Category: Publications

Nguyen Ba AN, Yeh BJ, van Dyk D, Davidson AR, Andrews BJ, Weiss EL, Moses AM

Sci Signal 2012 Mar;5(215):rs1

PubMed PMID: 22416277

Abstract

At least 30% of human proteins are thought to contain intrinsically disordered regions, which lack stable structural conformation. Despite lacking enzymatic functions and having few protein domains, disordered regions are functionally important for protein regulation and contain short linear motifs (short peptide sequences involved in protein-protein interactions), but in most disordered regions, the functional amino acid residues remain unknown. We searched for evolutionarily conserved sequences within disordered regions according to the hypothesis that conservation would indicate functional residues. Using a phylogenetic hidden Markov model (phylo-HMM), we made accurate, specific predictions of functional elements in disordered regions even when these elements are only two or three amino acids long. Among the conserved sequences that we identified were previously known and newly identified short linear motifs, and we experimentally verified key examples, including a motif that may mediate interaction between protein kinase Cbk1 and its substrates. We also observed that hub proteins, which interact with many partners in a protein interaction network, are highly enriched in these conserved sequences. Our analysis enabled the systematic identification of the functional residues in disordered regions and suggested that at least 5% of amino acids in disordered regions are important for function.

Lumba S, Tsuchiya Y, Delmas F, Hezky J, Provart NJ, Shi Lu Q, McCourt P, Gazzarrini S

BMC Biol. 2012;10:8

PubMed PMID: 22348746

Abstract

BACKGROUND: The embryonic temporal regulator FUSCA3 (FUS3) plays major roles in the establishment of embryonic leaf identity and the regulation of developmental timing. Loss-of-function mutations of this B3 domain transcription factor result in replacement of cotyledons with leaves and precocious germination, whereas constitutive misexpression causes the conversion of leaves into cotyledon-like organs and delays vegetative and reproductive phase transitions.

RESULTS: Herein we show that activation of FUS3 after germination dampens the expression of genes involved in the biosynthesis and response to the plant hormone ethylene, whereas a loss-of-function fus3 mutant shows many phenotypes consistent with increased ethylene signaling. This FUS3-dependent regulation of ethylene signaling also impinges on timing functions outside embryogenesis. Loss of FUS3 function results in accelerated vegetative phase change, and this is again partially dependent on functional ethylene signaling. This alteration in vegetative phase transition is dependent on both embryonic and vegetative FUS3 function, suggesting that this important transcriptional regulator controls both embryonic and vegetative developmental timing.

CONCLUSION: The results of this study indicate that the embryonic regulator FUS3 not only controls the embryonic-to-vegetative phase transition through hormonal (ABA/GA) regulation but also functions postembryonically to delay vegetative phase transitions by negatively modulating ethylene-regulated gene expression.

McCann HC, Nahal H, Thakur S, Guttman DS

Proc. Natl. Acad. Sci. U.S.A. 2012 Mar;109(11):4215-20

PubMed PMID: 22323605

Abstract

The innate immune system is an ancient and broad-spectrum defense system found in all eukaryotes. The detection of microbial elicitors results in the up-regulation of defense-related genes and the elicitation of inflammatory and apoptotic responses. These innate immune responses are the front-line barrier against disease because they collectively suppress the growth of the vast majority of invading microbes. Despite their critical role, we know remarkably little about the diversity of immune elicitors. To address this paucity, we reasoned that hosts are more likely to evolve recognition to “core” pathogen proteins under strong negative selection for the maintenance of essential cellular functions, whereas repeated exposure to host-defense responses will impose strong positive selective pressure for elicitor diversification to avoid host recognition. Therefore, we hypothesized that novel bacterial elicitors can be identified through these opposing forces of natural selection. We tested this hypothesis by examining the genomes of six bacterial phytopathogens and identifying 56 candidate elicitors that have an excess of positively selected residues in a background of strong negative selection. We show that these positively selected residues are atypically clustered, similar to patterns seen in the few well-characterized elicitors. We then validated selected candidate elicitors by showing that they induce Arabidopsis thaliana innate immunity in functional (virulence suppression) and cellular (callose deposition) assays. These finding provide targets for the study of host-pathogen interactions and applied research into alternative antimicrobial treatments.

Cutter AD, Wang GX, Ai H, Peng Y

Mol. Ecol. 2012 Mar;21(6):1345-59

PubMed PMID: 22320847

Abstract

Molecular hyperdiversity has been documented in viruses, prokaryotes and eukaryotes. Such organisms undermine the assumptions of the infinite-sites mutational model, because multiple mutational events at a site comprise a non-negligible portion of polymorphisms. Moreover, different sampling schemes of individuals from species with subdivided populations can profoundly influence resulting patterns and interpretations of molecular variation. Inspired by molecular hyperdiversity in the nematode Caenorhabditis sp. 5, which exhibits average pairwise differences among synonymous sites of >5% as well as modest population structure, we investigated via coalescent simulation the joint effects of a finite-sites mutation (FSM) process and population subdivision on the variant frequency spectrum. From many demes interconnected through a stepping-stone migration model, we constructed local samples from a single deme, pooled samples from several demes and scattered samples of a single individual from numerous demes. Compared with a single panmictic population at equilibrium, we find that high population mutation rates induce a deficit of rare variants (positive Tajima’s D) under a FSM model. Population structure also induces such a skew for local samples when migration is high and for pooled samples when migration is low. Contrasts of sampling schemes for C. sp. 5 imply high mutational input coupled with high migration. We propose that joint analysis of local, pooled and scattered samples for species with subdivided populations provides a means of improving inference of demographic history, by virtue of the partially distinct patterns of polymorphism that manifest when sequences are analyzed according to differing sampling schemes.

Krogan NT, Ckurshumova W, Marcos D, Caragea AE, Berleth T

New Phytol. 2012 Apr;194(2):391-401

PubMed PMID: 22320407

Abstract

Combinatorial interactions of AUXIN RESPONSE FACTORs (ARFs) and auxin/indole acetic acid (Aux/IAA) proteins through their common domains III and IV regulate auxin responses, but insight into the functions of individual proteins is still limited. As a new tool to explore this regulatory network, we generated a gain-of-function ARF genotype by eliminating domains III and IV from the functionally well-characterized ARF MONOPTEROS(MP)/ARF5. This truncated version of MP, termed MP?, conferred complementing MP activity, but also displayed a number of semi-dominant traits affecting auxin signaling and organ patterning. In MP?, the expression levels of many auxin-inducible genes, as well as rooting properties and vascular tissue abundance, were enhanced. Lateral organs were narrow, pointed and filled with parallel veins. This effect was epistatic over the vascular hypotrophy imposed by certain Aux/IAA mutations. Further, in MP? leaves, failure to turn off the procambium-selecting gene PIN1 led to the early establishment of oversized central procambial domains and very limited subsequent lateral growth of the leaf lamina. We conclude that MP? can selectively uncouple a single ARF from regulation by Aux/IAA proteins and can be used as a genetic tool to probe auxin pathways and explore leaf development.

Lee AH, Hurley B, Felsensteiner C, Yea C, Ckurshumova W, Bartetzko V, Wang PW, Quach V, Lewis JD, Liu YC, Börnke F, Angers S, Wilde A, Guttman DS, Desveaux D

PLoS Pathog. 2012 Feb;8(2):e1002523

PubMed PMID: 22319451

Abstract

The eukaryotic cytoskeleton is essential for structural support and intracellular transport, and is therefore a common target of animal pathogens. However, no phytopathogenic effector has yet been demonstrated to specifically target the plant cytoskeleton. Here we show that the Pseudomonas syringae type III secreted effector HopZ1a interacts with tubulin and polymerized microtubules. We demonstrate that HopZ1a is an acetyltransferase activated by the eukaryotic co-factor phytic acid. Activated HopZ1a acetylates itself and tubulin. The conserved autoacetylation site of the YopJ / HopZ superfamily, K289, plays a critical role in both the avirulence and virulence function of HopZ1a. Furthermore, HopZ1a requires its acetyltransferase activity to cause a dramatic decrease in Arabidopsis thaliana microtubule networks, disrupt the plant secretory pathway and suppress cell wall-mediated defense. Together, this study supports the hypothesis that HopZ1a promotes virulence through cytoskeletal and secretory disruption.

Weadick CJ, Chang BS

Mol. Biol. Evol. 2012 May;29(5):1297-300

PubMed PMID: 22319160

Abstract

Maximum likelihood codon substitution models have proven useful for studying when and how protein function evolves, but they have recently been criticized on a number of fronts. The strengths and weaknesses of such methods must therefore be identified and improved upon. Here, using simulations, we show that the Clade model C versus M1a test for functional divergence among clades is prone to false positives under simple evolutionary conditions. We then propose a new null model (M2a_rel) that better accounts for among-site variation in selective constraint. We show that the revised test has an improved false-positive rate and good power. Applying this test to previously analyzed data sets of primate ribonucleases and mammalian rhodopsins reveals that some conclusions may have been misled by the original method. The improved test should prove useful for identifying patterns of divergence in selective constraint among paralogous gene families and among orthologs from ecologically divergent species.

Lindsey BW, Darabie A, Tropepe V

J. Comp. Neurol. 2012 Jul;520(10):2275-316

PubMed PMID: 22318736

Abstract

A central goal of adult neurogenesis research is to characterize the cellular constituents of a neurogenic niche and to understand how these cells regulate the production of new neurons. Because the generation of adult-born neurons may be tightly coupled to their functional requirement, the organization and output of neurogenic niches may vary across different regions of the brain or between species. We have undertaken a comparative study of six (D, Vd, Vv, Dm, Dl, Ppa) periventricular zones (PVZs) harboring proliferative cells present in the adult forebrain of the zebrafish (Danio rerio), a species known to possess widespread neurogenesis throughout life. Using electron microscopy, we have documented for the first time the detailed cytoarchitecture of these zones, and propose a model of the cellular composition of pallial and subpallial PVZs, as well as a classification scheme for identifying morphologically distinct cell types. Immunolabeling of resin-embedded tissue confirmed the phenotype of three constitutively proliferating (bromodeoxyuridine [BrdU]+) cell populations, including a radial glial-like (type IIa) cell immunopositive for both S100? and glutamine synthetase (GS). Our data revealed rostrocaudal differences in the density of distinct proliferative populations, and cumulative labeling studies suggested that the cell cycle kinetics of these populations are not uniform between PVZs. Although the peak numbers of differentiated neurons were generated after ~2 weeks among most PVZs, niche-specific decline in the number of newborn neurons in some regions occurred after 4 weeks. Our data suggest that the cytoarchitecture of neurogenic niches and the tempo of neuronal production are regionally distinct in the adult zebrafish forebrain.

Chiu RS, Nahal H, Provart NJ, Gazzarrini S

BMC Plant Biol. 2012;12:15

PubMed PMID: 22279962

Abstract

BACKGROUND: Imbibed seeds integrate environmental and endogenous signals to break dormancy and initiate growth under optimal conditions. Seed maturation plays an important role in determining the survival of germinating seeds, for example one of the roles of dormancy is to stagger germination to prevent mass growth under suboptimal conditions. The B3-domain transcription factor FUSCA3 (FUS3) is a master regulator of seed development and an important node in hormonal interaction networks in Arabidopsis thaliana. Its function has been mainly characterized during embryonic development, where FUS3 is highly expressed to promote seed maturation and dormancy by regulating ABA/GA levels.

RESULTS: In this study, we present evidence for a role of FUS3 in delaying seed germination at supraoptimal temperatures that would be lethal for the developing seedlings. During seed imbibition at supraoptimal temperature, the FUS3 promoter is reactivated and induces de novo synthesis of FUS3 mRNA, followed by FUS3 protein accumulation. Genetic analysis shows that FUS3 contributes to the delay of seed germination at high temperature. Unlike WT, seeds overexpressing FUS3 (ML1:FUS3-GFP) during imbibition are hypersensitive to high temperature and do not germinate, however, they can fully germinate after recovery at control temperature reaching 90% seedling survival. ML1:FUS3-GFP hypersensitivity to high temperature can be partly recovered in the presence of fluridone, an inhibitor of ABA biosynthesis, suggesting this hypersensitivity is due in part to higher ABA level in this mutant. Transcriptomic analysis shows that WT seeds imbibed at supraoptimal temperature activate seed-specific genes and ABA biosynthetic and signaling genes, while inhibiting genes that promote germination and growth, such as GA biosynthetic and signaling genes.

CONCLUSION: In this study, we have uncovered a novel function for the master regulator of seed maturation, FUS3, in delaying germination at supraoptimal temperature. Physiologically, this is important since delaying germination has a protective role at high temperature. Transcriptomic analysis of seeds imbibed at supraoptimal temperature reveal that a complex program is in place, which involves not only the regulation of heat and dehydration response genes to adjust cellular functions, but also the activation of seed-specific programs and the inhibition of germination-promoting programs to delay germination.

Lee SJ, Lee J, Li KK, Holland D, Maughan H, Guttman DS, Yusta B, Drucker DJ

Endocrinology 2012 Mar;153(3):1141-51

PubMed PMID: 22253424

Abstract

Exogenous glucagon-like peptide-2 receptor (GLP-2R) activation elicits proliferative and cytoprotective responses in the gastrointestinal mucosa and ameliorates experimental small and large bowel gut injury. Nevertheless, the essential physiological role(s) of the endogenous GLP-2R remain poorly understood. We studied the importance of the GLP-2R for gut growth, epithelial cell lineage allocation, the response to mucosal injury, and host-bacterial interactions in Glp2r(-/-) and littermate control Glp2r(+/+) mice. Glp2r(-/-) mice exhibit normal somatic growth and preserved small and large bowel responses to IGF-I and keratinocyte growth factor. However, Glp2r(-/-) mice failed to up-regulate intestinal epithelial c-fos expression in response to acute GLP-2 administration and do not exhibit changes in small bowel conductance or small or large bowel growth after administration of GLP-2R agonists. The crypt and villus compartment and the numbers and localization of Paneth, enteroendocrine, and goblet cells were comparable in Glp2r(+/+) vs. Glp2r(-/-) mice. Although the severity and extent of colonic mucosal injury in response to 3% oral dextran sulfate was similar across Glp2r genotypes, Glp2r(-/-) mice exhibited significantly increased morbidity and mortality and increased bacterial translocation after induction of enteritis with indomethacin and enhanced mucosal injury in response to irinotecan. Moreover, bacterial colonization of the small bowel was significantly increased, expression of Paneth cell antimicrobial gene products was reduced, and mucosal bactericidal activity was impaired in Glp2r(-/-) mice. Although the Glp2r is dispensable for gut development and the response to colonic injury, Glp2r(-/-) mice exhibit enhanced sensitivity to small bowel injury, and abnormal host-bacterial interactions in the small bowel.