Prion-like domain mutations in hnRNPs cause multisystem proteinopathy and ALS

Summary Algorithms designed to identify canonical yeast prions predict that ~250 human proteins, including several RNA-binding proteins associated with neurodegenerative disease, harbor a distinctive prion-like domain (PrLD) enriched in uncharged polar amino acids and glycine. PrLDs in RNA-binding proteins are essential for the assembly of ribonucleoprotein granules. However, the interplay between human PrLD function and disease is not understood. Here, we define pathogenic mutations in PrLDs of hnRNPA2/B1 and hnRNPA1 in families with inherited degeneration affecting muscle, brain, motor neuron and bone, and a case of familial ALS. Wild-type hnRNPA2 and hnRNPA1 display an intrinsic tendency to assemble into self-seeding fibrils, which is exacerbated by the disease mutations. Indeed, the pathogenic mutations strengthen a ‘steric zipper’ motif in the PrLD, which accelerates formation of self-seeding fibrils that cross-seed polymerization of wild-type hnRNP. Importantly, the disease mutations promote excess incorporation of hnRNPA2 and hnRNPA1 into stress granules and drive the formation of cytoplasmic inclusions in animal models that recapitulate the human pathology. Thus, dysregulated polymerization caused by a potent mutant ‘steric zipper’ motif in a PrLD can initiate degenerative disease. Related proteins with PrLDs must be considered candidates for initiating and perhaps propagating proteinopathies of muscle, brain, motor neuron and bone

The Crystal Structure of the Escherichia coli Autoinducer-2 Processing Protein LsrF

Many bacteria produce and respond to the quorum sensing signal autoinducer-2 (AI-2). Escherichia coli and Salmonella typhimurium are among the species with the lsr operon, an operon containing AI-2 transport and processing genes that are up regulated in response to AI-2. One of the Lsr proteins, LsrF, has been implicated in processing the phosphorylated form of AI-2. Here, we present the structure of LsrF, unliganded and in complex with two phospho-AI-2 analogues, ribose-5-phosphate and ribulose-5-phosphate. The crystal structure shows that LsrF is a decamer of (αβ)8-barrels that exhibit a previously unseen N-terminal domain swap and have high structural homology with aldolases that process phosphorylated sugars. Ligand binding sites and key catalytic residues are structurally conserved, strongly implicating LsrF as a class I aldolase

Molecular Determinants and Genetic Modifiers of Aggregation and Toxicity for the ALS Disease Protein FUS/TLS

A combination of yeast genetics and protein biochemistry define how the fused in sarcoma (FUS) protein might contribute to Lou Gehrig's disease

Structure-based sequence alignment highlights conservation of binding-site and potential catalytic residues.

<p>Structure-based alignments <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0006820#pone.0006820-Lupyan1" target="_blank">[47]</a> were calculated for LsrF with rabbit FBPA (1J4E), <i>T. tenax</i> FBPA (1OK4), and <i>M. jannaschii</i> ADH synthase (2QJG). Identical residues are in white on red, conserved residues are in red (boxed). Secondary structure (from LsrF) is indicated above the sequence: blue bars are α-helices and red arrows are β-sheets. Residues implicated as either hydrogen bonding to the ligand phosphate or catalytic are indicated with an asterisk; these residues are disproportionately conserved. Numbering follows the LsrF sequence.</p

Structures of phospho-AI-2 and two analogues, ribulose-5-phosphate and ribose-5-phosphate.

<p>Structures of phospho-AI-2 and two analogues, ribulose-5-phosphate and ribose-5-phosphate.</p

Structure of the LsrF decamer.

<p>A. Surface representation of the LsrF decamer, viewed down the 5-fold symmetry axis, with each monomer a different color. The bound ligand (ribose-5-phosphate) is visible in the center of the (αβ)₈-barrel, and is shown in ball-and-stick format. B. Perpendicular view of the LsrF decamer along a two-fold axis.</p

Structure of a single LsrF chain.

<p>A. Stereoview of a single (α/β)8-barrel subunit with protein backbone in cartoon representation and bound P-AI-2 analogue (ribulose-5-phosphate) as ball-and-stick. The protein backbone is rainbow colored, with blue at the N-terminus and red at the C-terminus. B. Rotated view of the subunit (approximately 90°) highlighting the N-terminal residues that extend away from the (αβ)8-barrel and are swapped with the adjacent 2-fold related subunit. C. Identification of the components of the (αβ)8-barrel, with α-helices as blue cylinders and β-sheets as red arrows. The bound ligand (ribulose-5-phosphate) is shown in ball-and-stick format.</p

The LsrF ligand binding site and potential catalytic residues.

<p>A. Stereoview of ribulose-5-phosphate bound to LsrF showing 20-fold NCS averaged 2F₀-F_C electron density. Density was contoured at 4.0 (red) and 2.0 (blue) σ and truncated 2.0 Å from ligand atoms. The position of the phosphate is unambiguous, and the general path of the ligand is clear. B. Stereoview of ribose-5-phosphate bound to LsrF showing 20-fold NCS averaged 2F₀-F_C electron density. Density was contoured at 5.0 (red) and 2.0 (blue) σ and truncated 2.0 Å from the ligand. The position of the phosphate is unambiguous, and the general path of the ligand is clear. C. Structural alignment of key catalytic residues from rabbit (blue bonds; 1J4E) and <i>T. tenax</i> (red bonds; 1OK4) FBPA with LsrF (white bonds). Ribulose-5-phosphate from LsrF is shown in ball and stick form. Residue numbering follows LsrF.</p

Crystallographic data and refinement statistics.

<p>Crystallographic data and refinement statistics.</p

Inhibition of RNA lariat debranching enzyme suppresses TDP-43 toxicity in ALS disease models

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease primarily affecting motor neurons. Mutations in the gene encoding TDP-43 cause some forms of the disease, and cytoplasmic TDP-43 aggregates accumulate in degenerating neurons of most individuals with ALS. Thus, strategies aimed at targeting the toxicity of cytoplasmic TDP-43 aggregates may be effective. Here, we report results from two genome-wide loss-of-function TDP-43 toxicity suppressor screens in yeast. The strongest suppressor of TDP-43 toxicity was deletion of DBR1, which encodes an RNA lariat debranching enzyme. We show that, in the absence of Dbr1 enzymatic activity, intronic lariats accumulate in the cytoplasm and likely act as decoys to sequester TDP-43, preventing it from interfering with essential cellular RNAs and RNA-binding proteins. Knockdown of Dbr1 in a human neuronal cell line or in primary rat neurons is also sufficient to rescue TDP-43 toxicity. Our findings provide insight into TDP-43-mediated cytotoxicity and suggest that decreasing Dbr1 activity could be a potential therapeutic approach for ALS