GTPases are regulators of cell signaling acting as molecular switches. The translational GTPase EF-G stands out, as it uses GTP hydrolysis to generate force and promote the movement of the ribosome ...along the mRNA. The key unresolved question is how GTP hydrolysis drives molecular movement. Here, we visualize the GTPase-powered step of ongoing translocation by time-resolved cryo-EM. EF-G in the active GDP-Pi form stabilizes the rotated conformation of ribosomal subunits and induces twisting of the sarcin-ricin loop of the 23 S rRNA. Refolding of the GTPase switch regions upon Pi release initiates a large-scale rigid-body rotation of EF-G pivoting around the sarcin-ricin loop that facilitates back rotation of the ribosomal subunits and forward swiveling of the head domain of the small subunit, ultimately driving tRNA forward movement. The findings demonstrate how a GTPase orchestrates spontaneous thermal fluctuations of a large RNA-protein complex into force-generating molecular movement.
Conversion of a Singlet Silylene to a stable Biradical Mondal, Kartik Chandra; Roesky, Herbert W.; Schwarzer, Martin C. ...
Angewandte Chemie (International ed.),
February 4, 2013, Volume:
52, Issue:
6
Journal Article
Peer reviewed
Silicon becomes colored: Stable biradicals were prepared from an N‐heterocyclic carbene stabilized SiCl2 and a cyclic alkyl(amino)carbene, and characterized as two polymorphs. The deep‐blue crystals ...of one polymorph are stable upon exposure to air for about a week, while the solution in THF decomposes rapidly when exposed to air. In a side reaction, the different carbene species react with each other under CH activation and CC bond formation in the presence of the biradical.
Metal ions with radical centers in their coordination sphere are key participants in biological and catalytic processes. In the present study, we describe the synthesis of the cAAC:ZnCl2 adduct (1) ...using a cyclic alkylaminocarbene (cAAC) as donor ligand. Compound 1 was treated with 2 equiv of KC8 and LiB(sec-Bu)3H to yield a deep blue-colored dicarbene zinc compound (cAAC)2Zn (2) and the colorless hydrogenated zinc compound (cAACH)2Zn (3), respectively. Compounds 2 and 3 were well characterized by spectroscopic methods and single-crystal X-ray structural analysis. Density functional theory calculations were performed for 2 which indicate that this molecule possesses a singlet biradicaloid character. Moreover, we show the application of 2 in CO2 activation, which yields a zwitterionic cAAC·CO2 adduct.
GABA
A receptors are ligand-operated chloride channels assembled from five subunits in a heteropentameric manner. Using immunocytochemistry, we investigated the distribution of GABA
A receptor ...subunits deriving from 13 different genes (α1–α6, β1–β3, γ1–γ3 and δ) in the adult rat brain. Subunit α1-, β1-, β2-, β3- and γ2-immunoreactivities were found throughout the brain, although differences in their distribution were observed. Subunit α2-, α3-, α4-, α5-, α6-, γ1- and δ-immunoreactivities were more confined to certain brain areas. Thus, α2-subunit-immunoreactivity was preferentially located in forebrain areas and the cerebellum. Subunit α6-immunoreactivity was only present in granule cells of the cerebellum and the cochlear nucleus, and subunit γ1-immunoreactivity was preferentially located in the central and medial amygdaloid nuclei, in pallidal areas, the substantia nigra pars reticulata and the inferior olive. The α5-subunit-immunoreactivity was strongest in Ammon’s horn, the olfactory bulb and hypothalamus. In contrast, α4-subunit-immunoreactivity was detected in the thalamus, dentate gyrus, olfactory tubercle and basal ganglia. Subunit α3-immunoreactivity was observed in the glomerular and external plexiform layers of the olfactory bulb, in the inner layers of the cerebral cortex, the reticular thalamic nucleus, the zonal and superficial layers of the superior colliculus, the amygdala and cranial nerve nuclei. Only faint subunit γ3-immunoreactivity was detected in most areas; it was darkest in midbrain and pontine nuclei. Subunit δ-immunoreactivity was frequently co-distributed with α4 subunit-immunoreactivity, e.g. in the thalamus, striatum, outer layers of the cortex and dentate molecular layer. Striking examples of complementary distribution of certain subunit-immunoreactivities were observed. Thus, subunit α2-, α4-, β1-, β3- and δ-immunoreactivities were considerably more concentrated in the neostriatum than in the pallidum and entopeduncular nucleus. In contrast, labeling for the α1-, β2-, γ1- and γ2-subunits prevailed in the pallidum compared to the striatum. With the exception of the reticular thalamic nucleus, which was prominently stained for subunits α3, β1, β3 and γ2, most thalamic nuclei were rich in α1-, α4-, β2- and δ-immunoreactivities. Whereas the dorsal lateral geniculate nucleus was strongly immunoreactive for subunits α4, β2 and δ, the ventral lateral geniculate nucleus was predominantly labeled for subunits α2, α3, β1, β3 and γ2; subunit α1- and α5-immunoreactivities were about equally distributed in both areas. In most hypothalamic areas, immunoreactivities for subunits α1, α2, β1, β2 and β3 were observed. In the supraoptic nucleus, staining of conspicuous dendritic networks with subunit α1, α2, β2, and γ2 antibodies was contrasted by perykarya labeled for α5-, β1- and δ-immunoreactivities. Among all brain regions, the median emminence was most heavily labeled for subunit β2-immunoreactivity. In most pontine and cranial nerve nuclei and in the medulla, only subunit α1-, β2- and γ2-immunoreactivities were strong, whereas the inferior olive was significantly labeled only for subunits β1, γ1 and γ2.
In this study, a highly heterogeneous distribution of 13 different GABA
A receptor subunit-immunoreactivities was observed. This distribution and the apparently typical patterns of co-distribution of these GABA
A receptor subunits support the assumption of multiple, differently assembled GABA
A receptor subtypes and their heterogeneous distribution within the adult rat brain.
Vertebrates usually have three class V myosin paralogues (MyoV) to control membrane trafficking in the actin-rich cell cortex, but their functional overlapping or differentiation through cargoes ...selectivity is yet only partially understood. In this work, we reveal that the globular tail domain of MyoVc binds to the active form of small GTPase Rab3A with nanomolar affinity, a feature shared with MyoVa but not with MyoVb. Using molecular docking analyses guided by chemical cross-linking restraints, we propose a model to explain how Rab3A selectively recognizes MyoVa and MyoVc via a distinct binding site from that used by Rab11A. The MyoVa/c binding interface involves multiple residues from both lobules (I and II) and the short helix at the α2-α3 link region, which is conserved between MyoVa and MyoVc, but not in MyoVb. This motif is also responsible for the selective binding of RILPL2 by MyoVa and potentially MyoVc. Together, these findings support the selective recruitment of MyoVa and MyoVc to exocytic pathways via Rab3A and expand our knowledge about the functional evolution of class V myosins.
Hormone secretion, neurotransmitter release, and cytoplasm membrane recycling are examples of processes that rely on the interaction of molecular motors and Rab GTPases to regulate the intracellular trafficking and tethering of vesicles. Defects in these proteins may cause neurological impairment, immunodeficiency, and other severe disorders, being fatal in some cases. Despite their crucial roles, little is known about how these molecular motors are selectively recruited by specific members of the large family of Rab GTPases. In this study, we unveil the interaction between the actin-based molecular motor Myosin Vc and the small GTPase Rab3A, a key coordinator of vesicle trafficking and exocytosis in mammalian cells. Moreover, we propose a model for their recognition and demonstrate that Rab3A specifically binds to the globular tail of Myosins Va and Vc, but not of Myosin Vb, advancing our knowledge about the molecular basis for the selective recruitment of class V myosins by Rab GTPases.
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Valerenic acid (VA)-a β2/3-selective GABA type A (GABAA) receptor modulator-displays anxiolytic and anticonvulsive effects in mice devoid of sedation, making VA an interesting drug candidate. Here we ...analyzed β-subunit-dependent enhancement of GABA-induced chloride currents (IGABA) by a library of VA derivatives and studied their effects on pentylenetetrazole (PTZ)-induced seizure threshold and locomotion. Compound-induced IGABA enhancement was determined in oocytes expressing α1β1γ2S, α1β2γ2S, or α1β3γ2S receptors. Effects on seizure threshold and locomotion were studied using C57BL/6N mice and compared with saline-treated controls. β2/3-selective VA derivatives such as VA-amide (VA-A) modulating α1β3γ2S (VA-A: Emax = 972 ± 69%, n = 6, P < 0.05) and α1β2γ2S receptors (Emax = 1119 ± 72%, n = 6, P < 0.05) more efficaciously than VA (α1β3γ2S: VA: Emax = 632 ± 88%, n = 9 versus α1β2γ2S: VA: Emax = 721 ± 68%, n = 6) displayed significantly more pronounced seizure threshold elevation than VA (saline control: 40.4 ± 1.4 mg/kg PTZ versus VA 10 mg/kg: 49.0 ± 1.8 mg/kg PTZ versus VA-A 3 mg/kg: 57.9 ± 1.9 mg/kg PTZ, P < 0.05). Similarly, VA's methylamide (VA-MA) enhancing IGABA through β3-containing receptors more efficaciously than VA (Emax = 1043 ± 57%, P < 0.01, n = 6) displayed stronger anticonvulsive effects. Increased potency of IGABA enhancement and anticonvulsive effects at lower doses compared with VA were observed for VA-tetrazole (α1β3γ2S: VA-TET: EC50 = 6.0 ± 1.0 μM, P < 0.05; VA-TET: 0.3 mg/kg: 47.3 ± 0.5 mg/kg PTZ versus VA: 10 mg/kg: 49.0 ± 1.8 mg/kg PTZ, P < 0.05). At higher doses (≥10 mg/kg), VA-A, VA-MA, and VA-TET reduced locomotion. In contrast, unselective VA derivatives induced anticonvulsive effects only at high doses (30 mg/kg) or did not display any behavioral effects. Our data indicate that the β2/3-selective compounds VA-A, VA-MA, and VA-TET induce anticonvulsive effects at low doses (≤10 mg/kg), whereas impairment of locomotion was observed at doses ≥10 mg/kg.
This was a cross-sectional analytic study.
In relation to pain from the sacroiliac joint, this study sought to establish 1) its prevalence, 2) the validity of pain provocation, 3) whether any ...arthrographic abnormalities predict a response to joint block, and 4) whether certain pain patterns discriminate patients with this diagnosis.
The true prevalence of sacroiliac joint pain is unknown and despite a plethora of clinical tests, none of these tests has been validated against an established criterion standard. To our knowledge, arthrography of the sacroiliac joint had never been studied.
Forty-three consecutive patients with chronic low back pain maximal below L5-S1 were investigated with sacroiliac joint blocks under image intensifier using radiographic contrast followed by 2% lignocaine. Information was obtained on pain provocation, analgesia, and image pattern.
Thirteen patients (30%) obtained gratifying relief of their pain. Nine of these also exhibited tears of their ventral capsule. Groin pain was the only pain referral pattern found to be associated with response to sacroiliac joint block.
The sacroiliac joint is a significant source of pain in patients with chronic low back pain and warrants further study.
The postfledging period of a passerine bird's life is increasingly recognized as an understudied component of songbird conservation, because the habitat needs of fledglings can differ from nesting ...adults and survivorship during this period can substantially affect population growth rates. We studied postfledging survival of the Worthington's Marsh Wren (Cistothorus palustris griseus), a state-listed salt marsh songbird subspecies in northeastern Florida, USA, during 2016-2017. Postfledging survival was low during the first 21 d following fledging for the cohort of tracked birds (0.10; 95% confidence interval: 0.01, 0.37). The top a priori model indicated that heavier birds at the time of tagging exhibited greater survival than lighter birds, and birds from larger broods had greater survival than birds from smaller broods. Other covariates not in the top model that also influenced survival included the amount of tall-form smooth cordgrass (Spartina alterniflora) within the natal patch, which positively influenced survival, and the amount of black needlerush (Juncus roemerianus) within the natal patch, which negatively affected survival. The low rates of postfledging survival combined with low rates of nest survival reported elsewhere indicate that Worthington's Marsh Wrens in northeastern Florida require continued protection and warrant further research into management and restoration activities that will improve their habitat.
Tidal salt marshes face numerous pressures that threaten their ecological integrity, and many salt marsh vertebrates have exhibited declines concomitant with the degradation of salt marsh habitats. ...We studied nest survival of MacGillivray’s seaside sparrow Ammospiza maritima macgillivraii and Worthington’s marsh wren Cistothorus palustris griseus in smooth cordgrass Spartina alterniflora and black needlerush Juncus roemerianus dominated salt marshes in northeast Florida, USA, during 2015-2017 to understand why both species have undergone range contractions. We evaluated the effects of tides, habitat features, and weather on nest survival. The probability of a nest surviving from laying to fledging young was low for both species (seaside sparrow: 0.03, 95% CI: 0.00, 0.10; marsh wren: 0.10, 95% CI: 0.04, 0.18), suggesting that nest survival may limit population growth. The most substantial threat to marsh wren and seaside sparrow nest survival was from greater than normal high tides. However, there was limited evidence of direct nest loss due to flooding. An analysis of only fledged and depredated nests suggests that higher tides influenced nest survival by increasing nest predation, substantially so for marsh wrens. Other factors negatively associated with marsh wren nest survival, albeit less strongly, included precipitation and the amount of black needlerush at and within 50 m of the nest. The effect of tide height on nest survival suggests that salt marsh birds in northeast Florida are vulnerable to continued sea-level rise, which is exacerbated by upland development that will hinder marsh migration. Monitoring and protecting northeast Florida’s high-elevation, cordgrass-dominated marshes should be a priority.
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