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101.
Tai‐Wen Lin Chi‐Chih Chen Shu‐Mei Wu Yu‐Ching Chang Yi‐Chuan Li Yu‐Wang Su Chwan‐Deng Hsiao Hsin‐Yang Chang 《The Plant journal : for cell and molecular biology》2019,99(1):128-143
In mammals and yeast, tail‐anchored (TA) membrane proteins destined for the post‐translational pathway are safely delivered to the endoplasmic reticulum (ER) membrane by a well‐known targeting factor, TRC40/Get3. In contrast, the underlying mechanism for translocation of TA proteins in plants remains obscure. How this unique eukaryotic membrane‐trafficking system correctly distinguishes different subsets of TA proteins destined for various organelles, including mitochondria, chloroplasts and the ER, is a key question of long standing. Here, we present crystal structures of algal ArsA1 (the Get3 homolog) in a distinct nucleotide‐free open state and bound to adenylyl‐imidodiphosphate. This approximately 80‐kDa protein possesses a monomeric architecture, with two ATPase domains in a single polypeptide chain. It is capable of binding chloroplast (TOC34 and TOC159) and mitochondrial (TOM7) TA proteins based on features of its transmembrane domain as well as the regions immediately before and after the transmembrane domain. Several helices located above the TA‐binding groove comprise the interlocking hook‐like motif implicated by mutational analyses in TA substrate recognition. Our data provide insights into the molecular basis of the highly specific selectivity of interactions of algal ArsA1 with the correct sets of TA substrates before membrane targeting in plant cells. 相似文献
102.
Shih‐Fan Chan Wei‐Kai Shih An‐Yu Chang Sheng‐Feng Shen I‐Ching Chen 《Ecology letters》2019,22(10):1668-1679
How abiotic and biotic factors constrain distribution limits at the harsh and benign edges of species ranges is hotly debated, partly because macroecological experiments testing the proximate causes of distribution limits are scarce. It has long been recognized – at least since Darwin’s On the Origin of Species – that a harsh climate strengthens competition and thus sets species range limits. Using thorough field manipulations along a large elevation gradient, we show the mechanisms by which temperature determines competition type, resulting in a transition from interference to exploitative competition from the lower to the upper elevation limits in burying beetles (Nicrophorus nepalensis). This transition is an example of Darwin’s classic hypothesis that benign climates favor direct competition for highly accessible resources while harsh climates result in competition through resources of high rivalry. We propose that identifying the properties of these key resources will provide a more predictive framework to understand the interplay between biotic and abiotic factors in determining geographic range limits. 相似文献
103.
Takashi Yazawa Yoshitaka Imamichi Koh‐ichi Yuhki Junsuke Uwada Daisuke Mikami Masayuki Shimada Kaoru Miyamoto Takeshi Kitano Satoru Takahashi Toshio Sekiguchi Nobuo Suzuki Md. Rafiqul Islam Khan Fumitaka Ushikubi Akihiro Umezawa Takanobu Taniguchi 《Molecular reproduction and development》2019,86(7):786-797
104.
Liu P Sudhaharan T Koh RM Hwang LC Ahmed S Maruyama IN Wohland T 《Biophysical journal》2007,93(2):684-698
Single wavelength fluorescence cross-correlation spectroscopy (SW-FCCS), introduced to study biomolecular interactions, has recently been reported to monitor enzyme activity by using a newly developed fluorescent protein variant together with cyan fluorescent protein. Here, for the first time to our knowledge, SW-FCCS is applied to detect interactions between membrane receptors in vivo by using the widely used enhanced green fluorescent protein and monomeric red fluorescent protein. The biological system studied here is the epidermal growth factor/ErbB receptor family, which plays pivotal roles in the development of organisms ranging from worms to humans. It is widely thought that a ligand binds to the monomeric form of the receptor and induces its dimeric form for activation. By using SW-FCCS and F?rster resonance energy transfer, we show that the epidermal growth factor receptor and ErbB2 have preformed homo- and heterodimeric structures on the cell surface and quantitation of dimer fractions is performed by SW-FCCS. These receptors are major targets of anti-cancer drug development, and the receptors' homo- and heterodimeric structures are relevant for such developments. 相似文献
105.
Biochemical characterization of distinct regions of SPEC molecules and their role in phagocytosis 总被引:1,自引:0,他引:1
Cdc42 signaling pathways play important roles in immune cell polarization and cytoskeletal changes. Although the small Cdc42-binding proteins SPEC1 and SPEC2 play a role in F-actin accumulation in activated T lymphocytes, little is known about their precise activities in other cell types. Here, we mapped the Cdc42-binding activity of SPEC1 to the CRIB sequence and a downstream alpha helical region. Biochemical studies revealed that SPEC1 did not interact with a Rac1 switch-of-function mutant capable of inducing Cdc42-like filopodia, potentially eliminating a role for SPECs in this process. A phosphoinositide-binding region was identified within a basic region N-terminal to the CRIB sequence of SPEC1. Using an anti-SPEC2 antibody, we found that endogenous SPEC2 colocalized with Cdc42 at the phagocytic cup of macrophages internalizing zymosan A particles prior to significant F-actin accumulation. Overexpression studies of the related SPEC1 protein induced marked macrophage contraction and prevented particle binding and phagocytosis. Although a Cdc42-binding mutant of SPEC1 still caused macrophage contraction, mutations within the N-terminal cysteines and phosphoinositide-binding region reversed macrophage contraction but still resulted in impaired phagocytosis. These results identify three distinct structural and functional regions within SPECs and demonstrate their likely role in early contractile events in phagocytosis. 相似文献
106.
107.
The range of BRCA1/BRCA2 gene mutations is diverse and the mechanism accounting for this heterogeneity is obscure. To gain insight into the endogenous mutational mechanisms involved, we evaluated the association of specific sequences (i.e. CpG/CpNpG motifs, homonucleotides, short repeats) and mutations within the genes. We classified 1337 published mutations in BRCA1 (1765 BRCA2 mutations) for each specific sequence, and employed computer simulation combined with mathematical calculations to estimate the true underlying tendency of mutation occurrence. Interestingly, we found no mutational bias to homonucleotides and repeats in deletions/insertions and substitutions but striking bias to CpG/CpNpG in substitutions in both genes. This suggests that methylation-dependent DNA alterations would be a major mechanism for mutagenesis. 相似文献
108.
Human TRBP and PACT directly interact with each other and associate with dicer to facilitate the production of small interfering RNA 总被引:5,自引:0,他引:5
Mammalian Dicer interacts with double-stranded RNA-binding protein TRBP or PACT to mediate RNA interference and micro-RNA processing. TRBP and PACT are structurally related but exert opposite regulatory activities on PKR. It is not understood whether TRBP and PACT are simultaneously required for Dicer. Here we show that TRBP directly interacts with PACT in vitro and in mammalian cells. TRBP and PACT form a triple complex with Dicer and facilitate the production of small interfering RNA (siRNA) by Dicer. Knockdown of both TRBP and PACT in cultured cells leads to significant inhibition of gene silencing mediated by short hairpin RNA but not by siRNA, suggesting that TRBP and PACT function primarily at the step of siRNA production. Taken together, these findings indicate that human TRBP and PACT directly interact with each other and associate with Dicer to stimulate the cleavage of double-stranded or short hairpin RNA to siRNA. Our work significantly alters the current model for the assembly and function of the Dicer-containing complex that generates siRNA and micro-RNA in human. 相似文献
109.
Mesenchymal stromal cells (MSCs) have proven useful for cell and immune therapy, but the molecular constituents responsible for their functionalities, in particular, those on the plasma membrane, remain largely unknown. Here we employed both gel and nongel based MS to analyze human MSCs' membrane proteome before and after adipogenesis. 2-DE of cells that were pretreated with membrane impermeable fluorescent dyes revealed that both the whole cell proteome and the cell surface subproteome were independent of donors. LC coupled with tandem MS analysis of the plasma membrane-containing fraction allowed us to identify 707 proteins, approximately half of which could be annotated as membrane-related proteins. Of particular interest was a subset of ectodomain-containing membrane-bound proteins that encompass most known surface markers for MSCs, but also contain a multitude of solute carriers and ATPases. Upon adipogenic differentiation, this proteomic profile was amended to include several proteins involved in lipid metabolism and trafficking, at the expense of, most noticeably, ectoenzymes. Our results here provide not only a basis for future studies of MSC-specific molecular mechanisms, but also a molecular inventory for the development of antibody-based cell isolation and identification procedures. 相似文献
110.
Differential recovery of membrane proteins after extraction by aqueous methanol and trifluoroethanol
Zhang H Lin Q Ponnusamy S Kothandaraman N Lim TK Zhao C Kit HS Arijit B Rauff M Hew CL Chung MC Joshi SB Choolani M 《Proteomics》2007,7(10):1654-1663
Cell membrane proteome analysis is limited by inherent membrane hydrophobicity. Conventional membrane protein extraction techniques use detergents, chaotropes and organic acids that require sample clean-up or pH adjustment, and are associated with significant sample loss. We extracted membrane proteins from red blood cells (RBCs) using methanol (MeOH), trifluoroethanol (TFE) and urea, and identified membrane proteins using 2-D LC coupled with MALDI-TOF/TOF-MS. We show that organic solvents MeOH- and TFE-based methods have membrane protein analysis efficiencies comparable to urea, and are complementary for the recovery of both hydrophilic and hydrophobic peptides. The mean grand average of hydropathicity (GRAVY) value of identified peptides from the TFE-based method (-0.107) was significantly higher than that of the MeOH-based method (-0.465) (p<0.001). Sequential and adjunctive use of the organic solvents MeOH and TFE increases the number of proteins identified, and the confidence of their identification. We show that this strategy is effective for shotgun membrane proteome analysis. 相似文献