A chemical genomics-aggrephagy integrated method studying functional analysis of autophagy inducers.

Macroautophagy/autophagy plays a critical role in the pathogenesis of various human diseases including neurodegenerative disorders such as Parkinson disease (PD) and Huntington disease (HD). Chemical autophagy inducers are expected to serve as disease-modifying agents by eliminating cytotoxic/damaged proteins. Although many autophagy inducers have been identified, their precise molecular mechanisms are not fully understood because of the complicated crosstalk among signaling pathways. To address this issue, we performed several chemical genomic analyses enabling us to comprehend the dominancy among the autophagy-associated pathways followed by an aggresome-clearance assay. In a first step, more than 400 target-established small molecules were assessed for their ability to activate autophagic flux in neuronal PC12D cells, and we identified 39 compounds as autophagy inducers. We then profiled the autophagy inducers by testing their effect on the induction of autophagy by 200 well-established signal transduction modulators. Our principal component analysis (PCA) and clustering analysis using a dataset of "autophagy profiles" revealed that two Food and Drug Administration (FDA)-approved drugs, memantine and clemastine, activate endoplasmic reticulum (ER) stress responses, which could lead to autophagy induction. We also confirmed that SMK-17, a recently identified autophagy inducer, induced autophagy via the PRKC/PKC-TFEB pathway, as had been predicted from PCA. Finally, we showed that almost all of the autophagy inducers tested in this present work significantly enhanced the clearance of the protein aggregates observed in cellular models of PD and HD. These results, with the combined approach, suggested that autophagy-activating small molecules may improve proteinopathies by eliminating nonfunctional protein aggregates.Abbreviations: ADK: adenosine kinase; AMPK: AMP-activated protein kinase; ATF4: activating transcription factor 4; BECN1: beclin-1; DDIT3/CHOP: DNA damage inducible transcript 3; EIF2AK3/PERK: eukaryotic translation initiation factor 2 alpha kinase 3; EIF2S1/eIF2α: eukaryotic translation initiation factor 2 subunit alpha; ER: endoplasmic reticulum; ERN1/IRE1α: endoplasmic reticulum to nucleus signaling 1; FDA: Food and Drug Administration; GSH: glutathione; HD: Huntington disease; HSPA5/GRP78: heat shock protein family A (Hsp70) member 5; HTT: huntingtin; JAK: Janus kinase, MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MAP2K/MEK: mitogen-activated protein kinase kinase; MAP3K8/Tpl2: mitogen-activated protein kinase kinase kinase 8; MAPK: mitogen-activated protein kinase; MPP+: 1-methyl-4-phenylpyridinium; MTOR: mechanistic target of rapamycin kinase; MTORC: MTOR complex; NAC: N-acetylcysteine; NGF: nerve growth factor 2; NMDA: N-methyl-D-aspartate; PCA: principal component analysis; PD: Parkinson disease; PDA: pancreatic ductal adenocarcinoma; PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3; PMA: phorbol 12-myristate 13-acetate; PRKC/PKC: protein kinase C; ROCK: Rho-associated coiled-coil protein kinase; RR: ribonucleotide reductase; SIGMAR1: sigma non-opioid intracellular receptor 1; SQSTM1/p62: sequestosome 1; STK11/LKB1: serine/threonine kinase 11; TFEB: Transcription factor EB; TGFB/TGF-ß: Transforming growth factor beta; ULK1: unc-51 like autophagy activating kinase 1; XBP1: X-box binding protein 1.

In vitro Antimicrobial Effects of Virus Block, Which Contains Multiple Polyoxometalate Compounds, and Hygienic Effects of Virus Block-Supplemented Moist Hand Towels.

BACKGROUND: Historical evidence has verified the multifaceted antiviral efficacy of polyoxometalates (PMs). METHODS: We carried out a study to investigate the antimicrobial effects of each of the 5 substances comprising virus block (VB): 3 PMs that have antibacterial and antiviral activity, an antibiotic agent, and an antibacterial agent. We also investigated the effectiveness of the addition of VB to moist hand towels in a study involving 120 volunteers. The time-dependent changes in metal ion concentrations in aqueous VB solution were analyzed using inductively coupled plasma atomic emission spectroscopy. RESULTS: The metal elements in the aqueous VB solution remained stable for 12 weeks without undergoing time-dependent changes. DISCUSSION: Further investigations were performed to study hand hygiene using moist hand towels in daily life settings. To this end, 120 volunteers provided 240 specimens that were used to investigate the presence of antibacterial compounds on the volunteers' hands before and after hand towel use. An aliquot of each specimen was suspended in phosphate-buffered saline and plated on agar media, and the number of colonies formed was counted. Normal bacterial flora found on the hands of the volunteers was investigated before and after the use of 4 different moist hand towels. CONCLUSIONS: The effects of VB and PMs were superior to those of commercial moist hand towels, indicating that effective data were obtained that may be useful for the practical application of the tested items.

The bHLH transcription factor Hand2 regulates the expression of nanog in ANS differentiation.

The basic helix-loop-helix transcription factor Hand2 is induced by bone morphogenetic proteins (BMPs) in neural crest-derived precursor cells during the early stage of development of the autonomic nervous system (ANS). Previous studies showed that Hand2 was essential for the ANS differentiation. However, regulatory mechanism of pluripotent genes has not been elucidated in ANS differentiation. Here, we show that Hand2 regulated nanog expression in ANS differentiation. Our studies demonstrated that the forced expression of Hand2 promoted the ANS differentiation program in P19 embryonal carcinoma (EC) cells without aggregation. Furthermore, our results suggested that Hand2 bound to the promoter of nanog, a gene required for embryonic stem cells self-renewal, and suppressed nanog expression after Hand2 induction. The rapid downregulation of nanog mRNA during ANS differentiation correlated with the Hand2 transcriptional activity and nanog promoter methylation. These findings are evidence for a presence of the novel regulatory mechanism of nanog in ANS differentiation.