Adapted formaldehyde gradient cross-linking protocol implicates human eIF3d and eIF3c, k and l subunits in the 43S and 48S pre-initiation complex assembly, respectively

One of the key roles of the 12-subunit eukaryotic translation initiation factor 3 (eIF3) is to promote the formation of the 43S and 48S pre-initiation complexes (PICs). However, particular contributions of its individual subunits to these two critical initiation reactions remained obscure. Here, we adapted formaldehyde gradient cross-linking protocol to translation studies and investigated the efficiency of the 43S and 48S PIC assembly in knockdowns of individual subunits of human eIF3 known to produce various partial subcomplexes. We revealed that eIF3d constitutes an important intermolecular bridge between eIF3 and the 40S subunit as its elimination from the eIF3 holocomplex severely compromised the 43S PIC assembly. Similarly, subunits eIF3a, c and e were found to represent an important binding force driving eIF3 binding to the 40S subunit. In addition, we demonstrated that eIF3c, and eIF3k and l subunits alter the efficiency of mRNA recruitment to 43S PICs in an opposite manner. Whereas the eIF3c knockdown reduces it, downregulation of eIF3k or eIF3l increases mRNA recruitment, suggesting that the latter subunits possess a regulatory potential. Altogether this study provides new insights into the role of human eIF3 in the initial assembly steps of the translational machinery.

A Theoretical Model for Hydraulic Fracturing through Two Symmetric Radial Perforations Emanating from A Borehole

The production enhancement of oil, gas, or geothermal reservoirs through hydraulic fracturing requires an in-depth study on the fracture initiation and propagation from the borehole. According to the linear elastic fracture mechanics, a theoretical model is developed to calculate the stress intensity factors of two symmetric radial cracks emanating from a pressurized borehole. The maximum tangential stress criterion under the mix-mode condition is developed to investigate the hydraulic fracture initiation. The critical water pressure and critical initiation angle predicted by the theoretical model match closely the experimental results reported in the literature. The influence of the stress anisotropy coefficient, the perforation angle and length, the borehole radius, the ratio between the water pressures in the fracture and the borehole, and Biot’s coefficient are investigated. Moreover, the effects of the injected high water pressure (i.e., larger than the critical water pressure) on the fracture initiation angle are studied to further understand the characteristics of hydraulic fracture initiation. The results indicate that the perforation angle and length, the borehole radius, and the stress anisotropy coefficient have a relatively strong influence on the critical water pressure and critical initiation angle. During high-pressure water injection, the fracture initiation angle decreases as the ratio between the water pressure in the fracture and the borehole and Biot’s ratio increase. The theoretical model provides a comprehensive understanding of the fracture twist, the mixed-mode fracture propagation feature, and the hydraulic fracturing optimization.

Dynamic TMT-Based Quantitative Proteomics Analysis of Critical Initiation Process of Totipotency during Cotton Somatic Embryogenesis Transdifferentiation

The somatic embryogenesis (SE) process of plants, as one of the typical responses to abiotic stresses with hormone, occurs through the dynamic expression of different proteins that constitute a complex regulatory network in biological activities and promotes plant totipotency. Plant SE includes two critical stages: primary embryogenic calli redifferentiation and somatic embryos development initiation, which leads to totipotency. The isobaric labels tandem mass tags (TMT) large-scale and quantitative proteomics technique was used to identify the dynamic protein expression changes in nonembryogenic calli (NEC), primary embryogenic calli (PEC) and globular embryos (GEs) of cotton. A total of 9369 proteins (6730 quantified) were identified; 805, 295 and 1242 differentially accumulated proteins (DAPs) were identified in PEC versus NEC, GEs versus PEC and GEs versus NEC, respectively. Eight hundred and five differentially abundant proteins were identified, 309 of which were upregulated and 496 down regulated in PEC compared with NEC. Of the 295 DAPs identified between GEs and PEC, 174 and 121 proteins were up- and down regulated, respectively. Of 1242 differentially abundant proteins, 584 and 658 proteins were up- and down regulated, respectively, in GEs versus NEC. We have also complemented the authenticity and accuracy of the proteomic analysis. Systematic analysis indicated that peroxidase, photosynthesis, environment stresses response processes, nitrogen metabolism, phytohormone response/signal transduction, transcription/posttranscription and modification were involved in somatic embryogenesis. The results generated in this study demonstrate a proteomic molecular basis and provide a valuable foundation for further investigation of the roles of DAPs in the process of SE transdifferentiation during cotton totipotency.

Regulation of clathrin-mediated endocytosis by hierarchical allosteric activation of AP2

The critical initiation phase of clathrin-mediated endocytosis (CME) determines where and when endocytosis occurs. Heterotetrameric adaptor protein 2 (AP2) complexes, which initiate clathrin-coated pit (CCP) assembly, are activated by conformational changes in response to phosphatidylinositol-4,5-bisphosphate (PIP2) and cargo binding at multiple sites. However, the functional hierarchy of interactions and how these conformational changes relate to distinct steps in CCP formation in living cells remains unknown. We used quantitative live-cell analyses to measure discrete early stages of CME and show how sequential, allosterically regulated conformational changes activate AP2 to drive both nucleation and subsequent stabilization of nascent CCPs. Our data establish that cargoes containing Yxxφ motif, but not dileucine motif, play a critical role in the earliest stages of AP2 activation and CCP nucleation. Interestingly, these cargo and PIP2 interactions are not conserved in yeast. Thus, we speculate that AP2 has evolved as a key regulatory node to coordinate CCP formation and cargo sorting and ensure high spatial and temporal regulation of CME.

Numerical Simulation and Analysis on the Impact Effect of Cylindrical Projectile Impacting Shelled Explosive

The numerical simulation of shelled Comp.B explosive was studied following the Lee-Tarver ignition and growth model when it was impacted respectively by 4340 Steel, OFHC and 93#W projectile with the same mass; the influences on explosive detonation of the initiation process, the material of projectile and the L/D ratio of projectile were analyzed; the critical initiation speeds of the projectiles of three different materials with different L/D ratio were gained. In order to verify the simulation results, the questions were calculated by the theoretical simplified model, the results of the theoretical calculation and the numerical simulation accorded well based on critical energy criterion. The results show that the capability of igniting explosive, the first is 93#W, the second is OFHC, the last is 4340 Steel; The initiation point were not on the interface of shell and explosive, and the faster of the impacting velocity, the initiation point closer the interface; the bigger of the L/D ratio of projectile, the higher of the critical initiation speed.

Reading Saint Flannery: Modernism, Sexuality, and the Culture of Psychoanalysis

Because flannery O'CONNOR was a Christian writer who suffered considerably from a painful and debilitating disease, died young, and left a legacy of remarkable stories and novels, critics of many persuasions have canonized her in ways few writers have been canonized. Since it is hard to argue with a saint, the vast majority of readers have capitulated to O'Connor's pronouncements on how to read those works. Several critics — myself among them — have reminded us how effectively O'Connor has set the terms of the discourse about her. Working in the modernist age, she thought herself not a modernist, but a throwback to an earlier age. Consequently, one of the more interesting, but generally ignored, questions about Flannery O'Connor is how to periodize her work. Whatever her claims, it is clear she is modernist in important ways. We see that in her use of myth, for example, and in how she uses the devices of lyrical or poetic fiction such as powerful controlling metaphors and recurrent image-motifs to knit together a form in place of traditional emplotments. In her practice, moreover, she is a modernist easily allied with New Criticism. Of O'Connor's adherence to its tenets, Frederick Crews says, “As she freely admitted, she came into her own as an artist only after undergoing a full New Critical initiation at the University of Iowa's Writer's Workshop under the tutelage of Paul Engle and Andrew Lytle, with Brooks and Warren's then ubiquitous Understanding Fiction providing the models” (144). Indeed, says Crews, “Even the most impressive and original of her stories adhere to the classroom formula of her day: show, don't tell; keep the narrative voice distinct from those of your characters; cultivate understatement; develop a central image or symbol to convey your theme ‘objectively’; and point everything toward one neatly sprung ironic reversal. No one ever put it all together with greater deftness” (144–45).