Effect of subdomain interactions on methyl group dynamics in the hydrophobic core of villin headpiece protein

Thermostable villin headpiece protein (HP67) consists of the N‐terminal subdomain (residues 10–41) and the autonomously folding C‐terminal subdomain (residues 42–76) which pack against each other to form a structure with a unified hydrophobic core. The X‐ray structures of the isolated C‐terminal subdomain (HP36) and its counterpart in HP67 are very similar for the hydrophobic core residues. However, fine rearrangements of the free energy landscape are expected to occur because of the interactions between the two subdomains. We detect and characterize these changes by comparing the µs‐ms time scale dynamics of the methyl‐bearing side chains in isolated HP36 and in HP67. Specifically, we probe three hydrophobic side chains at the interface of the two subdomains (L42, V50, and L75) as well as at two residues far from the interface (L61 and L69). Solid‐state deuteron NMR techniques are combined with computational modeling for the detailed characterization of motional modes in terms of their kinetic and thermodynamic parameters. The effect of interdomain interactions on side chain dynamics is seen for all residues but L75. Thus, changes in dynamics because of subdomain interactions are not confined to the site of perturbation. One of the main results is a two‐ to threefold increase in the value of the activation energies for the rotameric mode of motions in HP67 compared with HP36. Detailed analysis of configurational entropies and heat capacities complement the kinetic view of the degree of the disorder in the folded state.     

Melatonin inhibits triple-negative breast cancer progression through the Lnc049808-FUNDC1 pathway

Melatonin has been reported to have tumor-suppressive effects via comprehensive molecular mechanisms, and long non-coding RNAs (lncRNAs) may participate in this process. However, the mechanism by which melatonin affects the function of lncRNAs in triple-negative breast cancer (TNBC), the most aggressive subtype of breast cancer, is still unknown. Therefore, we aimed to investigate the differentially expressed mRNAs and lncRNAs in melatonin-treated TNBC cells and the interaction mechanisms. Microarray analyses were performed to identify differentially expressed mRNAs and lncRNAs in TNBC cell lines after melatonin treatment. To explore the functions and underlying mechanisms of the mRNAs and lncRNAs candidates, a series of in vitro experiments were conducted, including CCK-8, Transwell, colony formation, luciferase reporter gene, and RNA immunoprecipitation (RIP) assays, and mouse xenograft models were established. We found that after melatonin treatment, FUNDC1 and lnc049808 downregulated in TNBC cell lines. Knockdown of FUNDC1 and lnc049808 inhibited TNBC cell proliferation, invasion, and metastasis. Moreover, lnc049808 and FUNDC1 acted as competing endogenous RNAs (ceRNAs) for binding to miR-101. These findings indicated that melatonin inhibited TNBC progression through the lnc049808-FUNDC1 pathway and melatonin could be used as a potential therapeutic agent for TNBC.Subject terms: Breast cancer, Non-coding RNAs     

Identification of blapsins A and B as potent small-molecule 14-3-3 inhibitors from the insect Blaps japanensis

In this study, we report three novel naturally occurring compounds, blapsins A (1) and B (2), and blapsamide (3) from the ethanol extract of the stink beetle, Blaps japanensis. The structures of these compounds were determined using spectroscopic methods. Compound 3 is a phenolic compound bearing a formamido group in the structure. Functional studies revealed that compounds 1 and 2 potently inhibited 14-3-3 protein-protein interactions (PPIs) with IC(50) values of 9.2 and 10.0 μM as determined by an ELISA assay, and 2.0 and 2.5 μM in an FP assay, respectively. These compounds represent the first example of natural small-molecule 14-3-3 inhibitors.     

Probe droplet arrays generated in the capillary for microarray analysis

Microarray technology is a useful tool for nucleic acid detection and has been widely used in biology and related research fields. However, the procedure is labor intensive and time consuming. Microfluidic chip-based microarrays save time with better performance, but the low spot density and probe number limit its applications. To develop high performance microarrays with high spot density within a microchannel, a method is reported here for preparing microarrays in a capillary by generating probe droplet arrays. The probes in droplets are immobilized onto the inner wall of the capillary to form a one-dimensional probe array, and then a sample solution is introduced to hybridize with the probe array. The effect of the capillary's inner diameter was evaluated to realize a high-density probe array. The processes of array generation and probe immobilization were studied to avoid possible cross contamination. The background from probe immobilization during the array generation and incubation was quantified to assure sensitivity. Multiple sample detection was also demonstrated within one capillary. The capillary based microarray assay had high spot density, easy fabrication, fast detection, high sensitivity and multiple sample capacity.     

Abundance, marker development and genetic mapping of microsatellites from unigenes in Brassica napus

Rapeseed (Brassica napus) is the second most important oil crop in the world after soybean. The repertoire of simple sequence repeat (SSR) markers for rapeseed is limited and warrants a search for a larger number of polymorphic SSRs for germplasm characterization and breeding applications. In this study, a total of 5,310 SSR-containing unigenes were identified from a set of 46,038 B. napus unigenes with an average density of one SSR every 5.75?kb. A set of 1,000 expressed sequence tag (EST)-SSR markers with repeat length ??18?bp were developed and tested for their ability to detect polymorphism among a panel of six rapeseed varieties. Of these SSR markers, 776 markers detected clear amplification products, and 511 displayed polymorphisms among the six varieties. Of these polymorphic markers, 195 EST-SSR markers, corresponding to 233 loci, were integrated into an existing B. napus linkage map. These EST-SSRs were randomly distributed on the 19 linkage groups of B. napus. Of the mapped loci, 166 showed significant homology to Arabidopsis genes. Based on the homology, 44 conserved syntenic blocks were identified between B. napus and Arabidopsis genomes. Most of the syntenic blocks were consistent with the duplication and rearrangement events identified previously. In addition, we also identified three previously unreported blocks in B. napus. A subset of 40 SSRs was used to assess genetic diversity in a collection of 192 rapeseed accessions. The polymorphism information content of these markers ranged from 0.0357 to 0.6753 with an average value of 0.3373. These results indicated that the EST-SSR markers developed in this study are useful for genetic mapping, molecular marker-assisted selection and comparative genomics.     

湛江高桥红树林和盐沼湿地的大型底栖动物次级生产力

为了比较湛江高桥潮间带不同植物生境的大型底栖动物次级生产力,根据2010年4个季度湛江高桥潮间带生境的大型底栖动物数据,运用Brey经验公式计算不同植物生境的大型底栖动物次级生产力.结果表明:湛江高桥红树林和盐沼湿地不同生境大型底栖动物平均次级生产力为11.77 g AFDM·m-2·a-1.其中,无瓣海桑生境次级生产力最高,为18.16 g AFDM·m-2·a-1,其次是桐花树、盐地鼠尾粟和木榄生境,分别为17.67、8.34和2.92 g AFDM·m-2·a-1.在4种生境中,木榄生境的年生产力/年均生物量(P/B)最高,为2.38,其次是无瓣海桑、盐地鼠尾粟和桐花树生境,分别为1.23、0.99和0.48.湛江高桥潮间带不同植物生境大型底栖动物次级生产力和P/B值的差异主要与总有机碳含量、食物类型和动物个体大小有关.