Firmly Planted, Always Moving Raikhel, Natasha V
Annual review of plant biology,
04/2017, Letnik:
68, Številka:
1
Journal Article
Recenzirano
Odprti dostop
I was a budding pianist immersed in music in Leningrad, in the Soviet Union (now Saint Petersburg, Russia), when I started over, giving up sheet music for the study of ciliates. In a second ...starting-over story, I emigrated to the United States, where I switched to studying carbohydrate-binding plant lectin proteins, dissecting plant vesicular trafficking, and isolating novel glycosyltransferases responsible for making cell wall polysaccharides. I track my journey as a plant biologist from student to principal investigator to founding director of the Center for Plant Cell Biology and then director of the Institute for Integrative Genome Biology at the University of California, Riverside. I discuss implementing a new vision as the first and (so far) only female editor in chief of
Plant Physiology
, as well as how my laboratory helped develop chemical genomics tools to study the functions of essential plant proteins. Always wanting to give back what I received, I discuss my present efforts to develop female scientist leadership in Chinese universities and a constant theme throughout my life: a love of art and travel.
Since the introduction of chemical genomics to plant biology as a tool for basic research, the field has advanced significantly. There are now examples of important basic discoveries that demonstrate ...the power and untapped potential of this approach. Given the combination of protein and small-molecule complexity, new phenotypes can be described through the perturbation of cellular functions that can be linked to growth and developmental phenotypes. There are now clear examples of overcoming functional redundancy in plants to dissect molecular mechanisms or critical pathways such as hormone signaling and dynamic intracellular processes. Owing to ongoing advances, including more sophisticated high-content screening and rapid approaches for target identification, the field is beginning to move forward. However, there are also challenges to improve automation, imaging, and analysis and provide chemical biology resources to the broader plant biology community.
Translation inhibition is a major but poorly understood mode of action of microRNAs (miRNAs) in plants and animals. In particular, the subcellular location where this process takes place is unknown. ...Here, we show that the translation inhibition, but not the mRNA cleavage activity, of Arabidopsis miRNAs requires ALTERED MERISTEM PROGRAM1 (AMP1). AMP1 encodes an integral membrane protein associated with endoplasmic reticulum (ER) and ARGONAUTE1, the miRNA effector and a peripheral ER membrane protein. Large differences in polysome association of miRNA target RNAs are found between wild-type and the amp1 mutant for membrane-bound, but not total, polysomes. This, together with AMP1-independent recruitment of miRNA target transcripts to membrane fractions, shows that miRNAs inhibit the translation of target RNAs on the ER. This study demonstrates that translation inhibition is an important activity of plant miRNAs, reveals the subcellular location of this activity, and uncovers a previously unknown function of the ER.
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•miRNA-mediated translation inhibition, but not translation in general, requires AMP1•AMP1 is dispensable for miRNA-mediated target RNA cleavage•AMP1 is an integral membrane protein that is localized to the rough ER•AMP1 inhibits the loading of miRNA target transcripts onto membrane-bound polysomes
The integral ER membrane protein AMP1 is required for translational repression of miRNA target transcripts, indicating that the ER is an important locus of translational repression.
The endomembrane system is a complex and dynamic intracellular trafficking network. It is very challenging to track individual vesicles and their cargos in real time; however, affinity purification ...allows vesicles to be isolated in their natural state so that their constituent proteins can be identified. Pioneering this approach in plants, we isolated the SYP61 trans-Golgi network compartment and carried out a comprehensive proteomic analysis of its contents with only minimal interference from other organelles. The proteome of SYP61 revealed the association of proteins of unknown function that have previously not been ascribed to this compartment. We identified a complete SYP61 SNARE complex, including regulatory proteins and validated the proteome data by showing that several of these proteins associated with SYP61 in planta. We further identified the SYP121-complex and cellulose synthases, sug- gesting that SYP61 plays a role in the exocytic trafficking and the transport of cell wall components to the plasma membrane. The presence of proteins of unknown function in the SYP61 proteome including ECHIDNA offers the opportunity to identify novel trafficking components and cargos. The affinity purification of plant vesicles in their natural state provides a basis for further analysis and dissection of complex endomembrane networks. The approach is widely applicable and can afford the study of several vesicle populations in plants, which can be compared with the SYP61 vesicle proteome.
Upstream ORFs are elements found in the 5′-leader sequences of specific mRNAs that modulate the translation of downstream ORFs encoding major gene products. In Arabidopsis , the translational control ...of auxin response factors (ARFs) by upstream ORFs has been proposed as a regulatory mechanism required to respond properly to complex auxin-signaling inputs. In this study, we identify and characterize the aberrant auxin responses in specific ribosomal protein mutants in which multiple ARF transcription factors are simultaneously repressed at the translational level. This characteristic lends itself to the use of these mutants as genetic tools to bypass the genetic redundancy among members of the ARF family in Arabidopsis . Using this approach, we were able to assign unique functions for ARF2, ARF3, and ARF6 in plant development.
The endomembrane system is an interconnected network required to establish signal transduction, cell polarity, and cell shape in response to developmental or environmental stimuli. In the model plant ...Arabidopsis thaliana, there are numerous markers to visualize polarly localized plasma membrane proteins utilizing endomembrane trafficking. Previous studies have shown that the large ARF-GEF GNOM plays a key role in the establishment of basal (rootward) polarity, whereas the apically (shootward) polarized membrane proteins undergo sorting via different routes. However, the mechanism that maintains apical polarity is largely unknown. Here, we used a chemical genomic approach and identified the compound endosidin 16 (ES16), which perturbed apically localized plasma membrane proteins without affecting basal polarity. We demonstrated that ES16 is an inhibitor for recycling of apical, lateral, and nonpolar plasma membrane proteins as well as biosynthetic secretion, leaving the basal proteins as the only exceptions not subject to ES16 inhibition. Further evidence from pharmaceutical and genetic data revealed that ES16 effects are mediated through the regulation of small GTPase RabA proteins and that RabA GTPases work in concert with the BIG clade ARF-GEF to modulate the nonbasal trafficking. Our results reveal that ES16 defines a distinct pathway for endomembrane sorting routes and is essential for the establishment of cell polarity.
Vacuoles play central roles in plant growth, development, and stress responses. To better understand vacuole function and biogenesis we have characterized the vegetative vacuolar proteome from ...Arabidopsis thaliana. Vacuoles were isolated from protoplasts derived from rosette leaf tissue. Total purified vacuolar proteins were then subjected either to multidimensional liquid chromatography/tandem mass spectrometry or to one-dimensional SDS-PAGE coupled with nano-liquid chromatography/tandem mass spectrometry (nano-LC MS/MS). To ensure maximum coverage of the proteome, a tonoplast-enriched fraction was also analyzed separately by one-dimensional SDS-PAGE followed by nano-LC MS/MS. Cumulatively, 402 proteins were identified. The sensitivity of our analyses is indicated by the high coverage of membrane proteins. Eleven of the twelve known vacuolar-ATPase subunits were identified. Here, we present evidence of four tonoplast-localized soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), representing each of the four groups of SNARE proteins necessary for membrane fusion. In addition, potential cargo of the N- and C-terminal propeptide sorting pathways, association of the vacuole with the cytoskeleton, and the vacuolar localization of 89 proteins of unknown function are identified. A detailed analysis of these proteins and their roles in vacuole function and biogenesis is presented.
Plant vacuoles are essential organelles for plant growth and development, and have multiple functions. Vacuoles are highly dynamic and pleiomorphic, and their size varies depending on the cell type ...and growth conditions. Vacuoles compartmentalize different cellular components such as proteins, sugars, ions and other secondary metabolites and play critical roles in plants response to different biotic/abiotic signaling pathways. In this review, we will summarize the patterns of changes in vacuole morphology in certain cell types, our understanding of the mechanisms of plant vacuole biogenesis, and the role of SNAREs and Rab GTPases in vacuolar trafficking.
The exocyst complex regulates the last steps of exocytosis, which is essential to organisms across kingdoms. In humans, its dysfunction is correlated with several significant diseases, such as ...diabetes and cancer progression. Investigation of the dynamic regulation of the evolutionarily conserved exocyst-related processes using mutants in genetically tractable organisms such as Arabidopsis thaliana is limited by the lethality or the severity of phenotypes. We discovered that the small molecule Endosidin2 (ES2) binds to the EXO70 (exocyst component of 70 kDa) subunit of the exocyst complex, resulting in inhibition of exocytosis and endosomal recycling in both plant and human cells and enhancement of plant vacuolar trafficking. An EXO70 protein with a C-terminal truncation results in dominant ES2 resistance, uncovering possible distinct regulatory roles for the N terminus of the protein. This study not only provides a valuable tool in studying exocytosis regulation but also offers a potentially new target for drugs aimed at addressing human disease.
Significance Auxin is an important phytohormone that regulates almost all aspects of plant growth and development. Ribosome proteins serve as translational regulators of auxin response. This work ...highlights the significance of lipid metabolism in the downstream of ribosome protein function to link auxin-regulated developmental patterning with endomembrane trafficking. Our work sheds light on a novel link between auxin signaling, ribosome function, and lipid metabolism.
The vacuole is the most prominent compartment in plant cells and is important for ion and protein storage. In our effort to search for key regulators in the plant vacuole sorting pathway, ribosomal large subunit 4 ( rpl4d ) was identified as a translational mutant defective in both vacuole trafficking and normal development. Polysome profiling of the rpl4d mutant showed reduction in polysome-bound mRNA compared with wild-type, but no significant change in the general mRNA distribution pattern. Ribsomal profiling data indicated that genes in the lipid metabolism pathways were translationally down-regulated in the rpl4d mutant. Live imaging studies by Nile red staining suggested that both polar and nonpolar lipid accumulation was reduced in meristem tissues of rpl4d mutants. Pharmacological evidence showed that sterol and sphingolipid biosynthetic inhibitors can phenocopy the defects of the rpl4d mutant, including an altered vacuole trafficking pattern. Genetic evidence from lipid biosynthetic mutants indicates that alteration in the metabolism of either sterol or sphingolipid biosynthesis resulted in vacuole trafficking defects, similar to the rpl4d mutant. Tissue-specific complementation with key enzymes from lipid biosynthesis pathways can partially rescue both vacuole trafficking and auxin-related developmental defects in the rpl4d mutant. These results indicate that lipid metabolism modulates auxin-mediated tissue differentiation and endomembrane trafficking pathways downstream of ribosomal protein function.
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