Direct proteasome binding and subsequent degradation of unspliced XBP-1 prevent its intracellular aggregation

The non-canonical splicing of XBP-1 mRNA is a hallmark of the mammalian unfolded protein response (UPR). The proteasomal degradation of unspliced XBP-1 (XBP-1u) facilitates the termination of the UPR. Thus, understanding the mechanism of XBP-1u degradation may allow control over UPR duration and intensity.We show that XBP-1u interacts with purified 20S proteasomes through its unstructured C-terminus, which leads to its degradation in a manner that autonomously opens the proteasome gate. In living cells, the C-terminus of XBP-1u accumulates in aggresome structures in the presence of proteasome inhibitors. We propose that direct proteasomal degradation of XBP-1u prevents its intracellular aggregation.

Structured summary

MINT-7302217: XBP1-u (uniprotkb:P17861-1) binds (MI:0407) to Proteasome subunit alpha 7.2 (uniprotkb:O14818) by pull down (MI:0096)MINT-7302148: Vimentin (uniprotkb:P08670) and XBP1-u (uniprotkb:P17861-1) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7302163: XBP1-u (uniprotkb:P17861-1) binds (MI:0407) to Proteasome subunit alpha 5 (uniprotkb:P28066) by pull down (MI:0096)MINT-7302186: XBP1-u (uniprotkb:P17861-1) binds (MI:0407) to Proteasome subunit alpha 6 (uniprotkb:P60900) by pull down (MI:0096)     

Factors affecting cell attachment, spreading, and growth on derivatized microcarriers. I. Establishment of working system and effect of the type of the amino-charged groups

A working system for studying the effects of factors involved in the chemical nature of microcarriers on cell attachment, spreading, and growth was established. The system is based on polyacrylamide beads, prepared by the emulsion polymerization technique. Sieved beads of desirable mean diameter were derivatized to generate controlled amounts of primary and tertiary amino groups. These microcarriers were used for the propagation of four different cell strains: BHK, MDCK, CEF, and MRC-5. It was found that BHK cells attach and spread significantly faster on primary amino-derivatized beads than those with tertiary amino groups, and at a lower degree of charging. Cell yields of MDCK cells (with pronounced epithelial morphology) propagated on primary amino-derivatized beads were higher than that obtained for the tertiary amino-derivatized microcarriers. On the other hand, CEF and MRC-5 cells (with pronounced fibroblast morphology) achieved higher cell yields on the tertiary amino-derivatized microcarriers.     

Charge delocalization in proton channels, II: the synthetic LS2 channel and proton selectivity

In this study, the minimalist synthetic LS2 channel is used as a prototype to examine the selectivity of protons over other cations. The free-energy profiles along the transport pathway of LS2 are calculated for three cation species: a realistic delocalized proton (including Grotthuss shuttling)--H(+), a classical (nonshuttling) hydronium--H(3)O(+), and a potassium cation--K(+). The overall barrier for K(+) is approximately twice as large as that for H(+), explaining the >100 times larger maximal ion conductance for the latter, in qualitative agreement with the experimental result. The profile for the classical hydronium is quantitatively intermediate between those of H(+) and K(+) and qualitatively more similar to that of H(+), for which the locations of the peaks are well correlated with the troughs of the pore radius profile. There is a strong correlation between the free-energy profiles and the very different characteristic hydration structures of the three cation species. This work suggests that the passage of various cations through ion channels cannot always be explained by simple electrostatic desolvation considerations.     

Coding of sweet,bitter, and umami tastes: different receptor cells sharing similar signaling pathways

Mammals can taste a wide repertoire of chemosensory stimuli. Two unrelated families of receptors (T1Rs and T2Rs) mediate responses to sweet, amino acids, and bitter compounds. Here, we demonstrate that knockouts of TRPM5, a taste TRP ion channel, or PLCbeta2, a phospholipase C selectively expressed in taste tissue, abolish sweet, amino acid, and bitter taste reception, but do not impact sour or salty tastes. Therefore, despite relying on different receptors, sweet, amino acid, and bitter transduction converge on common signaling molecules. Using PLCbeta2 taste-blind animals, we then examined a fundamental question in taste perception: how taste modalities are encoded at the cellular level. Mice engineered to rescue PLCbeta2 function exclusively in bitter-receptor expressing cells respond normally to bitter tastants but do not taste sweet or amino acid stimuli. Thus, bitter is encoded independently of sweet and amino acids, and taste receptor cells are not broadly tuned across these modalities.     

Disturbance of the rainforest has the potential to enhance egg parasitism of lepidopteran noctuid stemborers in Kisangani,DR Congo

Landscape context influences population dynamics of insects and impacts biological processes within communities. It was expected that anthropogenic disturbances of the rainforest landscape in DR Congo would lead to a decreased level of noctuid stemborer egg parasitism as a consequence of a decoupling between stemborers and their naturally occurring parasitoids through dispersal. To test this hypothesis, noctuid egg batches were collected in maize fields along an anthropogenic disturbance gradient to assess change in the rates of eggs parasitism and maize plant infestation with noctuid egg batches. Our results showed that, in contrast to what was initially expected, egg parasitism increased from less to highly disturbed landscape whereas maize infestation had an inverse tendency. Discovery efficiency and mean egg parasitism were 1.416 and 1.392 times higher, respectively, in the most than in the less disturbed landscape. The numbers of eggs and egg batches per 100 maize plants were 0.55 times and 0.532 times the value in the less disturbed landscape, suggesting a dilution of the stemborer population within a large habitat patch encompassing cultivated fields and the surrounding wild host plants. It was concluded that the presence of suitable host plants enhances noctuid stemborers egg parasitism in adjacent maize fields.     

PREDICT modeling and in-silico screening for G-protein coupled receptors

G-protein coupled receptors (GPCRs) are a major group of drug targets for which only one x-ray structure is known (the nondrugable rhodopsin), limiting the application of structure-based drug discovery to GPCRs. In this paper we present the details of PREDICT, a new algorithmic approach for modeling the 3D structure of GPCRs without relying on homology to rhodopsin. PREDICT, which focuses on the transmembrane domain of GPCRs, starts from the primary sequence of the receptor, simultaneously optimizing multiple 'decoy' conformations of the protein in order to find its most stable structure, culminating in a virtual receptor-ligand complex. In this paper we present a comprehensive analysis of three PREDICT models for the dopamine D2, neurokinin NK1, and neuropeptide Y Y1 receptors. A shorter discussion of the CCR3 receptor model is also included. All models were found to be in good agreement with a large body of experimental data. The quality of the PREDICT models, at least for drug discovery purposes, was evaluated by their successful utilization in in-silico screening. Virtual screening using all three PREDICT models yielded enrichment factors 9-fold to 44-fold better than random screening. Namely, the PREDICT models can be used to identify active small-molecule ligands embedded in large compound libraries with an efficiency comparable to that obtained using crystal structures for non-GPCR targets.     

Complexes of HLA-G protein on the cell surface are important for leukocyte Ig-like receptor-1 function

The nonclassical class I MHC molecule HLA-G is selectively expressed on extravillous cytotrophoblast cells at the maternal-fetal interface during pregnancy. HLA-G can inhibit the killing mediated by NK cells via interaction with the inhibitory NK cell receptor, leukocyte Ig-like receptor-1 (LIR-1). Comparison of the sequence of the HLA-G molecule to other class I MHC proteins revealed two unique cysteine residues located in positions 42 and 147. Mutating these cysteine residues resulted in a dramatic decrease in LIR-1 Ig binding. Accordingly, the mutated HLA-G transfectants were less effective in the inhibition of NK killing and RBL/LIR-1 induced serotonin release. Immunoprecipitation experiments demonstrated the involvement of the cysteine residues in the formation of HLA-G protein oligomers on the cell surface. The cysteine residue located at position 42 is shown to be critical for the expression of such complexes. These oligomers, unique among the class I MHC proteins, probably bind to LIR-1 with increased avidity, resulting in an enhanced inhibitory function of LIR-1 and an impaired killing function of NK cells.     

Inhaled nitric oxide increases surfactant protein gene expression in the intact lamb

Inhaled nitric oxide (iNO) is used to treat a number of disease processes. Although in vitro data suggest that nitric oxide (NO) alters surfactant protein gene expression, the effects in vivo have not been studied. The objective of this study was to evaluate the effects of iNO on surfactant protein (SP)-A, -B, and -C gene expression in the intact lamb. Thirteen 4-wk-old lambs were mechanically ventilated with 21% oxygen and received iNO at 40 ppm (n = 7) or vehicle gas (n = 6) for 24 h. Peripheral lung biopsies were obtained at 0, 12, and 24 h and analyzed for surfactant mRNA, protein, and total DNA content. Inhaled NO increased SP-A and SP-B mRNA content by 80% from 0 to 12 h and by 78 and 71%, respectively, from 0 to 24 h. There was an increase in SP-A and SP-B protein content by 45% from 0 to 12 h, and a decrease by 70 and 65%, respectively, from 0 to 24 h. DNA content was unchanged. The mechanisms and physiological effects of these findings warrant further investigation.     

Organogenesis in the vine cactus Selenicereus megalanthus using thidiazuron

Segments of distal and proximal parts of the cotyledons, hypocotyls and epicotyls of yellow pitaya [Selenicereus megalanthus (Cactaceae)], were cultured on Murashige and Skoog (MS) medium supplemented with thidiazuron (TDZ). The proximal part of the cotyledons was found to be the most responsive tissue, maximal bud regeneration being achieved in the presence of 200 M thidiazuron. However, TDZ does not change the pattern of regeneration competence, which remains restricted to the proximal part of the cotyledons. The best medium for shoot elongation was found to be MS lacking plant growth regulators. Rooting was induced on MS medium supplemented with 5.3 M naphthaleneacetic acid. Rooted plantlets were transferred to a soil medium and acclimatized successfully in a greenhouse.