Functional and Biochemical Characterization of Human Eukaryotic Translation Initiation Factor 3 in Living Cells

The main role of the translation initiation factor 3 (eIF3) is to orchestrate formation of 43S-48S preinitiation complexes (PICs). Until now, most of our knowledge on eIF3 functional contribution to regulation of gene expression comes from yeast studies. Hence, here we developed several novel in vivo assays to monitor the integrity of the 13-subunit human eIF3 complex, defects in assembly of 43S PICs, efficiency of mRNA recruitment, and postassembly events such as AUG recognition. We knocked down expression of the PCI domain-containing eIF3c and eIF3a subunits and of eIF3j in human HeLa and HEK293 cells and analyzed the functional consequences. Whereas eIF3j downregulation had barely any effect and eIF3a knockdown disintegrated the entire eIF3 complex, eIF3c knockdown produced a separate assembly of the a, b, g, and i subunits (closely resembling the yeast evolutionary conserved eIF3 core), which preserved relatively high 40S binding affinity and an ability to promote mRNA recruitment to 40S subunits and displayed defects in AUG recognition. Both eIF3c and eIF3a knockdowns also severely reduced protein but not mRNA levels of many other eIF3 subunits and indeed shut off translation. We propose that eIF3a and eIF3c control abundance and assembly of the entire eIF3 and thus represent its crucial scaffolding elements critically required for formation of PICs.     

Reforestations of Tropical Forests Alter Interactions Between Web-Building Spiders and Their Prey

Ecosystems - Immense effort has been devoted to mitigating the negative effect of deforestation, one of the main factors causing global change. However, the effect of reforestation management on...     

Restriction of an intron size en route to endothermy

Ca²⁺-insensitive and -sensitive E1 subunits of the 2-oxoglutarate dehydrogenase complex (OGDHC) regulate tissue-specific NADH and ATP supply by mutually exclusive OGDH exons 4a and 4b. Here we show that their splicing is enforced by distant lariat branch points (dBPs) located near the 5′ splice site of the intervening intron. dBPs restrict the intron length and prevent transposon insertions, which can introduce or eliminate dBP competitors. The size restriction was imposed by a single dominant dBP in anamniotes that expanded into a conserved constellation of four dBP adenines in amniotes. The amniote clusters exhibit taxon-specific usage of individual dBPs, reflecting accessibility of their extended motifs within a stable RNA hairpin rather than U2 snRNA:dBP base-pairing. The dBP expansion took place in early terrestrial species and was followed by a uridine enrichment of large downstream polypyrimidine tracts in mammals. The dBP-protected megatracts permit reciprocal regulation of exon 4a and 4b by uridine-binding proteins, including TIA-1/TIAR and PUF60, which promote U1 and U2 snRNP recruitment to the 5′ splice site and BP, respectively, but do not significantly alter the relative dBP usage. We further show that codons for residues critically contributing to protein binding sites for Ca²⁺ and other divalent metals confer the exon inclusion order that mirrors the Irving-Williams affinity series, linking the evolution of auxiliary splicing motifs in exons to metallome constraints. Finally, we hypothesize that the dBP-driven selection for Ca²⁺-dependent ATP provision by E1 facilitated evolution of endothermy by optimizing the aerobic scope in target tissues.     

Calreticulin and cancer

Calreticulin (CALR) is an endoplasmic reticulum (ER)-resident protein involved in a spectrum of cellular processes. In healthy cells, CALR operates as a chaperone and Ca²⁺ buffer to assist correct protein folding within the ER. Besides favoring the maintenance of cellular proteostasis, these cell-intrinsic CALR functions support Ca²⁺-dependent processes, such as adhesion and integrin signaling, and ensure normal antigen presentation on MHC Class I molecules. Moreover, cancer cells succumbing to immunogenic cell death (ICD) expose CALR on their surface, which promotes the uptake of cell corpses by professional phagocytes and ultimately supports the initiation of anticancer immunity. Thus, loss-of-function CALR mutations promote oncogenesis not only as they impair cellular homeostasis in healthy cells, but also as they compromise natural and therapy-driven immunosurveillance. However, the prognostic impact of total or membrane-exposed CALR levels appears to vary considerably with cancer type. For instance, while genetic CALR defects promote pre-neoplastic myeloproliferation, patients with myeloproliferative neoplasms bearing CALR mutations often experience improved overall survival as compared to patients bearing wild-type CALR. Here, we discuss the context-dependent impact of CALR on malignant transformation, tumor progression and response to cancer therapy.Subject terms: Oncogenes, Tumour immunology, Protein folding, Endoplasmic reticulum     

Three New Ichnogenera of Biting and Gnawing Traces on Reptilian and Mammalian Bones: A Case Study from the Miocene of the Czech Republic

The Ahníkov (Miocene, Czech Republic) site represents a concentration of vertebrate skeletal remains in a swamp setting. Autochthonous bone deposits were strongly altered by sedimentary processes and early diagenesis. Biting and gnawing traces recognized on hard animal tissues (bones, teeth, antlers, turtle thoraces) represent seven recurring morphotypes. The following ichnotaxa are erected: Nihilichnus nihilicus n. igen. et n. isp., Nihilichnus mortalis n. isp., Machichnus regularis n. igen. et n. isp., Machichnus multilineatus n. isp., Machichnus bohemicus n. isp., and Brutalichnus brutalis n. igen. et n. isp. Each kind of bones or similar substrates bears a specific proportion of various bite traces but no observed morphotype is specific for a single substrate. Ethologically, traces of sharpening of teeth are principally different from predation traces. The beavers Steneofiber eseri and Steneofiber depereti, the carnivore Amphicyon sp. and the crocodiles are presumed as the tracemakers.     

Trophic niche and diet composition of the northernmost population of the Mediterranean horseshoe bat (Rhinolophus euryale) with conservation implications

Mammal Research - The diet of Rhinolophus euryale was studied in southeastern Slovakia by an analysis of faeces collected from netted bats. Prey availability was studied using light traps, counting...     

Time-Resolved Fluorescence in Lipid Bilayers: Selected Applications and Advantages over Steady State

Fluorescence methods are versatile tools for obtaining dynamic and topological information about biomembranes because the molecular interactions taking place in lipid membranes frequently occur on the same timescale as fluorescence emission. The fluorescence intensity decay, in particular, is a powerful reporter of the molecular environment of a fluorophore. The fluorescence lifetime can be sensitive to the local polarity, hydration, viscosity, and/or presence of fluorescence quenchers/energy acceptors within several nanometers of the vicinity of a fluorophore. Illustrative examples of how time-resolved fluorescence measurements can provide more valuable and detailed information about a system than the time-integrated (steady-state) approach will be presented in this review: 1), determination of membrane polarity and mobility using time-dependent spectral shifts; 2), identification of submicroscopic domains by fluorescence lifetime imaging microscopy; 3), elucidation of membrane leakage mechanisms from dye self-quenching assays; and 4), evaluation of nanodomain sizes by time-resolved Förster resonance energy transfer measurements.     

An insect boring in an Early Cretaceous wood from Bornholm,Denmark

Abstract

An insect boring of unique shape is described from a lignitic layer within the Early Cretaceous (Berriasian) Skyttegård Member of the Rabekke Formation on Bornholm. Morphologically it cannot be compared to any modern or fossil wood borings, although some structures are reminiscent of Scolytidae, Platypodidae and Lymexylonidae. Most probably, however, the tracemaker was a female fungus-farming beetle, thus producing an agrichnion.     

Microbially induced wrinkle structures in Middle Devonian siliciclastics from the Prague Basin,Czech Republic

The occurrence of wrinkle structures in Middle Devonian deep‐water siliciclastic sequences of the Prague Basin (Roblín Member, Srbsko Formation, Givetian) is reported and interpreted to be microbially induced. Current and/or gravitational forces are considered the simplest explanation of the origin of these structures. Taking into account the tectonic activity linked with ongoing Variscan orogeny, the wrinkle structures could also be interpreted as soft sediment deformation structures originating due to exposure of mat‐bearing sediment to seismic shocks. The distribution of n‐alkanes and isoprenoids suggests two types of prevailing biological sources, namely phytoplankton, representing organic material from the water column, and benthic bacteria. Judging from the sedimentary facies and lack of petrographic characteristics suggestive of light competition, the microbial mats are interpreted to be formed by non‐autotrophic bacteria.