Protein degradation by the ubiquitin-proteasome pathway plays important roles in synaptic plasticity, but the molecular mechanisms by which proteolysis regulates synaptic strength are not well understood. We investigated the role of the proteasome in hippocampal late-phase long-term potentiation (L-LTP), a model for enduring synaptic plasticity. We show here that inhibition of the proteasome enhances the induction of L-LTP, but inhibits its maintenance. Proteasome inhibitor-mediated enhancement of the early part of L-LTP requires activation of NMDA receptors and the cAMP-dependent protein kinase. Augmentation of L-LTP induction by proteasome inhibition is blocked by a protein synthesis inhibitor anisomycin and is sensitive to the drug rapamycin. Our findings indicate that proteasome inhibition increases the induction of L-LTP by stabilizing locally translated proteins in dendrites. In addition, our data show that inhibition of the proteasome blocks transcription of brain-derived neurotrophic factor (BDNF), which is a cAMP-responsive element-binding protein (CREB)-inducible gene. Furthermore, our results demonstrate that the proteasome inhibitors block degradation of ATF4, a CREB repressor. Thus, proteasome inhibition appears to hinder CREB-mediated transcription. Our results indicate that blockade of proteasome activity obstructs the maintenance of L-LTP by interfering with transcription as well as translation required to sustain L-LTP. Thus, proteasome-mediated proteolysis has different roles during the induction and the maintenance of L-LTP.Several decades of investigation on synaptic plasticity underlying learning and memory have unearthed significant roles for posttranslational modifications such as phosphorylation in shortterm plasticity and for gene expression in causing long-lasting changes in synaptic strength (O'Dell et al. 1991;Kandel 2001;Deisseroth et al. 2003;Davis 2005;Mansuy and Shenolikar 2006). In recent years, a role for proteolysis by the ubiquitinproteasome pathway in synaptic plasticity has been discovered (Hegde and DiAntonio 2002;Murphey and Godenschwege 2002;Hegde 2004). In this pathway, proteins to be degraded are marked by covalent linkage to a small protein called ubiquitin for degradation by a proteolytic complex, the proteasome.Previous studies on long-term facilitation in Aplysia, which underlies a simple form of long-term memory, revealed a role for ubiquitin-proteasome-mediated degradation of the inhibitory regulatory subunit of cAMP-dependent protein kinase (PKA) (Hegde et al. 1993). Moreover, an enzyme of the ubiquitinproteasome pathway called ubiquitin C-terminal hydrolase (Apuch), which interacts with the proteasome, was found to be induced by serotonin (5-HT), the neurotransmitter that induces long-term facilitation. Ap-uch was found to be critical for induction of long-term facilitation (Hegde et al. 1997). Degradation of regulatory subunit of PKA suggested that the role of proteolysis is to remove inhibitory constraints on long-term synaptic plasticity (Hegde et al. 1997;Ch...

Alternative splicing (AS) of pre-mRNAs promotes transcriptome and proteome diversity and plays important roles in a wide range of biological processes. However, the role of AS in maintaining mineral nutrient homeostasis in plants is largely unknown. To clarify this role, we obtained whole transcriptome RNA sequencing data from rice (Oryza sativa) roots grown in the presence or absence of several mineral nutrients (Fe, Zn, Cu, Mn, and P). Our systematic analysis revealed 13,291 alternatively spliced genes, representing ∼53.3% of the multiexon genes in the rice genome. As the overlap between differentially expressed genes and differentially alternatively spliced genes is small, a molecular understanding of the plant's response to mineral deficiency is limited by analyzing differentially expressed genes alone. We found that the targets of AS are highly nutrient-specific. To verify the role of AS in mineral nutrition, we characterized mutants in genes encoding Ser/Arg (SR) proteins that function in AS. We identified several SR proteins as critical regulators of Zn, Mn, and P nutrition and showed that three SR protein-encoding genes regulate P uptake and remobilization between leaves and shoots of rice, demonstrating that AS has a key role in regulating mineral nutrient homeostasis in rice.

Mesenchymal stem cells (MSCs), which are poorly immunogenic and have potent immunosuppressive activities, have emerged as a promising candidate for cellular therapeutics for the treatment of disorders caused by abnormal immune responses. In this study we investigated whether human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) could ameliorate colitis in a trinitrobenzene sulfonic acid (TNBS)-induced colitis model. TNBS-treated colitic mice were infused with hUC-MSCs or vehicle control. The mice were sacrificed on day 1, 3, and 5 after infusion, and their clinical and pathological conditions were evaluated by body weight, colon length, and histological analysis. The expression levels of proinflammatory cytokine proteins in colon were examined by ELISA. The homing of hUC-MSCs was studied by live in vivo imaging and immunofluorescent microscopy. hUC-MSCs were found to migrate to the inflamed colon and effectively treated the colitic mice with improved clinical and pathological signs. The levels of IL-17 and IL-23 as well as IFN-γ and IL-6 were significantly lower in the colon tissues of the hUC-MSC-treated mice in comparison with the vehicle-treated mice. Coculture experiments showed that hUC-MSCs not only could inhibit IFN-γ expression but also significantly inhibit IL-17 production by lamina propria mononuclear cells (LPMCs) or splenocytes of the colitic mice or by those isolated from normal animals and stimulated with IL-23. Systemically infused hUC-MSCs could home to the inflamed colon and effectively ameliorate colitis. In addition to the known suppressive effects on Th1-type immune responses, hUC-MSC-mediated modulation of IL-23/IL-17 regulated inflammatory reactions also plays an important role in the amelioration of colitis.

Proteolysis by the ubiquitin-proteasome pathway appears to have a complex role in synaptic plasticity, but its various functions remain to be elucidated. Using late phase long-term potentiation (L-LTP) in the hippocampus of the mouse as a model for long-term synaptic plasticity, we previously showed that inhibition of the proteasome enhances induction but blocks maintenance of L-LTP. In this study, we investigated the possible mechanisms by which proteasome inhibition has opposite effects on L-LTP induction and maintenance. Our results show that inhibiting phosphatidyl inositol-3 kinase or blocking the interaction between eukaryotic initiation factors 4E (eIF4E) and 4G (eIF4G) reduces the enhancement of L-LTP induction brought about by proteasome inhibition suggesting interplay between proteolysis and the signaling pathway mediated by mammalian target of rapamycin (mTOR). Also, proteasome inhibition leads to accumulation of translational activators in the mTOR pathway such as eIF4E and eukaryotic elongation factor 1A (eEF1A) early during L-LTP causing increased induction. Furthermore, inhibition of the proteasome causes a buildup of translational repressors, such as polyadenylate-binding protein interacting protein 2 (Paip2) and eukaryotic initiation factor 4E-binding protein 2 (4E-BP2), during late stages of L-LTP contributing to the blockade of L-LTP maintenance. Thus, the proteasome plays a critical role in regulating protein synthesis during L-LTP by tightly controlling translation. Our results provide novel mechanistic insights into the interplay between protein degradation and protein synthesis in long-term synaptic plasticity.

This study aimed to examine HIF-2α, TWIST, and CXCR4 expression in papillary thyroid carcinoma (PTC) and assesses the association of their expression with clinicopathological indicators. HIF-2α, TWIST, and CXCR4 protein expression in 129 PTCs, 61 nodular hyperplasia, and 118 normal thyroid tissue specimens was analyzed using immunohistochemistry. The protein expression levels of these three molecules were upregulated in PTCs. High protein expression of HIF-2α, TWIST, and CXCR4 was significantly correlated with lymph node metastasis (LNM) (P < 0.001). Furthermore, HIF-2α, TWIST, and CXCR4 protein expression was correlated with one another. Concomitant high expression of these molecules had stronger correlation with LNM than did each alone (P = 0.032 for HIF-2α/TWIST, P < 0.001 for HIF-2α/CXCR4, P = 0.018 for TWIST/CXCR4, and P < 0.001 for HIF-2α/TWIST/CXCR4). Additionally, HIF-2α, TWIST, and CXCR4 mRNA expression were assessed in 30 PTCs, 10 nodular hyperplasia, and 10 normal thyroid tissue specimens using real-time RT-PCR. TWIST and CXCR4 mRNA expression levels were up-regulated in PTCs, and high mRNA expression of TWIST and CXCR4 was significantly correlated with LNM (P = 0.005 and P = 0.010, resp.). These results demonstrated that the evaluation of HIF-2α, TWIST, and CXCR4 expression in PTC may be useful in predicting the risk of LNM.

In contrast to hematopoietic stem cells, there is still a lack of definitive cell markers for specific isolation and identification of mesenchymal stem cells (MSCs). Thus a homogenous population of MSCs is only obtained after several passages, when multilineage potential or other distinctive features of very early progenitors may be already somewhat compromised. Recently a novel surface marker the neural ganglioside GD2 has been reported to distinguish MSCs from all other cells within marrow. Here, we found that MSCs derived from umbilical cord (UC-MSCs) also expressed this marker at early-passages. More importantly, UC-MSCs were the only cells within umbilical cord expressing this marker. Compared to unsorted cells, GD2⁺-sorted cells not only possessed much higher clonogenicity and proliferation capacity but also had significantly stronger multi-differentiation potentials. Flow cytometric analysis revealed that GD2⁺-sorted cells showed increased expression of SSEA-4, Oct-4, Sox-2 and Nanog, the typical markers expressed in embryonic stem cells, in comparison to unsorted or GD2-negative MSCs. Take together, our data demonstrate that the cells selected by GD2 are a subpopulation of MSCs with feature of primitive precursor cells and provide evidence that GD2 can be a cell surface marker suitable for the isolation and purification of UC-MSCs in early-passage culture.

Thioredoxin reductase 1 (TrxR1) is a pivotal intracellular redox sensor and antioxidant enzyme. On the other hand, overexpression of TrxR1 is closely correlated with the initiation of various tumors including breast cancer, though the detailed mechanism remains unclear. Here we investigated the role of TrxR1 in dysplastic transformation of human breast epithelial cell line MCF-10A induced by chronic oxidative stress. Not surprisingly, sustained exposure to H2O2 significantly augmented the expression and activity of TrxR1 in MCF-10A cells. The dysplastically transformed MCF-10A (MCF-10AT) cells undergoing 8-week H2O2 treatment exhibited a certain degree of malignancy in tumorigenicity evaluation. Moreover, TrxR1 inhibitor ethaselen (BBSKE) could partially reverse some malignant phenotypes including epithelial to mesenchymal transition (EMT) of MCF-10AT as well as MCF-7 cells. Collectively, our results supported the considerable involvement of TrxR1 in the onset of breast cancer and BBSKE may be a promising agent against breast cancer.

MicroRNAs (miRNAs) are a class of non-coding, small RNAs recognized as important regulators of gene expression. Although plant miRNAs have been extensively studied in model systems, less is known in other plants with limited genome sequence data, including Paeonia ostii. In this work, we used high-throughput sequencing to identify conserved and nonconserved miRNAs and other short RNAs in Paeonia ostii under control and copper stressed condition. 102 previously known plant miRNAs were identified and classified into 89 families according to their gene sequence identity. Some miRNAs were highly conserved in the plant kingdom suggesting that these miRNA play important and conserved roles. Combined our transcriptome sequencing data of Paeonia ostii under same conditions, 34 novel potential miRNAs were identified. The potential targets of the identified known and novel miRNAs were also predicted based on sequence homology search. Comparing the two libraries, it was observed that 12 conserved miRNAs and 18 novel miRNAs showed significantly changes in response to copper stress. Some of the new identified potential miRNAs might be involved in Paeonia ostii-specific regulating mechanisms under copper stress. These results provide a framework for further analysis of miRNAs and their role in regulating Paeonia ostii response to copper stress.