation of HSCs by suppressing CAV2 expression and subsequently increasing TGFβRI expression to promote TGF-β pathway. Our findings highlight the therapeutic potential of miR-199a-3p for hepatic fibrosis. Funding Statement: This study was supported by National Natural Science Foundation of China (81570554 and 81770568 to LY; 81572739 to YP), and by National Key R&D Program of China (2016YFA0502204 to YP). TS is supported by a joint PhD studentship by King’s College London, UK and China Scholarship Council, China. Declaration of Interests: None of the authors has any potential conflict of interests to disclose. Ethics Approval Statement: All animal studies were approved by the Medical Ethics Committee of the West China Hospital of Sichuan University. All specimens were collected upon obtaining informed consent from all patients, and the study was approved by the Human Ethics Committee of Sichuan University.

The accumulation of intracellular β-amyloid peptide (Aβ) is important pathological characteristic of Alzheimer’s disease (AD). However, the exact underlying molecular mechanism remains to be elucidated. Here, we reported that Nuclear Paraspeckle Assembly Transcript 1 (NEAT1), a long n on-coding RNA, exhibits repressed expression in the early stage of AD and its down-regulation declines neuroglial cell mediating Aβ clearance via inhibiting expression of endocytosis-related genes. We find that NEAT1 is associated with P300/CBP complex and its inhibition affects H3K27 acetylation (H3K27Ac) and H3K27 crotonylation (H3K27Cro) located nearby to the transcription start site of many genes, including endocytosis-related genes. Interestingly, NEAT1 inhibition down-regulates H3K27Ac but up-regulates H3K27Cro through repression of acetyl-CoA generation. NEAT1 also mediates the binding between STAT3 and H3K27Ac but not H3K27Cro. Therefore, the decrease of H3K27Ac and/or the increase of H3K27Cro declines expression of multiple related genes. Collectively, this study first reveals the different roles of H3K27Ac and H3K27Cro in regulation of gene expression and provides the insight of the epigenetic regulatory mechanism of NEAT1 in gene expression and AD pathology. Electronic supplementary material The online version of this article (10.1007/s00018-019-03074-9) contains supplementary material, which is available to authorized users.

Caveolins (Cav-1, -2 and -3) and Cavins (Cavin-1, -2, -3 and -4) are two protein families controlling the biogenesis and function of caveolae, plasma membrane omega-like invaginations representing the primary site of important cellular processes like endocytosis, cholesterol homeostasis and signal transduction. Caveolae are especially abundant in fat tissue, playing a consistent role in a number of processes, such as the insulin-dependent glucose uptake and transmembrane transport of lipids underlying differentiation, maintenance and adaptive hypertrophy of adipocytes. Based on this premise, in this work we have investigated the expression of caveolar protein components in liposarcoma (LPS), an adipocytic soft tissue sarcoma affecting adults categorized in well-differentiated, dedifferentiated, myxoid and pleomorphic histotypes. By performing an extensive microarray data analysis followed by immunohistochemistry on human LPS tumors, we demonstrated that Cav-1, Cav-2 and Cavin-1 always cluster in all the histotypes, reaching the highest expression in well-differentiated LPS, the least aggressive of the malignant forms composed by tumor cells with a morphology resembling mature adipocytes. In vitro experiments carried out using two human LPS cell lines showed that the expression levels of Cav-1, Cav-2 and Cavin-1 proteins were faintly detectable during cell growth, becoming consistently increased during the accumulation of intracellular lipid droplets characterizing the adipogenic differentiation. Moreover, in differentiated LPS cells the three proteins were also found to co-localize and form molecular aggregates at the plasma membrane, as shown via immunofluorescence and immunoprecipitation analysis. Overall, these data indicate that Cav-1, Cav-2 and Cavin-1 may be considered as reliable markers for identification of LPS tumors characterized by consistent adipogenic differentiation.

BK polyomavirus (BKPyV) is a human pathogen that causes polyomavirus-associated nephropathy and hemorrhagic cystitis in transplant patients. Gangliosides and caveolin proteins have previously been reported to be required for BKPyV infection in animal cell models. Recent studies from our lab and others, however, have indicated that the identity of the cells used for infection studies can greatly influence the behavior of the virus. We therefore wished to re-examine BKPyV entry in a physiologically relevant primary cell culture model, human renal proximal tubule epithelial cells. Using siRNA knockdowns, we interfered with expression of UDP-glucose ceramide glucosyltransferase (UGCG), and the endocytic vesicle coat proteins caveolin 1, caveolin 2, and clathrin heavy chain. The results demonstrate that while BKPyV does require gangliosides for efficient infection, it can enter its natural host cells via a caveolin- and clathrin-independent pathway. The results emphasize the importance of studying viruses in a relevant cell culture model.

Prefrontal cortical areas mediate flexible adaptive control of behavior, but the specific contributions of individual areas and the circuit mechanisms through which they interact to modulate learning have remained poorly understood. Using viral tracing and pharmacogenetic techniques, we show that prelimbic (PreL) and infralimbic cortex (IL) exhibit reciprocal PreL↔IL layer 5/6 connectivity. In set-shifting tasks and in fear/extinction learning, activity in PreL is required during new learning to apply previously learned associations, whereas activity in IL is required to learn associations alternative to previous ones. IL→PreL connectivity is specifically required during IL-dependent learning, whereas reciprocal PreL↔IL connectivity is required during a time window of 12–14 h after association learning, to set up the role of IL in subsequent learning. Our results define specific and opposing roles of PreL and IL to together flexibly support new learning, and provide circuit evidence that IL-mediated learning of alternative associations depends on direct reciprocal PreL↔IL connectivity., Prelimbic (PL) and infralimbic (IL) cortical areas are known to have complementary roles in learning and decision making. Here the authors report reciprocal connectivity between the two areas and elucidate their functional impact on different aspects of learning.

Caveolin-2 (Cav-2) is expressed in a variety of cell tissue, and it has also been found in renal tissue. The expression of Cav-2 in proximal tubules is still unclear. The aim of this study was to carry out a complete evaluation of the expression pattern of Cav-2 in rat renal cortex to clarify and deepen the knowledge about the localization of Cav-2 in the proximal tubules and also to evaluate its presence in urine. Male Wistar rats were used to assess Cav-2 expression by Western blot analysis in homogenates, apical, and basolateral membranes from kidney cortex, in lysates and total plasma membranes from renal cortical cell suspensions, in urine, and in urinary exosomes. Cav-2 was clearly expressed in renal cortex homogenates and in both apical and basolateral membranes isolated from kidney cortex, with a greater expression on the former membranes. It was also observed in lysates and in plasma membranes from cortical cell suspensions. Moreover, Cav-2 was found in urine and in its exosomal fraction. These results confirmed the presence of Cav-2 in proximal tubule cells in the kidney of healthy rats, and showed for the first time its expression at the apical membrane of these cells and in urine. Besides, urinary exosomal pathway could be involved in Cav-2 urinary excretion under normal conditions. We observed an increase in the urinary abundance of Cav-2 in two models of acute kidney injury, and thus we proposed the urinary excretion of Cav-2 as a potential biomarker of kidney injury.

Reducing insulin/IGF-1 signaling (IIS) extends lifespan, promotes protein homeostasis (proteostasis), and elevates stress resistance of worms, flies, and mammals. How these functions are orchestrated across the organism is only partially understood. Here, we report that in the nematode Caenorhabditis elegans, the IIS positively regulates the expression of caveolin-1 (cav-1), a gene which is primarily expressed in neurons of the adult worm and underlies the formation of caveolae, a subtype of lipid microdomains that serve as platforms for signaling complexes. Accordingly, IIS reduction lowers cav-1 expression and lessens the quantity of neuronal caveolae. Reduced cav-1 expression extends lifespan and mitigates toxic protein aggregation by modulating the expression of aging-regulating and signaling-promoting genes. Our findings define caveolae as aging-governing signaling centers and underscore the potential for cav-1 as a novel therapeutic target for the promotion of healthy aging.

The expression of caveolin-1 (CAV1) in both tumor cell and cancer-associated fibroblasts (CAFs) has been found to correlate with tumor aggressiveness in different epithelial tumor entities, whereas less is known for caveolin-2 (CAV2). The aim of this study was to investigate the clinicopathological significance and prognostic value of stromal CAV1 and CAV2 expression in lung cancer. The expression of these two genes was investigated at protein level on a tissue microarray (TMA) consisting of 161 primary tumor samples. 50.7% of squamous cell lung cancer (SCC) tumors showed strong expression of CAV1 in the tumor-associated stromal cells, whereas only 15.1% of adenocarcinomas (AC) showed a strong CAV1 expression (p<0.01). A strong CAV2 stromal expression was found in 46.0% of the lung tumor specimens, with no significant difference between the subtypes. Neither CAV1 nor CAV2 stromal expression was associated with any other clinicopathological factor including survival. When the stromal expression in matched primary tumors and lymph node metastases was compared, both CAV1 and CAV2 expressions were frequently found lost in the corresponding stroma of the lymph node metastasis (40.6%, p=0.003 and 38.4%, p=0.001, resp.). Loss of stromal CAV2 in the lymph node metastases was also significantly associated with earlier death (p=0.011). In conclusion, in contrast to the expression patterns in the tumor tissue of lung cancer, stromal expression of CAV1 in primary tumors was not associated with clinical outcome whereas the stromal expression of especially CAV2 in the metastatic lymph nodes could be associated with lung cancer pathogenesis.