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1.
J Cell Sci ; 135(7)2022 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-35194638

RESUMEN

Multicellular organisms use dedicator of cytokinesis (DOCK) family guanine nucleotide exchange factors (GEFs) to activate Rac/Rho-of-plants small GTPases and coordinate cell shape change. In developing tissues, DOCK signals integrate cell-cell interactions with cytoskeleton remodeling, and the GEFs cluster reversibly at specific organelle surfaces to orchestrate cytoskeletal reorganization. The domain organizations among DOCK orthologs are diverse, and the mechanisms of localization control are poorly understood. Here, we use combinations of transgene complementation and live-cell imaging assays to uncover an evolutionarily conserved and essential localization determinant in the DOCK-GEF named SPIKE1. The SPIKE1-DHR3 domain is sufficient for organelle association in vivo, and displays a complicated lipid-binding selectivity for both phospholipid head groups and fatty acid chain saturation. SPIKE1-DHR3 is predicted to adopt a C2-domain structure and functions as part of a tandem C2 array that enables reversible clustering at the cell apex. This work provides mechanistic insight into how DOCK GEFs sense compositional and biophysical membrane properties at the interface of two organelle systems.


Asunto(s)
Factores de Intercambio de Guanina Nucleótido , Proteínas de Unión al GTP Monoméricas , Dominios C2 , Citocinesis , Factores de Intercambio de Guanina Nucleótido/metabolismo , Proteínas de Unión al GTP Monoméricas/metabolismo , Orgánulos/metabolismo
2.
Plant Physiol ; 171(1): 42-61, 2016 05.
Artículo en Inglés | MEDLINE | ID: mdl-26983993

RESUMEN

Plant meristems, like animal stem cell niches, maintain a pool of multipotent, undifferentiated cells that divide and differentiate to give rise to organs. In Arabidopsis (Arabidopsis thaliana), the carpel margin meristem is a vital meristematic structure that generates ovules from the medial domain of the gynoecium, the female floral reproductive structure. The molecular mechanisms that specify this meristematic region and regulate its organogenic potential are poorly understood. Here, we present a novel approach to analyze the transcriptional signature of the medial domain of the Arabidopsis gynoecium, highlighting the developmental stages that immediately proceed ovule initiation, the earliest stages of seed development. Using a floral synchronization system and a SHATTERPROOF2 (SHP2) domain-specific reporter, paired with FACS and RNA sequencing, we assayed the transcriptome of the gynoecial medial domain with temporal and spatial precision. This analysis reveals a set of genes that are differentially expressed within the SHP2 expression domain, including genes that have been shown previously to function during the development of medial domain-derived structures, including the ovules, thus validating our approach. Global analyses of the transcriptomic data set indicate a similarity of the pSHP2-expressing cell population to previously characterized meristematic domains, further supporting the meristematic nature of this gynoecial tissue. Our method identifies additional genes including novel isoforms, cis-natural antisense transcripts, and a previously unrecognized member of the REPRODUCTIVE MERISTEM family of transcriptional regulators that are potential novel regulators of medial domain development. This data set provides genome-wide transcriptional insight into the development of the carpel margin meristem in Arabidopsis.


Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Proteínas de Dominio MADS/genética , Meristema/genética , Transcriptoma , Arabidopsis/anatomía & histología , Proteínas de Arabidopsis/aislamiento & purificación , Secuencia de Bases , Hidrato de Cloral , ADN sin Sentido , Flores/genética , Genoma de Planta , Hibridación in Situ , Ácidos Indolacéticos/farmacología , Proteínas de Dominio MADS/aislamiento & purificación , Meristema/crecimiento & desarrollo , Meristema/metabolismo , Microscopía Confocal , Óvulo Vegetal/citología , Óvulo Vegetal/crecimiento & desarrollo , Óvulo Vegetal/metabolismo , Isoformas de Proteínas , Protoplastos , ARN de Planta/química , ARN de Planta/aislamiento & purificación , Semillas/crecimiento & desarrollo , Alineación de Secuencia , Factores de Transcripción , Activación Transcripcional
3.
Plant Cell ; 23(11): 3961-73, 2011 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-22108406

RESUMEN

The effects of auxins on plant growth and development have been known for more than 100 years, yet our understanding of how plants synthesize this essential plant hormone is still fragmentary at best. Gene loss- and gain-of-function studies have conclusively implicated three gene families, CYTOCHROME P450 79B2/B3 (CYP79B2/B3), YUCCA (YUC), and TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1/TRYPTOPHAN AMINOTRANSFERASE-RELATED (TAA1/TAR), in the production of this hormone in the reference plant Arabidopsis thaliana. Each of these three gene families is believed to represent independent routes of auxin biosynthesis. Using a combination of pharmacological, genetic, and biochemical approaches, we examined the possible relationships between the auxin biosynthetic pathways defined by these three gene families. Our findings clearly indicate that TAA1/TARs and YUCs function in a common linear biosynthetic pathway that is genetically distinct from the CYP79B2/B3 route. In the redefined TAA1-YUC auxin biosynthetic pathway, TAA1/TARs are required for the production of indole-3-pyruvic acid (IPyA) from Trp, whereas YUCs are likely to function downstream. These results, together with the extensive genetic analysis of four pyruvate decarboxylases, the putative downstream components of the TAA1 pathway, strongly suggest that the enzymatic reactions involved in indole-3-acetic acid (IAA) production via IPyA are different than those previously postulated, and a new and testable model for how IAA is produced in plants is needed.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Ácidos Indolacéticos/metabolismo , Indoles/metabolismo , Oxigenasas/genética , Oxigenasas/metabolismo , Proteínas de Arabidopsis/genética , Sistema Enzimático del Citocromo P-450/genética , Sistema Enzimático del Citocromo P-450/metabolismo , Técnicas de Silenciamiento del Gen , Triptófano-Transaminasa/genética , Triptófano-Transaminasa/metabolismo
4.
J Exp Bot ; 63(5): 2231-41, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22238449

RESUMEN

As part of a continuing effort to elucidate mechanisms that regulate the magnitude of ethylene signalling, an Arabidopsis mutant with an enhanced ethylene response was identified. Subsequent characterization of this loss-of-function mutant revealed severe hypocotyl shortening in the presence of saturating ethylene along with increased expression in leaves of a subset of ethylene-responsive genes. It was subsequently determined by map-based cloning that the mutant (sar1-7) represents a loss-of-function mutation in the previously described nucleoporin AtNUP160 (At1g33410, SAR1). In support of previously reported results, the sar1-7 mutant partially restored auxin responsiveness to roots of an rce1 loss-of-function mutant, indicating that AtNUP160/SAR1 is required for proper expression of factors responsible for the repression of auxin signalling. Analysis of arf7-1/sar1-7 and arf19-1/sar1-7 double mutants revealed that mutations affecting either ARF7 or ARF19 function almost fully blocked manifestation of the sar1-7-dependent ethylene hypersensitivity phenotype, suggesting that ARF7- and ARF19-mediated auxin signalling is responsible for regulating the magnitude of and/or competence for the ethylene response in Arabidopsis etiolated hypocotyls. Consistent with this, addition of auxin to ethylene-treated seedlings resulted in severe hypocotyl shortening, reminiscent of that seen for other eer (enhanced ethylene response) mutants, suggesting that auxin functions in part synergistically with ethylene to control hypocotyl elongation and other ethylene-dependent phenomena.


Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Etilenos/metabolismo , Ácidos Indolacéticos/metabolismo , Proteínas R-SNARE/genética , Transducción de Señal/fisiología , Arabidopsis/crecimiento & desarrollo , Arabidopsis/fisiología , Proteínas de Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas/fisiología , Hipocótilo/genética , Hipocótilo/crecimiento & desarrollo , Hipocótilo/fisiología , Mutación , Proteínas de Complejo Poro Nuclear/genética , Proteínas de Complejo Poro Nuclear/metabolismo , Fenotipo , Hojas de la Planta/genética , Hojas de la Planta/crecimiento & desarrollo , Hojas de la Planta/fisiología , Raíces de Plantas/genética , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/fisiología , Plantas Modificadas Genéticamente , Mapeo de Interacción de Proteínas , Proteínas R-SNARE/metabolismo , Plantones/genética , Plantones/crecimiento & desarrollo , Plantones/fisiología , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
5.
Plant Biotechnol (Tokyo) ; 39(4): 367-379, 2022 Dec 25.
Artículo en Inglés | MEDLINE | ID: mdl-37283612

RESUMEN

It is estimated that multiple sclerosis (MS) affects over 2.8 million people worldwide, with a prevalence that is expected to continue growing over time. Unfortunately, there is no cure for this autoimmune disease. For several decades, antigen-specific treatments have been used in animal models of experimental autoimmune encephalomyelitis (EAE) to demonstrate their potential for suppressing autoimmune responses. Successes with preventing and limiting ongoing MS disease have been documented using a wide variety of myelin proteins, peptides, autoantigen-conjugates, and mimics when administered by various routes. While those successes were not translatable in the clinic, we have learned a great deal about the roadblocks and hurdles that must be addressed if such therapies are to be useful. Reovirus sigma1 protein (pσ1) is an attachment protein that allows the virus to target M cells with high affinity. Previous studies showed that autoantigens tethered to pσ1 delivered potent tolerogenic signals and diminished autoimmunity following therapeutic intervention. In this proof-of-concept study, we expressed a model multi-epitope autoantigen (human myelin basic protein, MBP) fused to pσ1 in soybean seeds. The expression of chimeric MBP-pσ1 was stable over multiple generations and formed the necessary multimeric structures required for binding to target cells. When administered to SJL mice prophylactically as an oral therapeutic, soymilk formulations containing MBP-pσ1 delayed the onset of clinical EAE and significantly reduced developing disease. These results demonstrate the practicality of soybean as a host for producing and formulating immune-modulating therapies to treat autoimmune diseases.

6.
Surg Neurol Int ; 13: 465, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-36324940

RESUMEN

Background: Due to the SARS-CoV-2 virus pandemic and its rapid spread worldwide, an early and effective detection strategy was the nasopharyngeal reverse transcription polymerase swab tests, a procedure still performed today. A relatively safe procedure when done correctly, however, one of the rare complications reported in the literature includes a cerebrospinal fluid (CSF) leak. Case Description: A 69-year-old female patient presented to the emergency department with clear fluid rhinorrhea, clinically diagnosed with a CSF fistula after a SARS-CoV-2 nasopharyngeal swab. Resulting computed tomography and magnetic resonance images did not report any abnormalities; however, persistence of clear fluid rhinorrhea obligated pharmacological treatment without resolution, requiring insertion of a lumbar catheter to achieve clinical resolution. Conclusion: It is essential to train staff to correctly administer nasopharyngeal swabs and thus reduce the rate of complications, as well as early recognition of symptoms and signs of CSF fistula.

7.
Plant J ; 55(3): 467-77, 2008 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-18429939

RESUMEN

An Arabidopsis mutant, eer5-1, which has an enhanced ethylene response in etiolated seedlings, including hypersensitivity and extreme exaggeration of response to ethylene, was isolated and characterized. As with other identified eer mutants, the enhanced response phenotype of eer5-1 was correlated with failure to induce appropriately a subset of ethylene-regulated genes, suggesting that proper ethylene-responsive gene expression is necessary for resetting the ethylene response pathway. eer5-1 represents a mutation that causes an amino acid substitution in a previously uncharacterized gene, which encodes a protein with a PAM [proteasome COP9 initiation factor (PCI/PINT)-associated module] domain similar to those found in components of the COP9 signalosome (CSN). Genetic analysis shows that manifestation of the eer5 mutant phenotype is solely dependent on ethylene signaling, as the ein2-5 eer5-1 double mutant was indistinguishable from ein2-5 in the presence of saturating ethylene concentrations. In contrast, the ein3-1 eer5-1 double mutant displayed characteristics of an enhanced ethylene response, and this suggests that EER5 regulates ethylene signaling independently of EIN3. Analysis of the EER5 protein indicates that it interacts with the C-terminus of EIN2 and with the CSN, suggesting that EER5 serves as a bridge between EIN2 and the modification or degradation of target proteins, including a proposed group of transcriptional repressors, as part of a resetting mechanism during or following ethylene signaling.


Asunto(s)
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/fisiología , Arabidopsis/metabolismo , Etilenos/metabolismo , Transducción de Señal , Arabidopsis/efectos de los fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Etilenos/farmacología , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Genotipo , Hipocótilo/efectos de los fármacos , Hipocótilo/genética , Mutación , Fenotipo , Complejo de la Endopetidasa Proteasomal/genética , Mapeo de Interacción de Proteínas , Estructura Terciaria de Proteína , Proteínas de Unión al ARN , Receptores de Superficie Celular/química , Receptores de Superficie Celular/metabolismo
8.
Dev Cell ; 47(3): 306-318.e5, 2018 11 05.
Artículo en Inglés | MEDLINE | ID: mdl-30415657

RESUMEN

Auxin is a major phytohormone that controls numerous aspects of plant development and coordinates plant responses to the environment. Morphogenic gradients of auxin govern cell fate decisions and underlie plant phenotypic plasticity. Polar auxin transport plays a central role in auxin maxima generation. The discovery of the exquisite spatiotemporal expression patterns of auxin biosynthesis genes of the WEI8/TAR and YUC families suggested that local auxin production may contribute to the formation of auxin maxima. Herein, we systematically addressed the role of local auxin biosynthesis in plant development and responses to the stress phytohormone ethylene by manipulating spatiotemporal patterns of WEI8. Our study revealed that local auxin biosynthesis and transport act synergistically and are individually dispensable for root meristem maintenance. In contrast, flower fertility and root responses to ethylene require local auxin production that cannot be fully compensated for by transport in the generation of morphogenic auxin maxima.


Asunto(s)
Ácidos Indolacéticos/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Raíces de Plantas/embriología , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Etilenos/metabolismo , Flores/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Meristema/embriología , Meristema/metabolismo , Morfogénesis , Oxigenasas , Desarrollo de la Planta/genética , Reguladores del Crecimiento de las Plantas/genética , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Análisis Espacio-Temporal
9.
Curr Biol ; 23(24): 2506-12, 2013 Dec 16.
Artículo en Inglés | MEDLINE | ID: mdl-24291089

RESUMEN

Establishment of the embryonic axis foreshadows the main body axis of adults both in plants and in animals, but underlying mechanisms are considered distinct. Plants utilize directional, cell-to-cell transport of the growth hormone auxin to generate an asymmetric auxin response that specifies the embryonic apical-basal axis. The auxin flow directionality depends on the polarized subcellular localization of PIN-FORMED (PIN) auxin transporters. It remains unknown which mechanisms and spatial cues guide cell polarization and axis orientation in early embryos. Herein, we provide conceptually novel insights into the formation of embryonic axis in Arabidopsis by identifying a crucial role of localized tryptophan-dependent auxin biosynthesis. Local auxin production at the base of young embryos and the accompanying PIN7-mediated auxin flow toward the proembryo are required for the apical auxin response maximum and the specification of apical embryonic structures. Later in embryogenesis, the precisely timed onset of localized apical auxin biosynthesis mediates PIN1 polarization, basal auxin response maximum, and specification of the root pole. Thus, the tight spatiotemporal control of distinct local auxin sources provides a necessary, non-cell-autonomous trigger for the coordinated cell polarization and subsequent apical-basal axis orientation during embryogenesis and, presumably, also for other polarization events during postembryonic plant life.


Asunto(s)
Arabidopsis/embriología , Ácidos Indolacéticos/metabolismo , Reguladores del Crecimiento de las Plantas/fisiología , Semillas/crecimiento & desarrollo , Proteínas de Arabidopsis/metabolismo , Tipificación del Cuerpo/efectos de los fármacos , Ácidos Indolacéticos/farmacología , Proteínas de Transporte de Membrana/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Reguladores del Crecimiento de las Plantas/farmacología , Transporte de Proteínas , Semillas/efectos de los fármacos
11.
J Exp Bot ; 58(10): 2627-39, 2007.
Artículo en Inglés | MEDLINE | ID: mdl-17526916

RESUMEN

eer4 was isolated as an Arabidopsis mutant with an extreme response to ethylene in dark-grown seedlings that was also found to have partial ethylene insensitivity at the level of ethylene-dependent gene expression, including ERF1. Subsequent cloning of eer4 revealed an inappropriate stop codon in a previously uncharacterized TFIID-interacting transcription factor homologous to human TAF12 and yeast TAF61. Genetic and pharmacological analysis demonstrated that the eer4 phenotype is strictly ethylene dependent in seedlings, yet a double mutant with the partially ethylene-insensitive Arabidopsis mutant, ein3-1, had restored ethylene responsiveness, indicating that eer4 also regulates a previously unknown resetting or dampening mechanism for the ethylene signalling pathway. Consistent with the absolute requirement of EER4 for ERF1 expression, biochemical analysis showed that EER4 is localized to the nucleus where it probably recruits EIN3 and probably other transcription factors along with components of the TFIID complex for expression of a subset of genes required for either manifestation or subsequent dampening of the response to ethylene.


Asunto(s)
Proteínas de Arabidopsis/fisiología , Arabidopsis/metabolismo , Etilenos/metabolismo , Factores de Terminación de Péptidos/metabolismo , Factor de Transcripción TFIID/fisiología , Secuencia de Aminoácidos , Arabidopsis/efectos de los fármacos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Codón sin Sentido , Oscuridad , Etilenos/farmacología , Proteínas Fluorescentes Verdes/análisis , Modelos Biológicos , Datos de Secuencia Molecular , Fenotipo , Análisis de Secuencia de Proteína , Transducción de Señal/genética , Factor de Transcripción TFIID/genética , Factor de Transcripción TFIID/metabolismo
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