Rustless translation

ATP binding cassette proteins are a large and diverse family of molecular machines and include transmembrane transporter, chromosome maintenance and DNA repair proteins, and translation factors. However, the function of the ABCE1, the only member of subfamily E of ABC proteins, remained mysterious for over a decade, even though it is perhaps the most conserved ABC protein in eukaryotes and archaea. Recent results have now identified ABCE1 as the ribosome-recycling factor of eukaryotes and archaea. Thus, two iron-sulfur clusters - the hallmark feature of ABCE1 - help catalyze an integral step of the translational cycle at the core of the protein synthesis machinery

DNA Double-Strand Breaks Come into Focus

The Mre11-Rad50-Nbs1 (MRN) complex senses DNA double-strand breaks and recruits different repair pathway and checkpoint proteins to break foci. Two new studies (Williams et al., 2009; Lloyd et al., 2009) identify Nbs1 as a key factor in this process and reveal how an N-terminal protein recruitment module in Nbs1 binds to different response factors through shared phosphopeptide motifs

ATP puts the brake on DNA double-strand break repair

DNA double-strand breaks (DSBs) are one of the most deleterious forms of DNA damage and can result in cell inviability or chromosomal aberrations. The Mre11-Rad50-Nbs1 (MRN) ATPase-nuclease complex is a central player in the cellular response to DSBs and is implicated in the sensing and nucleolytic processing of DSBs, as well as in DSB signaling by activating the cell cycle checkpoint kinase ATM. ATP binding to Rad50 switches MRN from an open state with exposed Mre11 nuclease sites to a closed state with partially buried nuclease sites. The functional meaning of this switch remained unclear. A new study shows that ATP binding to Rad50 promotes DSB recognition, tethering, and ATM activation, while ATP hydrolysis opens the nuclease active sites to promote processing of DSBs. MRN thus emerges as functional switch that may coordinate the temporal transition from signaling to processing of DSBs

Architectures and mechanisms of ATP binding cassette proteins

ATP binding cassette (ABC) ATPases form chemo-mechanical engines and switches that function in a broad range of biological processes. Most prominently, a very large family of integral membrane NTPases-ABC transporters-catalyzes the import or export of a diverse molecules across membranes. ABC proteins are also important components of the chromosome segregation, recombination, and DNA repair machineries and regulate or catalyze critical steps of ribosomal protein synthesis. Recent structural and mechanistic studies draw interesting architectural and mechanistic parallels between diverse ABC proteins. Here, I review this state of our understanding how NTP-dependent conformational changes of ABC proteins drive diverse biological processes. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 492-504, 2016

Chromosome Biology: The Crux of the Ring

SMC proteins are key components of large ring-shaped chromosomal protein complexes, such as cohesin and condensin. New evidence supports the idea that these rings topologically encircle DNA. Hints also emerge as to what it may take for DNA to enter the ring

Insulinomimetics and the vascular endothelin system

Insulin promotes vasodilatory, vasoconstrictor and atherogenic influences on vascular smooth muscle. Endothelin-1 (ET-1) is a potent, endothelium derived vasoconstrictor/mitogenic peptide whose production is stimulated by insulin both 'in vitro' and 'in vivo'. Consequently, ET-1 has been hypothesized to contribute to insulin evoked vasculopathy. The present study first characterized the effect of insulin and the insulinomimetic agent, vanadate, on ET receptor expression and action in rat aortic smooth muscle cells. Radioligand binding studies confirmed that insulin pretreatment selectively upregulated ETA receptor expression in a concentration and time dependent manner, whereas vanadate upregulated both ETA and ETB. Upregulated ET receptors were coupled to increased [Ca 2+]i responses to ET agonists. Upregulation by both compounds occurred at the mRNA level and required tyrosine kinase activation, active transcription, and new protein synthesis. In order to determine the ' in vivo' relevance of these effects, both insulin deficient diabetic and non-diabetic rats were treated with insulin and vanadate for 2-weeks. Strikingly, the diabetic state 'per se' was associated with both attenuated ET-1 plasma levels and exaggerated 'ex vivo' aortic vasoconstrictor responses to ET-1. Two weeks of treatment with either insulin or vanadate did not further increase ET-1 evoked vasoconstriction as expected based on our cellular data, but in fact restored altered ET-1 release and action to normal in the diabetic rats. It is concluded that while insulin and vanadate might increase ET receptor expression and subsequent adverse vascular responses to ET-1, their profound beneficial effects on metabolic function and the resulting beneficial vascular actions clearly minimize the significance of these actions

Crystal Structure of Human TWEAK in Complex with the Fab Fragment of a Neutralizing Antibody Reveals Insights into Receptor Binding.

The tumor necrosis factor-like weak inducer of apoptosis (TWEAK) is a multifunctional cytokine playing a key role in tissue regeneration and remodeling. Dysregulation of TWEAK signaling is involved in various pathological processes like autoimmune diseases and cancer. The unique interaction with its cognate receptor Fn14 makes both ligand and receptor promising targets for novel therapeutics. To gain insights into this important signaling pathway, we determined the structure of soluble human TWEAK in complex with the Fab fragment of an antibody selected for inhibition of receptor binding. In the crystallized complex TWEAK is bound by three Fab fragments of the neutralizing antibody. Homology modeling shows that Fab binding overlaps with the putative Fn14 binding site of TWEAK. Docking of the Fn14 cysteine rich domain (CRD) to that site generates a highly complementary interface with perfectly opposing charged and hydrophobic residues. Taken together the presented structure provides new insights into the biology of TWEAK and the TWEAK/Fn14 pathway, which will help to optimize the therapeutic strategy for treatment of related cancer types and autoimmune diseases

Vascular endothelial growth factor production and regulation in rodent and human pituitary tumor cells in vitro

Angiogenesis, the formation of a new blood supply, is an essential step in tumorigenesis. Although vascular endothelial growth factor (VEGF) is known to be a very potent angiogenic factor in most solid tumors, little is known about its production and regulation in pituitary adenomas. We have investigated basal and stimulated VEGF production by rodent pituitary tumor cells (mouse corticotrope AtT20, rat lactosomatotrope GH3, mouse gonadotrope alpha T3-1 and mouse folliculostellate TtT/GF cells), and by hormone-inactive (27), corticotrope (9), lactotrope (3) and somatotrope (21) human pituitary adenoma cell cultures. All 4 pituitary cell lines secreted VEGF, which in the case of AtT20, GH3 and TtT/GF cells was inhibited by approximately 50% by dexamethasone. TtT/GF cells were the most responsive to the different stimuli used since basal values were augmented by pituitary adenylate cyclase activating polypeptide-38 (PACAP-38), interleukin-6 (IL-6), transforming growth factor-cc (TGF-a), IGF-I and the somatostatin analogue ocreotide. However, in GH3, AtT20 and aT3-1 cells, basal VEGF levels where not enhanced with any of the stimuli tested. The majority of the human adenomas tested (92%) basally secreted measurable VEGF which was inhibited by dexamethasone in most cases (84%). VEGF levels were increased in hormone inactive adenomas, somatotrope tumors and prolactinomas by TGF-alpha, PACAP-38, and 17 beta -estradiol, respectively. In conclusion, pituitary tumor cells are capable of producing VEGF which may be involved in tumoral angiogenesis. Our results concerning the suppression of VEGF by dexamethasone suggest that glucocorticoids may have anti-angiogenic properties and therefore therapeutic relevance for the treatment of pituitary adenomas

Distinct requirements for the Rad32(Mre¹¹) nuclease and Ctp1(CtIP) in the removal of covalently bound topoisomerase I and II from DNA

For a cancer cell to resist treatment with drugs that trap topoisomerases covalently on the DNA, the topoisomerase must be removed. In this study, we provide evidence that the Schizosaccharomyces pombe Rad32Mre11 nuclease activity is involved in the removal of both Top2 from 5′ DNA ends as well as Top1 from 3′ ends in vivo. A ctp1CtIP deletion is defective for Top2 removal but overproficient for Top1 removal, suggesting that Ctp1CtIP plays distinct roles in removing topoisomerases from 5′ and 3′ DNA ends. Analysis of separation of function mutants suggests that MRN-dependent topoisomerase removal contributes significantly to resistance against topoisomerase-trapping drugs. This study has important implications for our understanding of the role of the MRN complex and CtIP in resistance of cells to a clinically important group of anticancer drugs