Several chemical compounds including natural products have been suggested as being effective against age-related diseases or as beneficial for a healthy life. On the other hand, forkhead box O (FOXO) ...proteins are emerging as key cellular components associated with extreme human longevity. FOXO proteins are mainly regulated by posttranslational modifications and as these modifications are reversible, activation and inactivation of FOXO are attainable through pharmacological treatment. Here, we questioned whether a panel of compounds with known health-beneficial properties has the capacity to induce the activity of FOXO factors. We show that resveratrol, a phytoalexin present in grapes and other food products, the amide alkaloid piperlongumine found in the fruit of the long pepper, and the plant-derived β-carboline compound harmine induced nuclear translocation of FOXO3. We also show that piperlongumine and harmine but not resveratrol activate FOXO-dependent transcription. We determined the half maximal effective concentration (EC50) values for resveratrol, piperlongumine, and harmine for FOXO translocation, and analyzed their inhibitory impact on chromosomal maintenance 1 (CRM1)-mediated nuclear export and the production of reactive oxygen species (ROS). We also used chemical biology approach and Western blot analysis to explore the underlying molecular mechanisms. We show that harmine, piperlongumine, and resveratrol activate FOXO3 independently of phosphoinositide 3-kinase (PI3K)/AKT signaling and the CRM1-mediated nuclear export. The effect of harmine on FOXO3 activity is at least partially mediated through the inhibition of dual-specificity tyrosine (Y) phosphorylationregulated kinase 1A (DYRK1A) and can be reverted by the inhibition of sirtuins (SIRTs).
Psoriasis is a common inflammatory skin disease involving a cross‐talk between epidermal and immune cells. The role of specific epidermal stem cell populations, including hair follicle stem cells ...(HF‐SCs) in psoriasis is not well defined. Here, we show reduced expression of c‐JUN and JUNB in bulge HF‐SCs in patients with scalp psoriasis. Using lineage tracing in mouse models of skin inflammation with inducible deletion of c‐Jun and JunB, we found that mutant bulge HF‐SCs initiate epidermal hyperplasia and skin inflammation. Mechanistically, thymic stromal lymphopoietin (TSLP) was identified in mutant cells as a paracrine factor stimulating proliferation of neighboring non‐mutant epidermal cells, while mutant inter‐follicular epidermal (IFE) cells are lost over time. Blocking TSLP in psoriasis‐like mice reduced skin inflammation and decreased epidermal proliferation, VEGFα expression, and STAT5 activation. These findings unravel distinct roles of HF‐SCs and IFE cells in inflammatory skin disease and provide novel mechanistic insights into epidermal cell interactions in inflammation.
Synopsis
Lineage tracing in models of skin inflammation reveals that thymic stromal lymphopoietin (TSLP) activates pro‐inflammatory cues in mutant hair follicle stem cells (HF‐SCs) and keratinocytes (KCs) (1). Primed non‐mutant HF‐SCs also produce TSLP, thus contributing to the disease (2).
Scalp psoriasis was associated with a reduction of c‐JUN/JUNB in bulge hair follicle stem cells (HF‐SCs) in psoriatic patients.
Specific deletion of c‐Jun/JunB in bulge HF‐SCs (mutantGFP HF‐SCs) was sufficient for the development of psoriasis‐like disease in mice.
Keratinocytes derived from mutantGFPHF‐SCs survived, while keratinocytes from mutantGFP inter‐follicular epidermal cells (IFE) were lost during psoriasis progression.
TSLP was an important mediator for epidermal hyper‐proliferation and pro‐inflammatory signaling in keratinocytes. Neutralization with anti‐TSLP reduced epidermal cell proliferation and psoriasis‐like progression in mice.
TSLP was highly expressed in the epidermis and hair follicles of scalp psoriasis.
Lineage tracing in models of skin inflammation reveals that thymic stromal lymphopoietin (TSLP) activates pro‐inflammatory cues in mutant hair follicle stem cells (HF‐SCs) and keratinocytes (KCs). Primed non‐mutant HF‐SCs also produce TSLP, thus contributing to the disease.
Cutaneous melanoma is a type of cancer with an inherent potential for lymph node colonization, which is generally preceded by neolymphangiogenesis. However, sentinel lymph node removal does not ...necessarily extend the overall survival of patients with melanoma. Moreover, lymphatic vessels collapse and become dysfunctional as melanomas progress. Therefore, it is unclear whether (and how) lymphangiogenesis contributes to visceral metastasis. Soluble and vesicle-associated proteins secreted by tumours and/or their stroma have been proposed to condition pre-metastatic sites in patients with melanoma. Still, the identities and prognostic value of lymphangiogenic mediators remain unclear. Moreover, our understanding of lymphangiogenesis (in melanomas and other tumour types) is limited by the paucity of mouse models for live imaging of distal pre-metastatic niches. Injectable lymphatic tracers have been developed, but their limited diffusion precludes whole-body imaging at visceral sites. Vascular endothelial growth factor receptor 3 (VEGFR3) is an attractive 'lymphoreporter' because its expression is strongly downregulated in normal adult lymphatic endothelial cells, but is activated in pathological situations such as inflammation and cancer. Here, we exploit this inducibility of VEGFR3 to engineer mouse melanoma models for whole-body imaging of metastasis generated by human cells, clinical biopsies or endogenously deregulated oncogenic pathways. This strategy revealed early induction of distal pre-metastatic niches uncoupled from lymphangiogenesis at primary lesions. Analyses of the melanoma secretome and validation in clinical specimens showed that the heparin-binding factor midkine is a systemic inducer of neo-lymphangiogenesis that defines patient prognosis. This role of midkine was linked to a paracrine activation of the mTOR pathway in lymphatic endothelial cells. These data support the use of VEGFR3 reporter mice as a 'MetAlert' discovery platform for drivers and inhibitors of metastasis.
Alteration of centrosome function and dynamics results in major defects during chromosome segregation and is associated with primary autosomal microcephaly (MCPH). Despite the knowledge accumulated ...in the last few years, why some centrosomal defects specifically affect neural progenitors is not clear. We describe here that the centrosomal kinase PLK1 controls centrosome asymmetry and cell fate in neural progenitors during development. Gain- or loss-of-function mutations in Plk1, as well as deficiencies in the MCPH genes Cdk5rap2 (MCPH3) and Cep135 (MCPH8), lead to abnormal asymmetry in the centrosomes carrying the mother and daughter centriole in neural progenitors. However, whereas loss of MCPH proteins leads to increased centrosome asymmetry and microcephaly, deficient PLK1 activity results in reduced asymmetry and increased expansion of neural progenitors and cortical growth during mid-gestation. The combination of PLK1 and MCPH mutations results in increased microcephaly accompanied by more aggressive centrosomal and mitotic abnormalities. In addition to highlighting the delicate balance in the level and activity of centrosomal regulators, these data suggest that human PLK1, which maps to 16p12.1, may contribute to the neurodevelopmental defects associated with 16p11.2-p12.2 microdeletions and microduplications in children with developmental delay and dysmorphic features.
Mammalian DNA replication origins are “licensed” by the loading of DNA helicases, a reaction that is mediated by CDC6 and CDT1 proteins. After initiation of DNA synthesis, CDC6 and CDT1 are inhibited ...to prevent origin reactivation and DNA overreplication before cell division. CDC6 and CDT1 are highly expressed in many types of cancer cells, but the impact of their deregulated expression had not been investigated in vivo. Here, we have generated mice strains that allow the conditional overexpression of both proteins. Adult mice were unharmed by the individual overexpression of either CDC6 or CDT1, but their combined deregulation led to DNA re-replication in progenitor cells and lethal tissue dysplasias. This study offers mechanistic insights into the necessary cooperation between CDC6 and CDT1 for facilitation of origin reactivation and describes the physiological consequences of DNA overreplication.
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•Transgenic mouse strains allow Cdc6- and Cdt1-inducible overexpression•Cdc6 and Cdt1 induce origin reactivation when expressed in combination•DNA re-replication causes lethal dysplasias in intestinal epithelia and other tissues•Levels of endogenous Cdc6 influence the sensitivity to Cdt1 deregulation in vivo
Muñoz et al. investigate the effects of deregulated DNA replication in vivo. The combined overexpression of CDC6 and CDT1, two key proteins that activate replication origins, resulted in aberrant origin reactivation, DNA overreplication, and lethal dysplasia in the intestine and other tissues.
Inappropriate drug delivery, secondary toxicities, and persistent chemo- and immunoresistance have traditionally compromised treatment response in melanoma. Using cellular systems and genetically ...engineered mouse models, we show that melanoma cells retain an innate ability to recognize cytosolic double-stranded RNA (dsRNA) and mount persistent stress response programs able to block tumor growth, even in highly immunosuppressed backgrounds. The dsRNA mimic polyinosine-polycytidylic acid, coadministered with polyethyleneimine as carrier, was identified as an unanticipated inducer of autophagy downstream of an exacerbated endosomal maturation program. A concurrent activity of the dsRNA helicase MDA-5 driving the proapoptotic protein NOXA resulted in an efficient autodigestion of melanoma cells. These results reveal tractable links for therapeutic intervention among dsRNA helicases, endo/lysosomes, and apoptotic factors.
The circadian clock is synchronized to the 24 h day by environmental light which is transmitted from the retina to the suprachiasmatic nucleus (SCN) primarily via the retinohypothalamic tract (RHT). ...Circadian rhythm abnormalities have been reported in neurodegenerative disorders such as Alzheimer's disease (AD). Whether these AD-related changes are a result of the altered clock gene expression, retina degeneration, including the dysfunction in RHT transmission, loss of retinal ganglion cells and its electrophysiological capabilities, or a combination of all of these pathological mechanisms, is not known. Here, we evaluated transgenic APP/PS1 mouse model of AD and wild-type mice at 6- and 12-month-old, as early and late pathological stage, respectively. We noticed the alteration of circadian clock gene expression not only in the hypothalamus but also in two extra-hypothalamic brain regions, cerebral cortex and hippocampus, in APP/PS1 mice. These alterations were observed in 6-month-old transgenic mice and were exacerbated at 12 months of age. This could be explained by the reduced RHT projections in the SCN of APP/PS1 mice, correlating with downregulation of hypothalamic GABAergic response in APP/PS1 mice in advanced stage of pathology. Importantly, we also report retinal degeneration in APP/PS1 mice, including Aβ deposits and reduced choline acetyltransferase levels, loss of melanopsin retinal ganglion cells and functional integrity mainly of inner retina layers. Our findings support the theory that retinal degeneration constitutes an early pathological event that directly affects the control of circadian rhythm in AD.
Background
Alzheimer’s disease (AD) is a progressive, chronic, and neurodegenerative disease, and the most common cause of dementia worldwide. Currently, the mechanisms underlying the disease are far ...from being elucidated. Thus, the study of proteins involved in its pathogenesis would allow getting further insights into the disease and identifying new markers for AD diagnosis.
Methods
We aimed here to analyze protein dysregulation in AD brain by quantitative proteomics to identify novel proteins associated with the disease. 10-plex TMT (tandem mass tags)-based quantitative proteomics experiments were performed using frozen tissue samples from the left prefrontal cortex of AD patients and healthy individuals and vascular dementia (VD) and frontotemporal dementia (FTD) patients as controls (CT). LC–MS/MS analyses were performed using a Q Exactive mass spectrometer.
Results
In total, 3281 proteins were identified and quantified using MaxQuant. Among them, after statistical analysis with Perseus (
p
value < 0.05), 16 and 155 proteins were defined as upregulated and downregulated, respectively, in AD compared to CT (Healthy, FTD and VD) with an expression ratio ≥ 1.5 (upregulated) or ≤ 0.67 (downregulated). After bioinformatics analysis, ten dysregulated proteins were selected as more prone to be associated with AD, and their dysregulation in the disease was verified by qPCR, WB, immunohistochemistry (IHC), immunofluorescence (IF), pull-down, and/or ELISA, using tissue and plasma samples of AD patients, patients with other dementias, and healthy individuals.
Conclusions
We identified and validated novel AD-associated proteins in brain tissue that should be of further interest for the study of the disease. Remarkably, PMP2 and SCRN3 were found to bind to amyloid-β (Aβ) fibers in vitro, and PMP2 to associate with Aβ plaques by IF, whereas HECTD1 and SLC12A5 were identified as new potential blood-based biomarkers of the disease.
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