Functional rejuvenation of aged neural stem cells by Plagl2 and anti-Dyrk1a activity

The regenerative potential of neural stem cells (NSCs) declines during aging, leading to cognitive dysfunctions. This decline involves up-regulation of senescence-associated genes, but inactivation of such genes failed to reverse aging of hippocampal NSCs. Because many genes are up-regulated or down-regulated during aging, manipulation of single genes would be insufficient to reverse aging. Here we searched for a gene combination that can rejuvenate NSCs in the aged mouse brain from nuclear factors differentially expressed between embryonic and adult NSCs and their modulators. We found that a combination of inducing the zinc finger transcription factor gene Plagl2 and inhibiting Dyrk1a, a gene associated with Down syndrome (a genetic disorder known to accelerate aging), rejuvenated aged hippocampal NSCs, which already lost proliferative and neurogenic potential. Such rejuvenated NSCs proliferated and produced new neurons continuously at the level observed in juvenile hippocampi, leading to improved cognition. Epigenome, transcriptome, and live-imaging analyses indicated that this gene combination induces up-regulation of embryo-associated genes and down-regulation of age-associated genes by changing their chromatin accessibility, thereby rejuvenating aged dormant NSCs to function like juvenile active NSCs. Thus, aging of NSCs can be reversed to induce functional neurogenesis continuously, offering a way to treat age-related neurological disorders.

Human Hepatocyte Nuclear Factors (HNF1 and LXRb) Regulates CYP7A1 in HIV-Infected Black South African Women with Gallstone Disease: A Preliminary Study

Background: Female sex, high estrogen levels, aging, obesity and dyslipidemia are some of the risk factors associated with gallstone formation. HIV infected patients on combination antiretroviral therapy (cART) are more prone to hypercholesterolemia. Bile acid synthesis is initiated by cholesterol 7-alpha hydroxylase (CYP7A1) and regulated by hepatocyte nuclear factors (HNF1α, HNF4α and LXRb).Aim/ Objective: To evaluate the expression of HNF1α, HNF4α, LXRb and miRNAs (HNF4α specific: miR-194-5p and miR-122*_1) that regulate CYP7A1 transcription in HIV-infected Black South African women on cART and presenting with gallstones relative to HIV negative patients with gallstone disease. Methods: Females (n = 96) presenting with gallstone disease were stratified based on HIV status. The gene expression of CYP7A1, HNF1α, HNF4α, LXRb, miR-194-5p and miR-122*_1 was determined using RT-qPCR. Messenger RNA and miRNA levels were reported as fold change expressed as 2-ΔΔCt (RQ min; RQ max). Fold changes >2 and <0.5 were considered significant. Results: HIV-infected females were older in age (p = 0.0267) and displayed higher low-density lipoprotein cholesterol (LDL-c) (p = 0.0419), CYP7A1 [2.078-fold (RQ min: 1.278; RQ max: 3.381)], LXRb [2.595-fold (RQ min: 2.001; RQ max: 3.000)] and HNF1α [3.428 (RQ min: 1.806; RQ max: 6.507] levels. HNF4α [0.642-fold (RQ min: 0.266; RQ max: 1.55)], miR-194-5p [0.527-fold (RQ min: 0.37; RQ max: 0.752)] and miR-122*_1 [0.595-fold (RQ min: 0.332; RQ max: 1.066)] levels were lower in HIV-infected females. Conclusions: HIV-infected women with gallstone disease displayed higher LDL-c levels and increased bile acid synthesis which was evident by the elevated expression of CYP7A1, HNF1α and LXRb. This could have been further influenced by cART and aging.

PRMT6 physically associates with nuclear factor Y to regulate photoperiodic flowering in Arabidopsis

The timing of floral transition is critical for reproductive success in flowering plants. In long-day (LD) plant Arabidopsis, the floral regulator gene FLOWERING LOCUS T (FT) is a major component of the mobile florigen. FT expression is rhythmically activated by CONSTANS (CO), and specifically accumulated at dusk of LDs. However, the underlying mechanism of adequate regulation of FT transcription in response to day-length cues to warrant flowering time still remains to be investigated. Here, we identify a homolog of human protein arginine methyltransferases 6 (HsPRMT6) in Arabidopsis, and confirm AtPRMT6 physically interacts with three positive regulators of flowering Nuclear Factors YC3 (NF-YC3), NF-YC9, and NF-YB3. Further investigations find that AtPRMT6 and its encoding protein accumulate at dusk of LDs. PRMT6-mediated H3R2me2a modification enhances the promotion of NF-YCs on FT transcription in response to inductive LD signals. Moreover, AtPRMT6 and its homologues proteins AtPRMT4a and AtPRMT4b coordinately inhibit the expression of FLOWERING LOCUS C, a suppressor of FT. Taken together, our study reveals the role of arginine methylation in photoperiodic pathway and how the PRMT6-mediating H3R2me2a system interacts with NF-CO module to dynamically control FT expression and facilitate flowering time.

DEAD-box RNA helicase 18 disrupts IRF3-binding to the interferon-β promoter

The production of type I interferons (IFN-α/β) requires strict control to avoid excessive activation during viral infections. The binding of interferon regulatory factor 3 (IRF3) to the IFN-β promoter region in the nucleus is essential for IFN-β transcription; however, whether nuclear factors have important negative-regulatory roles in this process is largely unknown. By screening for IRF3-interacting partners in the nucleus, we identified DEAD-box RNA helicase 18 (DDX18) as an important negative regulator of intranuclear IRF3. Overexpression of DDX18 suppressed virus- and IRF3-induced IFN-β production, whereas knockdown of DDX18 expression or knockout of the DDX18 gene had opposite effects. Mechanistically, DDX18 interacts with IRF3 and decreases the binding of IRF3 to the IFN-β promoter after viral infection. DDX18 knockdown mice (Ddx18+/-) further demonstrated that DDX18 suppressed antiviral innate immunity in vivo. Thus, despite many members of the DDX family act as important positive regulators in the cytoplasm, DDX18 plays a unique "braking" role in balancing virus-induced type I IFN production.

Beta-Cell Dysfunction Induced by Tacrolimus: A Way to Explain Type 2 Diabetes?

The combination of insulin resistance and β-cells dysfunction leads to the onset of type-2 diabetes mellitus (T2DM). This process can last for decades, as β-cells are able to compensate the demand for insulin and maintain normoglycemia. Understanding the adaptive capacity of β-cells during this process and the causes of its failure is essential to the limit onset of diabetes. Post-transplant diabetes mellitus (PTDM) is a common and serious disease that affects 30% of renal transplant recipients. With the exception of immunosuppressive therapy, the risk factors for T2D are the same as for PTDM: obesity, dyslipidaemia, insulin resistance and metabolic syndrome. Tacrolimus (TAC) is the immunosuppressant of choice after renal transplantation but it has the highest rates of PTDM. Our group has shown that insulin resistance and glucolipotoxicity, without favouring the appearance of apoptosis, modify key nuclear factors for the maintenance of identity and functionality of β-cells. In this context, TAC accelerates or enhances these changes. Our hypothesis is that the pathways that are affected in the progression from pre-diabetes to diabetes in the general population are the same pathways that are affected by TAC. So, TAC can be considered a tool to study the pathogenesis of T2DM. Here, we review the common pathways of β-cells dysfunction on T2DM and TAC-induced diabetes.

Lipid chaperone LBP-8 coordinates with nuclear factors to promote longevity in Caenorhabditis elegans

Eukaryotic cells are composed of a variety of organelles. Their coordination plays crucial roles in cellular homeostasis and organism longevity and is mediated by proteins with the ability to transport between different organelles. In Caenorhabditis elegans, LBP-8 is a pro-longevity lipid chaperone that can localize to both lysosomes and the nucleus. Here we profiled LBP-8’s binding partners using immunoprecipitation and mass spectrometry. From the 45 identified candidates, we discovered four nuclear factors that are required for the LBP-8-induced longevity. Among them, RPC-2, an RNA Polymerase III core subunit, is also necessary for the nuclear localization of LBP-8. Moreover, we have screened nuclear transport machinery components, and revealed the requirement of the nuclear import, not export, for the LBP-8 longevity effects. Together, these results suggest that the lipid chaperone LBP-8 relies on specific nuclear factors to retain in the nucleus and regulate longevity.

CREB5 reprograms nuclear interactions to promote resistance to androgen receptor targeting therapies

Metastatic castration resistant prostate cancers (mCRPC) are treated with therapies that antagonize the androgen receptor (AR). Nearly all patients develop resistance to AR-targeted therapies (ART). Our previous work identified CREB5 as an upregulated target gene in human mCRPC that promoted resistance to all clinically-approved ART. The mechanisms by which CREB5 promotes progression of mCRPC or other cancers remains elusive. Integrating ChIP-seq and rapid immunoprecipitation and mass spectroscopy of endogenous proteins (RIME), we report that cells overexpressing CREB5 demonstrate extensive reprogramming of nuclear protein-protein interactions in response to the ART agent enzalutamide. Specifically, CREB5 physically interacts with AR, the pioneering actor FOXA1, and other known co-factors of AR and FOXA1 at transcription regulatory elements recently found to be active in mCRPC patients. We identified a subset of CREB5/FOXA1 co-interacting nuclear factors that have critical functions for AR transcription (GRHL2, HOXB13) while others (TBX3, NFIC) regulated cell viability and ART resistance and were amplified or overexpressed in mCRPC. Upon examining the nuclear protein interactions and the impact of CREB5 expression on the mCRPC patient transcriptome, we found CREB5 was associated with TGF-beta; and Wnt signaling and epithelial to mesenchymal transitions, implicating these pathways in ART resistance. Overall, these observations define the molecular interactions among CREB5, FOXA1, and pathways that promote ART resistance.

Ex Vivo Study on the Antioxidant Activity of a Winemaking By-Product Polyphenolic Extract (Taurisolo®) on Human Neutrophils

Oxidative stress (OxS) has been linked to several chronic diseases and is recognized to have both major causes and consequences. The use of antioxidant-based nutraceuticals has been licensed as an optimal tool for management of OxS-related diseases. Currently, great interest is focused on the valorization of agri-food by-products as a source of bioactive compounds, including polyphenols. In this sense, we evaluated the efficacy of a novel nutraceutical formulation based on polyphenolic extract from Aglianico cultivar grape pomace (registered as Taurisolo®). In particular, we tested both native and in vitro gastrointestinal digested forms. The two extracts have been used to treat ex vivo neutrophils from subjects with metabolic syndrome, reporting a marked antioxidant activity of Taurisolo®, as shown by its ability to significantly reduce both the levels of reactive oxygen species (ROS) and the activities of catalase and myeloperoxidase in the cell medium after stimulation of neutrophils with phorbol 12-myristate 13-acetate (PMA). Interestingly, we observed an increase in intracellular enzymatic activities in PMA-treated cells, suggesting that Taurisolo® polyphenols might be able to activate nuclear factors, up-regulating the expression of this target antioxidant gene. In addition, Taurisolo® reversed the increase in malondialdehyde induced by PMA; reduced the expression of pro-inflammatory genes such as cyclooxygenase 2 (COX-2), tumor necrosis factor alpha (TNFα) and myeloperoxidase (MPO); and induced the expression of the anti-inflammatory cytokine IL-10. Overall, these results suggest the efficacy of Taurisolo® in contrasting the OxS at blood level, providing evidence for its therapeutic potential in the management of OxS-related pathological conditions in humans.

Rifampicin Induces Gene, Protein, and Activity of P-Glycoprotein (ABCB1) in Human Precision-Cut Intestinal Slices

P-glycoprotein (ABCB1), an ATP-binding cassette efflux transporter, limits intestinal absorption of its substrates and is a common site of drug–drug interactions. Drug-mediated induction of intestinal ABCB1 is a clinically relevant phenomenon associated with significantly decreased drug bioavailability. Currently, there are no well-established human models for evaluating its induction, so drug regulatory authorities provide no recommendations for in vitro/ex vivo testing drugs’ ABCB1-inducing activity. Human precision-cut intestinal slices (hPCISs) contain cells in their natural environment and express physiological levels of nuclear factors required for ABCB1 induction. We found that hPCISs incubated in William’s Medium E for 48 h maintained intact morphology, ATP content, and ABCB1 efflux activity. Here, we asked whether rifampicin (a model ligand of pregnane X receptor, PXR), at 30 μM, induces functional expression of ABCB1 in hPCISs over 24- and 48-h incubation (the time to allow complete induction to occur). Rifampicin significantly increased gene expression, protein levels, and efflux activity of ABCB1. Moreover, we described dynamic changes in ABCB1 transcript levels in hPCISs over 48 h incubation. We also observed that peaks of induction are achieved among donors at different times, and the extent of ABCB1 gene induction is proportional to PXR mRNA levels in the intestine. In conclusion, we showed that hPCISs incubated in conditions comparable to those used for inhibition studies can be used to evaluate drugs’ ABCB1-inducing potency in the human intestine. Thus, hPCISs may be valuable experimental tools that can be prospectively used in complex experimental evaluation of drug–drug interactions.