RNF144A Sustains EGFR Signaling to Promote EGF-Dependent Cell Proliferation [Protein Synthesis and Degradation]

August 31st, 2018 by Shiuh-Rong Ho, Weei-Chin Lin

RNF144A is a single-pass transmembrane RBR E3 ligase that interacts with and degrades cytoplasmic DNA-PKcs, which is an EGFR-interacting partner. Interestingly, RNF144A expression is positively correlated with EGFR mRNA and protein levels in several types of cancer. However, the relationship between RNF144A and EGFR is poorly understood. This study reports an unexpected role for RNF144A in the regulation of EGF/EGFR signaling and EGF-dependent cell proliferation. EGFR ligands, but not DNA-damaging agents, induce a DNA-PKcs-independent interaction between RNF144A and EGFR. RNF144A promotes EGFR ubiquitination, maintains EGFR protein and prolongs EGF/EGFR signaling during EGF stimulation. Moreover, depletion of RNF144A by multiple independent approaches results in a decrease in EGFR expression and EGF/EGFR signaling. RNF144A knockout cells also fail to mount an immediate response to EGF for activation of G1/S progression genes. Consequently, depletion of RNF144A reduces EGF-dependent cell proliferation. These defects may be at least in part due to a role for RNF144A in regulating EGFR transport in the intracellular vesicles during EGF treatment.

Identification of a Kdn biosynthesis pathway in the haptophyte Prymnesium parvum suggests widespread sialic acid biosynthesis among microalgae [Enzymology]

August 31st, 2018 by Ben A Wagstaff, Martin Rejzek, Robert A. Field

Sialic acids are a family of more than 50 structurally distinct acidic sugars on the surface of all vertebrate cells where they terminate glycan chains and are exposed to many interactions with the surrounding environment. In particular, sialic acids play important roles in cell-cell and host-pathogen interactions. The sialic acids or related nonulosonic acids have been observed in Deuterostome lineages, Eubacteria, and Archaea, but are notably absent from plants. However, the structurally related C8 acidic sugar, 3-deoxy-D-manno-2-octulosonic acid (Kdo), is present in Gram-negative bacteria and plants as a component of bacterial lipopolysaccharide and pectic rhamnogalacturonan-II in the plant cell wall. Until recently, sialic acids were not thought to occur in algae, but as in plants, Kdo has been observed in algae. Here, we report the de novo biosynthesis of the deaminated sialic acid, 3-deoxy-D-glycero-D-galacto-2-nonulosonic acid (Kdn), in the toxin-producing microalga Prymnesium parvum. Using biochemical methods, we show that this alga contains CMP–Kdn and identified and recombinantly expressed the P. parvum genes encoding Kdn-9-P synthetase and CMP-Kdn synthetase enzymes that convert mannose-6-P to CMP–Kdn. Bioinformatics analysis revealed sequences related to those of the two P. parvum enzymes, suggesting that sialic acid biosynthesis is likely more widespread among microalgae than previously thought and that this acidic sugar may play a role in host-pathogen interactions involving microalgae. Our findings provide evidence that P. parvum has the biosynthetic machinery for de novo production of the deaminated sialic acid Kdn and that sialic acid biosynthesis may be common among microalgae.
  • Posted in Journal of Biological Chemistry, Publications
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Acetate-dependent tRNA acetylation required for decoding fidelity in protein synthesis

August 27th, 2018 by Takaaki Taniguchi

Acetate-dependent tRNA acetylation required for decoding fidelity in protein synthesis

Acetate-dependent tRNA acetylation required for decoding fidelity in protein synthesis, Published online: 27 August 2018; doi:10.1038/s41589-018-0119-z

A comparative genomic approach identified a novel acetate-dependent tRNA-modifying enzyme that catalyzes RNA acetylation with a mechanism similar to tRNA aminoacylation. This modification maintains decoding fidelity in protein synthesis.
  • Posted in Nat Chem Biol, Publications
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Nuclear RNR-α antagonizes cell proliferation by directly inhibiting ZRANB3

August 27th, 2018 by Yuan Fu

Nuclear RNR-α antagonizes cell proliferation by directly inhibiting ZRANB3

Nuclear RNR-α antagonizes cell proliferation by directly inhibiting ZRANB3, Published online: 27 August 2018; doi:10.1038/s41589-018-0113-5

The large subunit of ribonucleotide reductase RNR-α downregulates DNA replication in the nucleus by directly disrupting PCNA and ZRANB3 interactions. RNR-α nuclear entry is regulated by an interplay between IRBIT and importin-α1.
  • Posted in Nat Chem Biol, Publications
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Are N- and C-terminally Truncated A{beta} species key pathological triggers in Alzheimer’s disease? [Neurobiology]

August 24th, 2018 by Julie Dunys, Audrey Valverde, Frederic Checler

The histopathology of Alzheimer’s disease (AD) is characterized by neuronal loss, neurofibrillary tangles, and senile plaque formation. The latter results from an exacerbated production (familial AD cases) or altered degradation (sporadic cases) of 40/42 amino-acid-long β-amyloid peptides (Aβ peptides) that are produced by sequential cleavages of Aβ precursor protein (βAPP) by β- and γ-secretases. The amyloid cascade hypothesis proposes a key role for the full-length Aβ42 and the Aβ40/42 ratio in AD etiology, in which soluble Aβ oligomers lead to neurotoxicity, tau hyperphosphorylation, aggregation, and, ultimately, cognitive defects. However, following this postulate, during the last decade, several clinical approaches aimed at decreasing full-length Aβ42 production or neutralizing it by immunotherapy have failed to reduce or even stabilize AD-related decline. Thus, the Aβ peptide (Aβ40/42)-centric hypothesis is probably a simplified view of a much more complex situation involving a multiplicity of APP fragments and Aβ catabolites. Indeed, biochemical analyses of AD brain deposits and fluids have unraveled an Aβ peptidome consisting of additional Aβ-related species. Such Aβ catabolites could be due to either primary enzymatic cleavages of βAPP or secondary processing of Aβ itself by exopeptidases. Here, we review the diversity of N- and C-terminally truncated Aβ peptides and their biosynthesis and outline their potential function/toxicity. We also highlight their potential as new pharmaceutical targets and biomarkers.

p25 of the dynactin complex plays a dual role in cargo binding and dynactin regulation [Cell Biology]

August 24th, 2018 by Rongde Qiu, Jun Zhang, Xin Xiang

Cytoplasmic dynein binds its cargoes via the dynactin complex and cargo adapters, and the dynactin pointed-end protein p25 is required for dynein-dynactin binding to the early endosomal dynein adapter HookA (Hook in the fungus Aspergillus nidulans). However, it is unclear whether the HookA-dynein-dynactin interaction requires p27, another pointed-end protein forming heterodimers with p25 within vertebrate dynactin. Here, live-cell imaging and biochemical pull-down experiments revealed that although p27 is a component of the dynactin complex in A. nidulans, it is dispensable for dynein-dynactin to interact with ΔC-HookA (cytosolic HookA lacking its early endosome-binding C terminus) and not critical for dynein-mediated early endosome transport. Using mutagenesis, imaging, and biochemical approaches, we found that several p25 regions are required for the ΔC-HookA-dynein-dynactin interaction, with the N terminus and Loop1 being the most critical regions. Interestingly, p25 was also important for the microtubule (MT) plus-end accumulation of dynactin. This p25 function in dynactin localization also involved p25’s N terminus and Loop1 critical for the ΔC-HookA-dynein-dynactin interaction. Given that dynactin’s MT plus-end localization does not require HookA and that the kinesin-1-dependent plus-end accumulation of dynactin is unnecessary for the ΔC-HookA-dynein-dynactin interaction, our results indicate that p25 plays a dual role in cargo binding and dynactin regulation. As cargo adapters are implicated in dynein activation via binding to dynactin’s pointed end to switch the conformation of p150, a major dynactin component, our results suggest p25 as a critical pointed-end protein involved in this process.

[ASAP] Variant Bacterial Riboswitches Associated with Nucleotide Hydrolase Genes Sense Nucleoside Diphosphates

August 23rd, 2018 by Madeline E. Sherlock, Harini Sadeeshkumar, Ronald R. Breaker

TOC Graphic

Biochemistry
DOI: 10.1021/acs.biochem.8b00617

[ASAP] Generation and Comparative Kinetic Analysis of New Glycosynthase Mutants from <italic toggle=”yes”>Streptococcus pyogenes</italic> Endoglycosidases for Antibody Glycoengineering

August 21st, 2018 by Xin Tong, Tiezheng Li, Chao Li, Lai-Xi Wang

TOC Graphic

Biochemistry
DOI: 10.1021/acs.biochem.8b00719

Thioridazine inhibits self-renewal in breast cancer cells via DRD2-dependent STAT3 inhibition, but induces a G1 arrest independent of DRD2 [Signal Transduction]

August 21st, 2018 by Matthew Tegowski, Cheng Fan, Albert S. Baldwin

Thioridazine is an antipsychotic that has been shown to induce cell death and inhibit self-renewal in a broad spectrum of cancer cells. The mechanisms by which these effects are mediated are currently unknown but are presumed to result from the inhibition of dopamine receptor 2 (DRD2). Here we show that the self-renewal of several, but not all, triple-negative breast cancer cell lines is inhibited by thioridazine. The inhibition of self-renewal by thioridazine in these cells is mediated by DRD2 inhibition. Further, we demonstrate that DRD2 promotes self-renewal in these cells via a STAT3 and IL-6-dependent mechanism. We also show that thioridazine induces a G1 arrest and a loss in cell viability in all tested cell lines. However, the reduction in proliferation and cell viability is independent of DRD2 and STAT3. Our results indicate that while there are cell types in which DRD2 inhibition results in inhibition of STAT3 and self-renewal, the dramatic block in cancer cell proliferation across many cell lines caused by thioridazine treatment is independent of DRD2 inhibition.
  • Posted in Journal of Biological Chemistry, Publications
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Continuous directed evolution of proteins with improved soluble expression

August 20th, 2018 by Tina Wang

Continuous directed evolution of proteins with improved soluble expression

Continuous directed evolution of proteins with improved soluble expression, Published online: 20 August 2018; doi:10.1038/s41589-018-0121-5

Through use of a split-intein pIII, soluble expression phage-assisted continuous evolution (SE-PACE) enables two simultaneous positive selections to rapidly evolve proteins with improved expression while maintaining their desired activities.
  • Posted in Nat Chem Biol, Publications
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