Rory Degenhardt

Arabidopsis Ribosomal Proteins RPL23aA and RPL23aB Are Differentially Targeted to the Nucleolus and Are Disparately Required for Normal Development

Protein synthesis is catalyzed by the ribosome, a two-subunit enzyme comprised of four ribosomal RNAs and, in Arabidopsis (Arabidopsis thaliana), 81 ribosomal proteins (r-proteins). Plant r-protein genes exist as families of multiple expressed members, yet only one r-protein from each family is incorporated into any given ribosome, suggesting that many r-protein genes may be functionally redundant or development/tissue/stress specific. Here, we characterized the localization and gene-silencing phenotypes of a large subunit r-protein family, RPL23a, containing two expressed genes (RPL23aA and RPL23aB). Live cell imaging of RPL23aA and RPL23aB in tobacco with a C-terminal fluorescent-protein tag demonstrated that both isoforms accumulated in the nucleolus; however, only RPL23aA was targeted to the nucleolus with an N-terminal fluorescent protein tag, suggesting divergence in targeting efficiency of localization signals. Independent knockdowns of endogenous RPL23aA and RPL23aB transcript levels using RNA interference determined that an RPL23aB knockdown did not alter plant growth or development. Conversely, a knockdown of RPL23aA produced a pleiotropic phenotype characterized by growth retardation, irregular leaf and root morphology, abnormal phyllotaxy and vasculature, and loss of apical dominance. Comparison to other mutants suggests that the phenotype results from reduced ribosome biogenesis, and we postulate a link between biogenesis, microRNA-target degradation, and maintenance of auxin homeostasis. An additional RNA interference construct that coordinately silenced both RPL23aA and RPL23aB demonstrated that this family is essential for viability.

Transcript profiling demonstrates absence of dosage compensation in Arabidopsis following loss of a single RPL23a paralog

Translation of nucleus-encoded messages in plants is conducted by the cytoplasmic ribosome, an enzyme that is comprised of two RNA/protein subunits. In Arabidopsis thaliana, the 81 different ribosomal proteins (r-proteins) of the cytosolic ribosome belong to gene families with multiple expressed members. Given that ribosomes generally contain only one copy of each r-protein, regulatory mechanisms must exist to ensure their stoichiometric accumulation. These mechanisms must be dynamic, allowing for adjustments to ribosome biogenesis to fulfill biological requirements for protein synthesis during development, and following stress induction of global changes in gene expression. In this study, we investigated whether r-protein paralogs are feedback regulated at the transcript level by obtaining a T-DNA knockout of one member, RPL23aB, from the two-member RPL23a family. Expression of the lone functional paralog in this line, RPL23aA, was compared to the expression of both paralogs in wildtype plants under non-stressed, low temperature-, and high light stresses. RPL23aA expression was not upregulated in RPL23aB knockouts to compensate for paralog-loss, and consequently knockouts showed reduced total abundance of RPL23a transcripts. However, no phenotype developed in RPL23aB knockouts, suggesting that this paralog is dispensable under experimental conditions examined, or that compensation by RPL23aA may occur post-transcriptionally. Patterns of RPL23aA and RPL23aB transcript accumulation in wildtype plants suggest that paralogs respond coordinately to developmental and stress stimuli.

Organic Farming in Central Alberta: Current Trends, Production Constraints and Research Needs

ABSTRACT An estimated one-third of organic farmers in Alberta are located within a 150 km radius of Edmonton, a commercial center with a population of close to one million. A random sample of these producers were administered and an in-depth survey in 2002 to obtain knowledge of their crop and commodity selection practices, land usage, fertility management, perceived research needs and recognized constraints to sector viability. The study area contained a 4-fold greater proportion of land cropped to cereals (40%) and double the proportion of farms raising organic cattle (45%), but had less than half the amount of pasture and forage land (35%) than the province as a whole. Albertan organic farmers felt they were constrained by markets, soil fertility, weeds, and production costs. They would like to see more research focused on improving and protecting soil productivity, developing profitable and sustainable crop rotations, identifying alternative livestock vaccines and feed supplements, and comparing organic and conventional farming systems. Overcoming challenges facing this emerging sector will require the combined assistance of government, industry and research sectors.

Plasticity, life cycle and interference potential of field violet (Viola arvensis Murr.) in direct-seeded wheat and canola in central Alberta

Field violet is an established weed of field crops in Europe and eastern North America. It has recently been identified in field crops of central Alberta. Natural infestations of field violet in direct-seeded wheat and canola in central Alberta were observed in 2002 and 2003. Field violet exhibited summer annual, winter annual and short-lived perennial life cycles. Plants emerged in variable flushes, with peaks in early-June and September. Very few annuals produced seed during the season if they emerged after June, but some (< 45%) of these plants successfully overwintered. Summer annuals began dispersing mature seed 7 wk after emergence, but were also capable of persisting in a quiescent state for extended periods (up to 19 wk). Wheat and canola yield loss attributable to field violet was minimal. We compared domestic and European field violet accessions to determine if the study population represented a unique agroecotype. Plants from both cohorts grown in the greenhouse were near-identical, suggesting ...

Effect of Herbicides on Field Violet (Viola arvensis) in Direct-Seeded Spring Wheat

The agrestal field violet, a pervasive weed in Europe, has been identified in reduced-tillage cereal fields in Alberta. The efficacy of herbicides in direct-seeded spring wheat was assessed on natural field violet infestations in Alberta in 2002 and 2003. Only fluroxypyr + 2,4-D, applied postemergence, provided control of field violet in 2002 when rainfall was limiting. Over both years, this herbicide combination reduced biomass by 59 to 69% and plant density by 83 to 91%, relative to nontreated plots. The herbicides metsulfuron, sulfosulfuron, and thifensulfuron + tribenuron only suppressed weed growth under drought conditions in 2002 but controlled the weed in 2003 when rainfall was greater, reducing plant density by 82 to 92% and rendering remaining plants sterile. Suppression was also observed with MCPA + mecoprop + dicamba in 2002 and 2003 and with metribuzin only in 2003. Effective control of field violet was conferred by a pre–crop emergence application of glyphosate at 445 g ae/ha in 2003, the only year that this treatment was evaluated. Activity of herbicides on three- to four-leaf seedlings was also evaluated in a greenhouse dose– response assay. All herbicides had greater efficacy in the greenhouse, and those that provided control in situ reduced field violet dry weight by 85% at less than the recommended rate used in field experiments. Management of field violet is possible with herbicides registered for use on spring wheat in Alberta. However, the weed does not appear to cause significant crop production losses; hence, herbicide selection should be based on knowledge of all weed species present within the field. Nomenclature: 2,4-D; dicamba; fluroxypyr; glyphosate; MCPA; mecoprop; metribuzin; metsulfuron; sulfosulfuron; thifensulfuron; tribenuron; field violet, Viola arvensis Murr. #3 VIOAR; hard red spring wheat, Triticum aestivum L. ‘AC Barrie’. Additional index words: Bentazon, direct seeded, herbicide evaluation, linuron, metribuzin, weed control. Abbreviations: ALS, acetolactate synthase (EC 4.1.3.18); ED50, effective herbicide dose necessary to cause a 50% reduction in weed dry weight; ED85, effective herbicide dose necessary to cause an 85% reduction in weed dry weight; POST, postemergence; PREPLANT, pre–crop emergence; RU, reproductive units; WAT, weeks after treatment.

Evolutionary divergence of ribosomal protein paralogs in Arabidopsis

The majority of proteins in plants are synthesized by the cytoplasmic ribosome. In Arabidopsis, this massive (~3.2 MDa), two-subunit (40S small subunit [SSU] and 60S large subunit [LSU]) enzyme is comprised of 81 ribosomal proteins (RPs; 33 SSU, 48 LSU) that assemble around a catalytic core of 4 ribosomal RNAs (rRNA; 1 SSU, 3 LSU).1,2 None of the Arabidopsis RPs are encoded by single genes, but rather derive from families of 2–7 members that encode nearly-identical isoforms, are independently regulated, dispersed throughout the genome, and largely all transcriptionally active.2 To gain an understanding of why so many plant RP paralogs exist, we recently analyzed function and localization of the two-member Arabidopsis RPL23a family (RPL23aA and RPL23aB).3 RPL23aB was found to be unnecessary for normal development, while a relatively small decrease in transcript levels of RPL23aA resulted in development of a severe phenotype. Isoforms exhibited differences in nucleolar-targeting, which may result from disparity in putative nuclear/nucleolar localization signals (NLS/NoLSs). We postulate a role for ribosome biogenesis in the primary regulation of auxin homeostasis and plant development, and discuss properties of high efficiency NoLSs. Addendum to: Degenhardt RF, Bonham-Smith PC. Arabidopsis ribosomal proteins RPL23aA and -B are differentially targeted to the nucleolus and are disparately required for normal development. Plant Physiol 2008; In press.