Susan Lindquist

Bio: Susan Lindquist is an academic researcher from Massachusetts Institute of Technology. The author has contributed to research in topics: Heat shock protein & Saccharomyces cerevisiae. The author has an hindex of 147, co-authored 440 publications receiving 81067 citations. Previous affiliations of Susan Lindquist include University of Illinois at Chicago & Howard Hughes Medical Institute.

Prion formation by a yeast GLFG nucleoporin.

Abstract: The self-assembly of proteins into higher order structures is both central to normal biology and a dominant force in disease. Certain glutamine/asparagine (Q/N)-rich proteins in the budding yeast Saccharomyces cerevisiae assemble into self-replicating amyloid-like protein polymers, or prions, that act as genetic elements in an entirely protein-based system of inheritance. The nuclear pore complex (NPC) contains multiple Q/N-rich proteins whose self-assembly has also been proposed to underlie structural and functional properties of the NPC. Here we show that an essential sequence feature of these proteins—repeating GLFG motifs—strongly promotes their self-assembly into amyloids with characteristics of prions. Furthermore, we demonstrate that Nup100 can form bona fide prions, thus establishing a previously undiscovered ability of yeast GLFG nucleoporins to adopt this conformational state in vivo.

Analysis of the AAA sensor-2 motif in the C-terminal ATPase domain of Hsp104 with a site-specific fluorescent probe of nucleotide binding

Abstract: Hsp104 from Saccharomyces cerevisiae is a hexameric protein with two AAA ATPase domains (N- and C-terminal nucleotide-binding domains NBD1 and NBD2, respectively) per monomer. Our previous analysis of the Hsp104 ATP hydrolysis cycle revealed that NBD1 and NBD2 have very different catalytic properties, but each shows positive cooperativity in hydrolysis. There is also communication between the two domains, in that ATP hydrolysis at NBD1 depends on the nucleotide that is bound to NBD2. Here, we extend our understanding of the Hsp104 ATP hydrolysis cycle through mutagenesis of the AAA sensor-2 motif in NBD2. To do so, we took advantage of the lack of tryptophan residues in Hsp104 to place a single tryptophan in the C-terminal domain (Y819W). The Y819W substitution has no significant effects on folding stability of the C-terminal domain or on ATP hydrolysis by NBD1 or NBD2. The fluorescence of this tryptophan changes in response to ATP and ADP binding, allowing the Kd and Hill coefficient to be determined for each nucleotide. By using this site-specific probe of binding, we analyze the effect of mutating the conserved arginine residue in the sensor-2 motif in Hsp104 NBD2. An R826M mutation causes nearly equal decreases in affinity of NBD2 for both ATP and ADP, indicating that at this site, the sensor-2 provides binding energy, but does not act to sense the difference between these nucleotides. In addition, the rate of ATP hydrolysis at NBD1 is decreased by the R826M mutation, providing further evidence for interdomain communication in the Hsp104 ATP hydrolysis cycle.

Structural basis for species-selective targeting of Hsp90 in a pathogenic fungus.

Abstract: New strategies are needed to counter the escalating threat posed by drug-resistant fungi. The molecular chaperone Hsp90 affords a promising target because it supports survival, virulence and drug-resistance across diverse pathogens. Inhibitors of human Hsp90 under development as anticancer therapeutics, however, exert host toxicities that preclude their use as antifungals. Seeking a route to species-selectivity, we investigate the nucleotide-binding domain (NBD) of Hsp90 from the most common human fungal pathogen, Candida albicans. Here we report structures for this NBD alone, in complex with ADP or in complex with known Hsp90 inhibitors. Encouraged by the conformational flexibility revealed by these structures, we synthesize an inhibitor with >25-fold binding-selectivity for fungal Hsp90 NBD. Comparing co-crystals occupied by this probe vs. anticancer Hsp90 inhibitors revealed major, previously unreported conformational rearrangements. These insights and our probe’s species-selectivity in culture support the feasibility of targeting Hsp90 as a promising antifungal strategy. The chaperone Hsp90 is a potential target for the development of drugs against fungal pathogens. Here the authors determine the structure of the Hsp90 nucleotide-binding domain from Candida albicans, which they use to design an inhibitor and demonstrate its selectivity for the fungal enzyme, both biochemically and in cells.

Biochemical, Cell Biological, and Genetic Assays to Analyze Amyloid and Prion Aggregation in Yeast

Abstract: Protein aggregates are associated with a variety of debilitating human diseases, but they can have functional roles as well. Both pathological and nonpathological protein aggregates display tremendous diversity, with substantial differences in aggregate size, morphology, and structure. Among the different aggregation types, amyloids are particularly remarkable, because of their high degree of order and their ability to form self-perpetuating conformational states. Amyloids form the structural basis for a group of proteins called prions, which have the ability to generate new phenotypes by a simple switch in protein conformation that does not involve changes in the sequence of the DNA. Although protein aggregates are notoriously difficult to study, recent technological developments and, in particular, the use of yeast prions as model systems, have been very instrumental in understanding fundamental aspects of aggregation. Here, we provide a range of biochemical, cell biological and yeast genetic methods that are currently used in our laboratory to study protein aggregation and the formation of amyloids and prions.

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Abstract: 抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

STRING v11: protein-protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets.

Abstract: Proteins and their functional interactions form the backbone of the cellular machinery. Their connectivity network needs to be considered for the full understanding of biological phenomena, but the available information on protein-protein associations is incomplete and exhibits varying levels of annotation granularity and reliability. The STRING database aims to collect, score and integrate all publicly available sources of protein-protein interaction information, and to complement these with computational predictions. Its goal is to achieve a comprehensive and objective global network, including direct (physical) as well as indirect (functional) interactions. The latest version of STRING (11.0) more than doubles the number of organisms it covers, to 5090. The most important new feature is an option to upload entire, genome-wide datasets as input, allowing users to visualize subsets as interaction networks and to perform gene-set enrichment analysis on the entire input. For the enrichment analysis, STRING implements well-known classification systems such as Gene Ontology and KEGG, but also offers additional, new classification systems based on high-throughput text-mining as well as on a hierarchical clustering of the association network itself. The STRING resource is available online at https://string-db.org/.

Genome engineering using the CRISPR-Cas9 system

Abstract: Targeted nucleases are powerful tools for mediating genome alteration with high precision. The RNA-guided Cas9 nuclease from the microbial clustered regularly interspaced short palindromic repeats (CRISPR) adaptive immune system can be used to facilitate efficient genome engineering in eukaryotic cells by simply specifying a 20-nt targeting sequence within its guide RNA. Here we describe a set of tools for Cas9-mediated genome editing via nonhomologous end joining (NHEJ) or homology-directed repair (HDR) in mammalian cells, as well as generation of modified cell lines for downstream functional studies. To minimize off-target cleavage, we further describe a double-nicking strategy using the Cas9 nickase mutant with paired guide RNAs. This protocol provides experimentally derived guidelines for the selection of target sites, evaluation of cleavage efficiency and analysis of off-target activity. Beginning with target design, gene modifications can be achieved within as little as 1-2 weeks, and modified clonal cell lines can be derived within 2-3 weeks.

A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118; iso-2; iso-3

Abstract: We describe a new computer program, SnpEff, for rapidly categorizing the effects of variants in genome sequences. Once a genome is sequenced, SnpEff annotates variants based on their genomic locations and predicts coding effects. Annotated genomic locations include intronic, untranslated region, upstream, downstream, splice site, or intergenic regions. Coding effects such as synonymous or non-synonymous amino acid replacement, start codon gains or losses, stop codon gains or losses, or frame shifts can be predicted. Here the use of SnpEff is illustrated by annotating ~356,660 candidate SNPs in ~117 Mb unique sequences, representing a substitution rate of ~1/305 nucleotides, between the Drosophila melanogaster w1118; iso-2; iso-3 strain and the reference y1; cn1 bw1 sp1 strain. We show that ~15,842 SNPs are synonymous and ~4,467 SNPs are non-synonymous (N/S ~0.28). The remaining SNPs are in other categories, such as stop codon gains (38 SNPs), stop codon losses (8 SNPs), and start codon gains (297 SNPs) in...

Principles of Neural Science

Abstract: I have developed "tennis elbow" from lugging this book around the past four weeks, but it is worth the pain, the effort, and the aspirin. It is also worth the (relatively speaking) bargain price. Including appendixes, this book contains 894 pages of text. The entire panorama of the neural sciences is surveyed and examined, and it is comprehensive in its scope, from genomes to social behaviors. The editors explicitly state that the book is designed as "an introductory text for students of biology, behavior, and medicine," but it is hard to imagine any audience, interested in any fragment of neuroscience at any level of sophistication, that would not enjoy this book. The editors have done a masterful job of weaving together the biologic, the behavioral, and the clinical sciences into a single tapestry in which everyone from the molecular biologist to the practicing psychiatrist can find and appreciate his or