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Peroxisome proliferator-activated receptor γ (PPARγ) is the master transcriptional regulator of adipocytes and the cellular target of thiazolidinedione (TZD) drugs. Suppression of pro-inflammatory actions, including pro-inflammatory gene expression and lipolysis in adipocytes, contributes to PPARγ-mediated anti-diabetic effects of TZDs. However, adverse side effects largely limited the clinical use of TZDs, despite their potent insulin-sensitizing effects. Therefore, it is important to understand how PPARγ is regulated. Thyroid hormone receptor-associated protein 3 (THRAP3) was previously reported to promote diabetic gene expression by acting as a transcriptional coregulator of PPARγ in adipocytes. Therefore, we tested if THRAP3 modulated anti-inflammatory functions of PPARγ in 3T3-L1 adipocytes. THRAP3 depletion increased basal and tumor necrosis factor α (TNFα)-induced lipolysis, pro-inflammatory gene expression, and phosphorylation of extracellular signal-regulated kinases (ERKs), suggesting elevated pro-inflammatory response after THRAP3 depletion in adipocytes. Moreover, TZD-mediated suppression of TNFα-induced lipolysis, pro-inflammatory gene expression, and ERK phosphorylation was attenuated or alleviated after THRAP3 depletion. Interestingly, the mRNA and protein levels of PPARγ were greatly reduced in THRAP3-depleted adipocytes. Actinomycin D treatment revealed that the stability of PPARγ mRNA was greatly reduced by THRAP3 depletion in adipocytes. Thus, in addition to modulating PPARγ function, THRAP3 may directly regulate the transcript of PPARγ in differentiated adipocytes.

MicroRNAs (miRNAs), a class of non-coding single-strand RNA molecules encoded by endogenous genes, are about 21-24 nucleotides long and are involved in the post-transcriptional regulation of gene expression in plants and animals. Generally, the types and quantities of miRNAs in the different tissues of an organism are diverse, and these divergences may be related to their specific functions. Here we have identified 296 known miRNAs and 46 novel miRNAs in the antennae of the parasitoid wasp Microplitis mediator by high-throughput sequencing. Thirty-three miRNAs were predicted to target olfactory-associated genes, including odorant binding proteins (OBPs), chemosensory proteins, odorant receptors (ORs), ionotropic receptors (IRs) and gustatory receptors. Among these, 17 miRNAs were significantly highly expressed in the antennae, four miRNAs were highly expressed both in the antennae and head or wings, while the remaining 12 miRNAs were mainly expressed in the head, thorax, abdomen, legs and wings. Notably, miR-9a-5p and miR-2525-3p were highly expressed in male antennae, whereas miR-1000-5p and novel-miR-13 were enriched in female antennae. The 17 miRNAs highly expressed in antennae are likely to be associated with olfaction, and were predicted to target one OBP (targeted by miR-3751-3p), one IR (targeted by miR-7-5p) and 14 ORs (targeted by 15 miRNAs including miR-6-3p, miR-9a-5p, miR-9b-5p, miR-29-5p, miR-71-5p, miR-275-3p, miR-1000-5p, miR-1000-3p, miR-2525-3p, miR-6012-3p, miR-9719-3p, novel-miR-10, novel-miR-13, novel-miR-14 and novel-miR-28). These candidate olfactory-associated miRNAs are all likely to be involved in chemoreception through the regulation of chemosensory gene expression in the antennae of M. mediator.

The E3 ubiquitin ligase ring finger protein 115 (RNF115) is overexpressed in more than half of human breast tumors and is implicated in the pathogenesis and progression of breast cancer. However, the mechanism behind RNF115 overexpression in breast tumors remains largely unknown. Here we report that ubiquitin-specific protease 9X (USP9X), a substrate-specific deubiquitinating enzyme, stabilizes RNF115 and thereby regulates its biological functions in breast cancer cells. Immunoprecipitation and GST pull-down assays showed that USP9X interacted with RNF115. Depletion of RNF115 by siRNAs or overexpression of RNF115 did not significantly affect USP9X expression. In contrast, knockdown of USP9X in breast cancer cells by siRNAs reduced RNF115 protein abundance, which was partially restored following treatment with proteasome inhibitor MG-132. Moreover, depletion of USP9X reduced the half-life of RNF115 and increased its ubiquitination. Conversely, overexpression of USP9X resulted in an accumulation of RNF115 protein, accompanied by a decrease in its ubiquitination. RNF115 mRNA levels were unaffected by overexpression or knockdown of USP9X. Furthermore, USP9X protein expression levels correlated positively with RNF115 in breast cancer cell lines and breast tumor samples. Importantly, reintroduction of RNF115 in USP9X-depleted cells partially rescued the reduced proliferation, migration, and invasion of breast cancer cells by USP9X knockdown. Collectively, these findings indicate that USP9X is a stabilizer of RNF115 protein and that the USP9X-RNF115 signaling axis is implicated in the breast cancer malignant phenotype.

Genetically modified (GM) organisms have been developed for decades. However, unintended effects are the main concerns of safety assessment that needs to be carefully investigated. Here, eight varieties of GM rice that were developed in China were selected to assess the unintended effects through transcriptome and metabolism. There are 2892–8758 differentially expressed genes (DEGs) and 7–50 metabolites at significant level between GM varieties and their isogenic counterparts, which were far fewer than that between traditional rice varieties. The function enrichment analysis showed altered transcription in stress-related pathway and starch and sucrose metabolism. DEGs shared among eight GM samples constitute less than 1% of the genes in the genome, and none of them is reported more than four times. The insertion effect on the nearby gene expression and the associated metabolism is only restricted to 50 genes. All the results provide a comprehensive analysis of unintended effects and indication of difference in Chinese transgenic rice based on their backgrounds, transformation, and insertion elements.

We previously identified a causal link between a rare patient mutation in DISC1 (disrupted-in-schizophrenia 1) and synaptic deficits in cortical neurons differentiated from isogenic patient-derived induced pluripotent stem cells (iPSCs). Here we find that transcripts related to phosphodiesterase 4 (PDE4) signaling are significantly elevated in human cortical neurons differentiated from iPSCs with the DISC1 mutation and that inhibition of PDE4 or activation of the cAMP signaling pathway functionally rescues synaptic deficits. We further generated a knock-in mouse line harboring the same patient mutation in the Disc1 gene. Heterozygous Disc1 mutant mice exhibit elevated levels of PDE4s and synaptic abnormalities in the brain, and social and cognitive behavioral deficits. Pharmacological inhibition of the PDE4 signaling pathway rescues these synaptic, social and cognitive behavioral abnormalities. Our study shows that patient-derived isogenic iPSC and humanized mouse disease models are integral and complementary for translational studies with a better understanding of underlying molecular mechanisms., Previous work has shown in iPSC derived neurons that synaptic impairments are associated with a 4bp DISC1 deletion. Here the authors demonstrate a role for the PDE4 signalling pathway in these synaptic impairments.

Background: Chronic intermittent hypoxia is the most remarkable feature of obstructive sleep apnea/hypopnea syndrome and it can induce the change of hypoxia-inducible factor-1α (HIF-1α) expression and contractile properties in the genioglossus. To clarify the role of HIF-1α in contractile properties of the genioglossus, this study generated and compared high-throughput RNA-sequencing data from genioglossus between HIF-1α conditional knockout (KO) mice and littermate wild-type (WT) mice. Methods: KO mice were generated with cre-loxP strategy. Gene expression profile analysis was performed using gene enrichment analysis. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses of differently expressed messenger RNAs were performed to identify the related pathways and biological functions. Six differentially expressed genes (DEGs) were validated by qualitative reverse transcription polymerase chain reaction. Results: A total of 142 (77 upregulated and 65 downregulated) transcripts were found to exhibit statistically significant difference between the HIF-1α-KO and WT mice. GO and KEGG analyses indicated that DEGs included genes involved in "skeletal muscle cell differentiation," "muscle organ development," "glucose metabolic process," "glycogen biosynthetic and metabolic process," etc. Conclusion: This study might provide evidence that HIF-1α affects the expression of multiple genes involved in the myogenesis, muscle development, and carbohydrate metabolism through transcriptome analysis in conditional HIF-1α-KO mice. Key words: Carbohydrate Metabolism; Genioglossus; Hypoxia-inducible Factor-1α; Myogenesis; Transcriptomic Analysis

Splicing expands, reshapes, and regulates the transcriptome of eukaryotic organisms. Despite its importance, key questions remain unanswered, including the following: Can splicing evolve when organisms adapt to new challenges? How does evolution optimize inefficiency of introns’ splicing and of the splicing machinery? To explore these questions, we evolved yeast cells that were engineered to contain an inefficiently spliced intron inside a gene whose protein product was under selection for an increased expression level. We identified a combination of mutations in Cis (within the gene of interest) and in Trans (in mRNA-maturation machinery). Surprisingly, the mutations in Cis resided outside of known intronic functional sites and improved the intron’s splicing efficiency potentially by easing tight mRNA structures. One of these mutations hampered a protein’s domain that was not under selection, demonstrating the evolutionary flexibility of multi-domain proteins as one domain functionality was improved at the expense of the other domain. The Trans adaptations resided in two proteins, Npl3 and Gbp2, that bind pre-mRNAs and are central to their maturation. Interestingly, these mutations either increased or decreased the affinity of these proteins to mRNA, presumably allowing faster spliceosome recruitment or increased time before degradation of the pre-mRNAs, respectively. Altogether, our work reveals various mechanistic pathways toward optimizations of intron splicing to ultimately adapt gene expression patterns to novel demands., An experimental evolution study involving an inefficiently spliced intron reveals that the splicing machinery, introns, and RNA quality control factors evolve in Cis and in Trans when cells optimize their transcriptome to new challenges.

Ionotropic receptors (IRs) mainly detect the acids and amines having great importance in many insect species, representing an ancient olfactory receptor family in insects. In the present work, we performed RNAseq of Microplitis mediator antennae and identified seventeen IRs. Full-length MmedIRs were cloned and sequenced. Phylogenetic analysis of the Hymenoptera IRs revealed that ten MmedIR genes encoded "antennal IRs" and seven encoded "divergent IRs". Among the IR25a orthologous groups, two genes, MmedIR25a.1 and MmedIR25a.2, were found in M. mediator. Gene structure analysis of MmedIR25a revealed a tandem duplication of IR25a in M. mediator. The tissue distribution and development specific expression of the MmedIR genes suggested that these genes showed a broad expression profile. Quantitative gene expression analysis showed that most of the genes are highly enriched in adult antennae, indicating the candidate chemosensory function of this family in parasitic wasps. Using immunocytochemistry, we confirmed that one co-receptor, MmedIR8a, was expressed in the olfactory sensory neurons. Our data will supply fundamental information for functional analysis of the IRs in parasitoid wasp chemoreception.

SummaryAlternative splicing is a major mechanism of controlling gene expression and protein diversity in higher eukaryotes. We report that the splicing factor SRSF10 functions during striated muscle development, myoblast differentiation, and glucose production both in cells and in mice. A combination of RNA-sequencing and molecular analysis allowed us to identify muscle-specific splicing events controlled by SRSF10 that are critically involved in striated muscle development. Inclusion of alternative exons 16 and 17 of Lrrfip1 is a muscle-specific event that is activated by SRSF10 and essential for myoblast differentiation. On the other hand, in mouse primary hepatocytes, PGC1α is a key target of SRSF10 that regulates glucose production by fasting. SRSF10 represses inclusion of PGC1α exon 7a and facilitates the production of functional protein. The results highlight the biological significance of SRSF10 and regulated alternative splicing in vivo.

Chronic cerebral hypoperfusion (CCH) causes learning and memory impairments and increases the risk of Alzheimer disease (AD) and vascular dementia (VD) through several biologically plausible pathways, yet the mechanisms underlying the disease process remained unclear particularly in a temporal manner. We performed permanent bilateral occlusion of the common carotid arteries (two-vessel occlusion, 2VO) to induce CCH. To determine whether hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are altered at different stages of cognitive impairment caused by CCH, adult male SD rats were randomly distributed into sham-operated 4, 8 and 12weeks group, 2VO 4, 8 and 12weeks group. Learning and memory performance were evaluated with Morris water maze (MWM) and long-term potentiation (LTP) was used to address the underlying synaptic mechanisms. Expression of NeuN, HCN1 and HCN2 in hippocampal CA1, DG and CA3 areas was quantified by immunohistochemistry and western blotting. Our data showed that CCH induced a remarkable spatial learning and memory deficits in rats of 2VO 4, 8, and 12weeks group although neuronal loss only occurred after 4weeks of 2VO surgery in CA1. In addition, a significant reduction of HCN1 surface expression in CA1 was observed in the group that suffered 4weeks ischemia but neither 8 nor 12weeks. However, HCN2 surface expression in CA1 increased throughout the ischemia time-scales (4, 8 and 12w). Our findings indicate spatial learning and memory deficits in the CCH model are associated with disturbed HCN1 and HCN2 surface expression in hippocampal CA1. The altered patterns of both HCN1 and HCN2 surface expression may be implicated in the early stage (4w) of spatial learning and memory impairments; and the stable and long-lasting impairments of spatial learning and memory may partially attribute to the up-regulated HCN2 surface expression.

Nm23 is a family of genes encoding the nucleoside diphosphate (NDP) kinase, which functions in a wide variety of biological processes, including growth, development, differentiation and tumor metastasis. In this study, a novel nm23 gene, designated as Mrnm23, was identified from the freshwater giant prawn Macrobrachium rosenbergii. The full-length cDNA was 776bp in length, encoding for a protein of 176 amino acids with one typical NDP kinase domain that harbored all the crucial residues for nucleotide binding and enzymatic activity. Like human novel nm23-H1B, the putative protein contained a unique 21-amino-acid NH2-terminal extension as compared to human nm23 (nm23-H1) homologs. Further, 3 extra amino acid residues prolonged the COOH-terminus. The Mrnm23 was ubiquitously expressed in all tissues examined, including androgenic gland, gill, heart, liver, muscle, ovary, and testis. In situ hybridization to gonad sections indicated that the Mrnm23 mRNA was localized in the cytoplasm of cup-base of differentiating spermatids, in the spike of the umbrella-shaped spermatozoa and in the cytoplasm of the early previtellogenic oocytes, suggesting that the Mrnm23 has potential roles in spermiogenesis and early differentiation of oocyte.

The G0/G1 switch gene 2 (G0S2) is rapidly induced by all-trans-retinoic acid (RA)-treatment of acute promyelocytic leukemia (APL) and other cells. G0S2 regulates lipolysis via inhibition of adipose triglyceride lipase (ATGL). This study found that retinoic acid receptor (RAR), but not retinoid X receptor (RXR) agonists induced G0S2 expression in APL cells. Novel G0S2 functions were uncovered that included repression of exogenous gene expression and transcriptional activity. Transient G0S2 transfection repressed the activities of multiple reporter constructs (including the retinoid-regulated species RARβ, UBE1L and G0S2); this occurred in diverse cell contexts. This inhibition was antagonized by siRNA-mediated G0S2 knockdown. To determine the inhibitory effects were not due to transient G0S2 expression, G0S2 was stably overex-pressed in cells without appreciable basal G0S2 expression. As expected, this repressed transcriptional activities. Intriguingly, transfection of G0S2 did not affect endogenous RARβ, UBE1L or G0S2 expression. Hence, only exogenously expressed genes were affected by G0S2. The domain responsible for this repression was localized to the G0S2 hydrophobic domain (HD). This was the same region responsible for the ability of G0S2 to inhibit ATGL activity. Whether an interaction with ATGL accounted for this new G0S2 activity was studied. Mimicking the inhibition of ATGL by oleic acid treatment that increased lipid droplet size or ATGL siRNA knockdown did not recapitulate G0S2 repressive effects. Engineered gain of ATGL expression did not rescue G0S2 transcriptional repression either. Thus, transcriptional repression by G0S2 did not depend on the ability of G0S2 to inhibit ATGL. Subcellular localization studies revealed that endogenous and exogenously-expressed G0S2 proteins were localized to the cytoplasm, particularly in the perinuclear region. Expression of a mutant G0S2 species that lacked the HD domain altered cytosolic G0S2 localization. This linked G0S2 subcellular localization to G0S2 transcriptional repression. The potential mechanisms responsible for this G0S2 repression are examined.

The purpose of this study was to examine the MAC30 expression in non–small cell lung cancer and to evaluate its prognostic value on therapeutic response in patients with non–small cell lung cancer receiving postoperative chemotherapy. Among a total of 218 retrospective Chinese patients with non–small cell lung cancer, 164 patients receiving adjuvant chemotherapy were enrolled in this study. Real-time polymerase chain reaction was performed to confirm the expression of MAC30 messenger RNA in 32 cases of non–small cell lung cancer tumors with the corresponding nontumor lung tissues. The MAC30 protein expression in all specimens was analyzed by immunohistochemical staining. Moreover, we assessed the correlation of MAC30 expression with clinicopathological features, therapeutic response, and survival of patients. Here, we observed the increased expression of MAC30 messenger RNA in patients with non–small cell lung cancer compared to those in control samples. The overexpression of MAC30 was strongly associated with poor tumor differentiation, high tumor–node–metastasis stage, and lymph node metastasis. In addition, we observed that patients with increased MAC30 expression showed gloomy overall survival and disease-free survival. A multivariate analysis explicated that higher MAC30 expression was a valuable independent prognostic factor of poorer tumor differentiation, shorter overall survival, and disease-free survival in patients receiving chemotherapy. MAC30 could be a useful biomarker of tumor differentiation and outcome of patients with non–small cell lung cancer. Overexpression of MAC30 predicts a worse tumor differentiated stage and prognosis in patients with non–small cell lung cancer receiving adjuvant chemotherapy.

Immunity and metabolism are closely linked. The liver is an important metabolic organ in the body. However, the interactions between hepatocytes and the immune system are poorly understood. In mice developing concanavalin A (ConA)-induced hepatitis (CIH), we found extensive lipid accumulation in hepatocytes. Critical enzyme involved in fat synthesis such as stearoyl-CoA desaturase 1 (SCD1) was upregulated. When we injected ConA to SCD1-deficient mice, we found these mice to be highly resistant to CIH. The mechanisms of the protective effect of SCD1 deficiency might be attributed to the reduced leptin levels in those mice, which modulated critical cytokines and signaling pathways in CIH pathogenesis. In conclusion, our study suggests that SCD1 deficiency protects mice from liver injury in a leptin-dependent manner.

Objective To explore the role of P2X 3 receptor in pain induced by experimental tooth movement. Design Male Sprague-Dawley rats weighing 200–300 g were used. P2X 3 receptor distribution in the caudal one-third portion of the trigeminal ganglion (TG) was studied by IHC. Next, the changes of P2X 3 expression were detected by Western blotting 4 h, 1 d, 2 d, 3 d, 5 d, 7 d, 14 d after tooth movement. We then developed a behaviour pain model associated with directed mouth wiping. Finally, the effect of TNP-ATP on nociceptive-like behaviour was evaluated. Results Our results showed that P2X 3 receptors were expressed mainly in small- and medium-sized cells and experimental tooth movement led to an increase in staining of mandibular P2X 3 receptors. In addition, following experimental tooth movement, the expression of P2X 3 receptor in TG was statistically significantly up-regulated from days 1 to 5, with a peak on day 3. It was also found that the time spent on directed mouth wiping was dramatically increased by experimental tooth movement from days 1 to 7. The rhythm change of P2X 3 receptor expression in TG and the mouth wiping behaviour were in concert with the initial orthodontic pain responses. The directed mouth wiping behaviour was modulated in a force-dependent manner and could be attenuated by peripheral and systemic morphine. Furthermore, peripherally administered TNP-ATP could exert an analgesic effect on this pain model. Conclusion These results suggest that directed mouth wiping behaviour can be a reliable measurement of pain following experimental tooth movement in rats. The P2X 3 receptor is important in the development and maintenance of tooth movement pain and thus may be peripheral targets for analgesics in orthodontic pain control.

Sensory organs are composed of neurons, which convert environmental stimuli to electrical signals, and glia-like cells, whose functions are not well understood. To decipher glial roles in sensory organs, we ablated the sheath glial cell of the major sensory organ of Caenorhabditis elegans . We found that glia-ablated animals exhibit profound sensory deficits and that glia provide activities that affect neuronal morphology, behavior generation, and neuronal uptake of lipophilic dyes. To understand the molecular bases of these activities, we identified 298 genes whose messenger RNAs are glia-enriched. One gene, fig-1 , encodes a labile protein with conserved thrombospondin TSP1 domains. FIG-1 protein functions extracellularly, is essential for neuronal dye uptake, and also affects behavior. Our results suggest that glia are required for multiple aspects of sensory organ function.

GABA receptors play an important role in ischemic brain injury. Studies have indicated that autophagy is closely related to neurodegenerative diseases. However, during chronic cerebral hypoperfusion, the changes of autophagy in the hippocampal CA1 area, the correlation between GABA receptors and autophagy and their influences on hippocampal neuronal apoptosis have not been well established. Here, we found that chronic cerebral hypoperfusion resulted in rat hippocampal atrophy, neuronal apoptosis, enhancement and redistribution of autophagy, down-regulation of Bcl-2/Bax ratio, elevation of cleaved caspase-3 levels, reduction of surface expression of GABAA receptor α1 subunit and an increase in surface and mitochondrial expression of connexin 43 (CX43) and CX36. Chronic administration of GABAB receptors agonist baclofen significantly alleviated neuronal damage. Meanwhile, baclofen could up-regulate the ratio of Bcl-2/Bax and increase the activation of Akt, GSK-3β and ERK which suppressed cytodestructive autophagy. The study also provided evidence that baclofen could attenuate the decrease in surface expression of GABAA receptor α1 subunit and down-regulate surface and mitochondrial expression of CX43 and CX36, which might enhance protective autophagy. The current findings suggested that, under chronic cerebral hypoperfusion, the effects of GABAB receptors activation on autophagy regulation could reverse neuronal damage.

Item does not contain fulltext Hfi is a dominant cataract mutation where heterozygotes show hydropic lens fibers and homozygotes show total lens opacity. The Hfi locus was mapped to the distal part of mouse chromosome 10 close to the major intrinsic protein (Mip), which is expressed only in cell membranes of lens fibers. Molecular analysis of Mip revealed a 76-bp deletion that resulted in exon 2 skipping in Mip mRNA. In Hfi/Hfi this deletion resulted in a complete absence of the wildtype Mip. In contrast, Hfi/+ animals had the same amount of wildtype Mip as +/+. Results from pulse-chase expression studies excluded hetero-oligomerization of wildtype and mutant Mip as a possible mechanism for cataract formation in the Hfi/+. We propose that the cataract phenotype in the Hfi heterozygote mutant is due to a detrimental gain of function by the mutant Mip resulting in either cytotoxicity or disruption in processing of other proteins important for the lens. Cataract formation in the Hfi/Hfi mouse is probably a combined result of both the complete loss of wildtype Mip and a gain of function of the mutant Mip.

Prion diseases are a group of transmissible fatal neurodegenerative disorders of humans and animals, including bovine spongiform encephalopathy, scrapie, and Creutzfeldt-Jakob disease. Microglia, the resident macrophages of the central nervous system, are exquisitely sensitive to pathological tissue alterations, altering their morphology and phenotype to adopt a so-called activated state and perform immunological functions in response to pathophysiological brain insults. Although recent findings have provided valuable insights into the role microglia play in the proinflammatory events observed in prion, the intracellular signaling molecules responsible for the initiation of these responses remain to be elucidated. It seems that microglial activation involve PrP106-126 binding and the activation of cell surface immune and adhesion molecules such as CD36 and integrins, with the subsequent recruitment of Src family tyrosine kinases such as Fyn, Lyn, and Syk kinases. In the present study, we show that CD36 is involved in PrP106-126-induced microglial activation and that PP2 and piceatannol (Pic) can abrogate neurotoxic prion peptides-induced inducible nitric oxide synthase activation in microglia. These findings unveil a previously unrecognized role of PP2 and Pic as Src family kinase Fyn and the tyrosine kinase Syk inhibitor involved in neurotoxic prion peptides-microglia interactions, thus providing new insights into mechanisms underlying the activation of microglia by neurotoxic prion peptides.

New pharmacologic targets are needed for lung cancer. One candidate pathway to target is composed of the E1-like ubiquitin-activating enzyme (UBE1L) that associates with interferon-stimulated gene 15 (ISG15), which complexes with and destabilizes cyclin D1. Ubiquitin protease 43 (UBP43/USP18) removes ISG15 from conjugated proteins. This study reports that gain of UBP43 stabilized cyclin D1, but not other D-type cyclins or cyclin E. This depended on UBP43 enzymatic activity; an enzymatically inactive UBP43 did not affect cyclin D1 stability. As expected, small interfering RNAs that reduced UBP43 expression also decreased cyclin D1 levels and increased apoptosis in a panel of lung cancer cell lines. Forced cyclin D1 expression rescued UBP43 apoptotic effects, which highlighted the importance of cyclin D1 in conferring this. Short hairpin RNA-mediated reduction of UBP43 significantly increased apoptosis and reduced murine lung cancer growth in vitro and in vivo after transplantation of these cells into syngeneic mice. These cells also exhibited increased response to all-trans-retinoic acid, interferon, or cisplatin treatments. Notably, gain of UBP43 expression antagonized these effects. Normal-malignant human lung tissue arrays were examined independently for UBP43, cyclin D1, and cyclin E immunohistochemical expression. UBP43 was significantly (P < 0.01) increased in the malignant versus normal lung. A direct relationship was found between UBP43 and cyclin D1 (but not cyclin E) expression. Differential UBP43 expression was independently detected in a normal-malignant tissue array with diverse human cancers. Taken together, these findings uncovered UBP43 as a previously unrecognized antineoplastic target. Mol Cancer Ther; 11(9); 1968–77. ©2012 AACR.

Microglial activation is a characteristic feature of the pathogenesis of prion diseases. The molecular mechanisms that underlie prion-induced microglial activation are not very well understood. In the present study, we investigated the role of the class B scavenger receptor CD36 in microglial activation induced by neurotoxic prion protein (PrP) fragment 106-126 (PrP(106-126)). We first examined the time course of CD36 mRNA expression upon exposure to PrP(106-126) in BV2 microglia. We then analyzed different parameters of microglial activation in PrP(106-126)-treated cells in the presence or not of anti-CD36 monoclonal antibody (mAb). The cells were first incubated for 1 h with CD36 monoclonal antibody to block the CD36 receptor, and were then treated with neurotoxic prion peptides PrP(106-126). The results showed that PrP(106-126) treatment led to a rapid yet transitory increase in the mRNA expression of CD36, upregulated mRNA and protein levels of proinflammatory cytokines (IL-1β, IL-6 and TNF-α), increased iNOS expression and nitric oxide (NO) production, stimulated the activation of NF-κB and caspase-1, and elevated Fyn activity. The blockade of CD36 had no effect on PrP(106-126)-stimulated NF-κB activation and TNF-α protein release, abrogated the PrP(106-126)-induced iNOS stimulation, downregulated IL-1β and IL-6 expression at both mRNA and protein levels as well as TNF-α mRNA expression, decreased NO production and Fyn phosphorylation, reduced caspase-1 cleavage induced by moderate PrP(106-126)-treatment, but had no effect on caspase-1 activation after treatment with a high concentration of PrP(106-126). Together, these results suggest that CD36 is involved in PrP(106-126)-induced microglial activation and that the participation of CD36 in the interaction between PrP(106-126) and microglia may be mediated by Src tyrosine kinases. Our findings provide new insights into the mechanisms underlying the activation of microglia by neurotoxic prion peptides and open perspectives for new therapeutic strategies for prion diseases by modulation of CD36 signaling.

Background— Pathological stresses induce heart failure in animal models through activation of multiple cardiac transcription factors (TFs) working cooperatively. However, interactions among TFs in human heart failure are less understood. Here, we use genomic data to examine the evidence that 5 candidate TF families coregulate gene expression in human heart failure. Methods and Results— RNA isolates from failing (n=86) and nonfailing (n=16) human hearts were hybridized with Affymetrix HU133A arrays. For each gene on the array, we determined conserved MEF2, NFAT, NKX , GATA , and FOX binding motifs within the −1-kb promoter region using human-murine sequence alignments and the TRANSFAC database. Across 9076 genes expressed in the heart, TF-binding motifs tended to cluster together in nonrandom patterns within promoters of specific genes ( P values ranging from 10 −2 to 10 −21 ), suggesting coregulation. We then modeled differential expression as a function of TF combinations present in promoter regions. Several combinations predicted increased odds of differential expression in the failing heart, with the highest odds ratios noted for genes containing both MEF2 and NFAT binding motifs together in the same promoter region (peak odds ratio, 3.47; P =0.005). Conclusions— These findings provide genomic evidence for coregulation of myocardial gene expression by MEF2 and NFAT in human heart failure. In doing so, they extend the paradigm of combinatorial regulation of gene expression to the human heart and identify new target genes for mechanistic study. More broadly, we demonstrate how integrating diverse sources of genomic data yields novel insight into human cardiovascular disorders.

Research has revealed a shift towards Th2 in many types of malignant tumor, but the state of Th1/Th2 is not clear in patients with primary hepatic cancer (PHC). This study was designed to determine the expression of Th1- versus Th2-type cytokines in primary hepatic cancer and the adjacent liver tissue in order to provide evidence for treatment of the Th1/Th2 shift.Samples were collected from 11 patients with PHC. The gene expression of Th1/Th2 cytokines was detected by reverse transcriptase polymerase chain reaction (RT-PCR) using IFN-gamma and IL-2 as Th1-type cytokine genes, and IL-4 and IL-10 as Th2-type cytokine genes.Th1-type cytokines were expressed in 7/11 PHCs and 9/11 adjacent liver tissues, while Th0 type cytokines occurred in 4/11 PHCs and 2/11 adjacent liver tissues.Th1-type cytokines are expressed predominantly in primary hepatic cancer and the adjacent liver tissue.

mahoganoid is a mouse coat-color mutation whose pigmentary phenotype and genetic interactions resemble those of Attractin ( Atrn ). Atrn mutations also cause spongiform neurodegeneration. Here, we show that a null mutation for mahoganoid causes a similar age-dependent neuropathology that includes many features of prion diseases but without accumulation of protease-resistant prion protein. The gene mutated in mahoganoid encodes a RING-containing protein with E3 ubiquitin ligase activity in vitro. Similarities in phenotype, expression, and genetic interactions suggest that mahoganoid and Atrn genes are part of a conserved pathway for regulated protein turnover whose function is essential for neuronal viability.

The expression of glutamate receptor/subunit mRNAs was examined 3 weeks after discontinuing 1 week of daily injections of saline or cocaine. The level of mRNA for GluR1-4, NMDAR1, and mGluR5 receptors was measured with in situ hybridization and RT-PCR. In nucleus accumbens, acute cocaine treatment significantly reduced the mRNA level for GluR3, GluR4, and NMDAR1 subunits, whereas repeated cocaine reduced the level for GluR3 mRNA. Acute cocaine treatment also reduced the NMDAR1 mRNA level in dorsolateral striatum and ventral tegmental area. In prefrontal cortex, repeated cocaine treatment significantly increased the level of GluR2 mRNA. The GluR2 mRNA level was not changed by acute or repeated cocaine in any other brain regions examined. Repeated cocaine treatment also significantly increased mGluR5 mRNA levels in nucleus accumbens shell and dorsolateral striatum. Functional properties of the ionotropic glutamate receptors are determined by subunit composition. In addition, metabotropic glutamate receptors can modulate synaptic transmission and the response to stimulation of ionotropic receptors. Thus, the observed changes in levels of AMPA and NMDA receptor subunits and the mGluR5 metabotropic receptor may alter excitatory neurotransmission in the mesocorticolimbic dopamine system, which could play a significant role in the enduring biochemical and behavioral effects of cocaine.