Mitochondrial haplotypes may modulate the phenotypic manifestation of the LHON-associated m.14484T>C (MT-ND6) mutation in Chinese families

Mitochondrial m.14484T>C (MT-ND6) mutation has been associated with Leber's hereditary optic neuropathy. Previous investigations revealed that the m.14484T>C mutation is a primary factor underlying the development of optic neuropathy but is not sufficient to produce a clinical phenotype. However, mitochondrial haplogroups have been proposed to modulate the phenotypic manifestation of the m.14484T>C mutation. Here, we performed the clinical, genetic evaluation and complete mitochondrial genome sequence analysis of 41 Han Chinese pedigrees carrying the m.14484T>C mutation. These families exhibited a wide range of penetrances and expressivities of optic neuropathy. The average ratio between affected male/female matrilineal relatives from 41 families was 2:1. The penetrance of optic neuropathy in these Chinese pedigrees ranged from 5.6% to 100%, with the average of 23.8%. Furthermore, the age-of-onset for optic neuropathy varied from 4 to 44 years, with the average of 19.3 years. Sequence analysis of their mitochondrial genomes identified distinct sets of polymorphisms belonging to ten Eastern Asian haplogroups, indicating that the m.14484T>C mutation occurred through recurrent origins and founder events. We showed that mitochondrial haplogroups M9, M10 and N9 increased the penetrance of optic neuropathy in these Chinese families. In particular, these mitochondrial haplogroup specific variants: m.3394T>C (MT-ND1), m.14502T>C (MT-ND4) and m.14693A>G (MT-TE) enhanced the penetrance of visual loss in these Chinese families. These data provided the direct evidence that mitochondrial modifiers modulate the variable penetrance and expressivity of optic neuropathy among Chinese pedigrees carrying the m.14484T>C mutation.     

Formation of the death domain complex between FADD and RIP1 proteins in vitro

Fas-associated death domain (FADD) protein is an adapter molecule that bridges the interactions between membrane death receptors and initiator caspases. The death receptors contain an intracellular death domain (DD) which is essential to the transduction of the apoptotic signal. The kinase receptor-interacting protein 1 (RIP1) is crucial to programmed necrosis. The cell type interplay between FADD and RIP1, which mediates both necrosis and NF-κB activation, has been evaluated in other studies, but the mechanism of the interaction of the FADD and RIP1 proteins remain poorly understood. Here, we provided evidence indicating that the DD of human FADD binds to the DD of RIP1 in vitro. We developed a molecular docking model using homology modeling based on the structures of FADD and RIP1. In addition, we found that two structure-based mutants (G109A and R114A) of the FADD DD were able to bind to the RIP1 DD, and two mutations (Q169A and N171A) of FADD DD and four mutations (G595, K596, E620, and D622) of RIP1 DD disrupted the FADD–RIP1 interaction. Six mutations (Q169A, N171A, G595, K596, E620, and D622) lowered the stability of the FADD–RIP1 complex and induced aggregation that structurally destabilized the complex, thus disrupting the interaction.     

Complexes of Neutralizing and Non-Neutralizing Affinity Matured Fabs with a Mimetic of the Internal Trimeric Coiled-Coil of HIV-1 gp41

A series of mini-antibodies (monovalent and bivalent Fabs) targeting the conserved internal trimeric coiled-coil of the N-heptad repeat (N-HR) of HIV-1 gp41 has been previously constructed and reported. Crystal structures of two closely related monovalent Fabs, one (Fab 8066) broadly neutralizing across a wide panel of HIV-1 subtype B and C viruses, and the other (Fab 8062) non-neutralizing, representing the extremes of this series, were previously solved as complexes with 5-Helix, a gp41 pre-hairpin intermediate mimetic. Binding of these Fabs to covalently stabilized chimeric trimers of N-peptides of HIV-1 gp41 (named (CCIZN36)₃ or 3-H) has now been investigated using X-ray crystallography, cryo-electron microscopy, and a variety of biophysical methods. Crystal structures of the complexes between 3-H and Fab 8066 and Fab 8062 were determined at 2.8 and 3.0 Å resolution, respectively. Although the structures of the complexes with the neutralizing Fab 8066 and its non-neutralizing counterpart Fab 8062 were generally similar, small differences between them could be correlated with the biological properties of these antibodies. The conformations of the corresponding CDRs of each antibody in the complexes with 3-H and 5-Helix are very similar. The adaptation to a different target upon complex formation is predominantly achieved by changes in the structure of the trimer of N-HR helices, as well as by adjustment of the orientation of the Fab molecule relative to the N-HR in the complex, via rigid-body movement. The structural data presented here indicate that binding of three Fabs 8062 with high affinity requires more significant changes in the structure of the N-HR trimer compared to binding of Fab 8066. A comparative analysis of the structures of Fabs complexed to different gp41 intermediate mimetics allows further evaluation of biological relevance for generation of neutralizing antibodies, as well as provides novel structural insights into immunogen design.     

高通量测序分析重庆地区茅苍术根际丛枝菌根真菌多样性

【背景】丛枝菌根真菌(arbuscular mycorrhizal fungi,AMF)是土壤微生物区系中分布最广泛的一类菌根真菌,能与地球上90%的维管植物形成共生体,可通过调节植物的生理代谢过程增强植物的抗逆性。【目的】揭示重庆地区中药材茅苍术根际土壤中AMF的结构与组成,解析土壤因子对AMF类群的影响。【方法】以重庆地区茅苍术主产地彭水县、秀山县、石柱县、南川区和酉阳县2-3年生茅苍术根际土壤为材料,利用Illumina MiSeq 2500测序平台进行真菌扩增子测序,分析不同地点土壤的茅苍术根际AMF类群组成和多样性的差异。【结果】茅苍术的菌根侵染率均在50%以上,每10g风干土壤中孢子含量在50个以上,最高达到144个。根际土壤共检测到球囊菌门Glomeromycota的3纲4目8科9属AMF,包括Glomus、Claroideoglomus、Gigaspora、Paraglomus、Archaeospora、Ambispora、Acaulospora、Diversispora和Scutellospora,其中前6属为5个区县土样共有。球囊霉属(Glomus)相对丰度最高,达67%,为所有地区茅苍术根际样本中的优势类群。冗余度分析(redundancy analysis,RDA)表明,土壤pH对AMF群落组成影响最大。pH、有机质、碱解氮、速效钾与Shannon指数呈正相关,有效磷与之呈负相关;各土壤因子与Simpson指数的相关性则相反。5个土壤因子均与丰度(Chao1)指数呈负相关。另外,pH、有机质与均匀度(ACE)指数呈正相关;碱解氮、速效钾、有效磷与之呈负相关。【结论】茅苍术根际土壤中AMF资源丰富,土壤因子对AMF群落组成和丰度影响显著,是导致AMF群落结构地理分布格局差异的重要原因之一。     

Effects of different 2A peptides on transgene expression mediated by tricistronic vectors in transfected CHO cells

Molecular Biology Reports - Multicistronic vectors can increase transgene expression and decrease the imbalance of gene expression in the Chinese hamster ovary (CHO) cell expression system. Small,...     

Long Noncoding RNA SNHG5 Knockdown Alleviates Neuropathic Pain by Targeting the miR-154-5p/CXCL13 Axis

Neurochemical Research - Neuropathic pain is an unneglectable pain condition with limited treatment options owing to its enigmatic underlying mechanisms. Long noncoding RNA small nucleolar RNA host...     

SARNP,a participant in mRNA splicing and export,negatively regulates E-cadherin expression via interaction with pinin

Triple-negative breast cancer (TNBC) is associated with a high mortality rate, which is related to the insufficient number of appropriate biomarkers and targets. Therefore, there is an urgent need to discover appropriate biomarkers and targets for TNBC. SARNP (Hcc-1 and CIP29) is highly expressed in several cancers. It binds to UAP56, an RNA helicase component of the TREX complex in messenger RNA (mRNA) splicing and export. However, the role of SARNP in mRNA splicing and export and in the progression of breast cancer, especially of TNBC, remains unknown. Therefore, we examined the role of SARNP in mRNA splicing and export and progression of TNBC. We confirmed that SARNP binds to UAP56 and Aly and that SARNP overexpression enhances mRNA splicing, whereas its knockdown suppressed mRNA export. The SARNP overexpression induced the proliferation of MCF7 cells, whereas its knockdown induced E-cadherin expression and downregulated vimentin and N-cadherin expressions in SK-BR-3 and MDA-MB-231 cells. SARNP downregulates E-cadherin expression by interaction with pinin. Mice injected with MDA-MB-231_shSARNP cells exhibited a significant reduction in tumor growth and lung metastasis compared with those injected with MDA-MB-231_shCon cells in vivo. These findings suggested that SARNP is involved in mRNA splicing and export. SARNP maintains mesenchymal phenotype by escaping from inhibitory interaction with pinin leading to the downregulation of E-cadherin expression.     

Reprogramming of sugar transport pathways in Escherichia coli using a permeabilized SecY protein-translocation channel

In the initial step of sugar metabolism, sugar-specific transporters play a decisive role in the passage of sugars through plasma membranes into cytoplasm. The SecY complex (SecYEG) in bacteria forms a membrane channel responsible for protein translocation. The present work shows that permeabilized SecY channels can be used as nonspecific sugar transporters in Escherichia coli. SecY with the plug domain deleted allowed the passage of glucose, fructose, mannose, xylose, and arabinose, and, with additional pore-ring mutations, facilitated lactose transport, indicating that sugar passage via permeabilized SecY was independent of sugar stereospecificity. The engineered E. coli showed rapid growth on a wide spectrum of monosaccharides and benefited from the elimination of transport saturation, improvement in sugar tolerance, reduction in competitive inhibition, and prevention of carbon catabolite repression, which are usually encountered with native sugar uptake systems. The SecY channel is widespread in prokaryotes, so other bacteria may also be engineered to utilize this system for sugar uptake. The SecY channel thus provides a unique sugar passageway for future development of robust cell factories for biotechnological applications.