Publisher Correction: Science diplomacy for plant health.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Vitamin D receptor (VDR) polymorphisms and severe RSV bronchiolitis: a systematic review and meta-analysis.

BACKGROUND: A number of small studies have suggested a relationship between vitamin D status and severe acute lower respiratory tract infection (ALRI), including RSV-bronchiolitis. The objective of this study was to evaluate the relationship between vitamin D receptor (VDR) polymorphism and severe RSV-bronchiolitis through a systemic literature review and meta-analysis. METHODS: A comprehensive electronic literature search was conducted to identify all studies published before January 2013. Two reviewers independently screened all abstracts, followed by the full text of potential articles to evaluate eligibility. Study methodological quality was evaluated using the Newcastle Ottawa scale and individual component analysis. Meta-analysis evaluated associations at the allele and genotype levels. RESULTS: Of 803 studies identified from our literature search, three met eligibility criteria. Two VDR polymorphisms were included in more than one study: TaqI (rs731236) and FokI (rs2228570). All three reported a positive relationship between the FokI minor allele and disease with random effects meta-analyses demonstrating a statistically significant relationship (OR 1.52, CI: 1.12, 2.05). Genotype analysis was highly suggestive of a dominant or incomplete dominance model with combined odds ratios for fF (OR 1.73, CI: 0.92-3.36) and ff (OR 2.24, CI: 0.98-5.14) compared to the FF genotype. No association between TaqI and severe RSV-bronchiolitis was evident at the allele or genotype level. CONCLUSIONS: Available literature supports an association between the FokI polymorphism and severe RSV disease. Determination of VDR receptor polymorphism status could help predict high-risk infants who might benefit from preventive measures.

Ethics application protocols for multicentre clinical studies in Canada: A paediatric rheumatology experience.

INTRODUCTION: Individual institutions govern research ethics applications and each must administer and regulate their own protocols. Variations in ethics review procedures and expectations among centres impose impediments to efficiently conducting multicentre studies. METHODS: Observations relating to preparing multisite ethics documents for a study conducted by Canadian paediatric rheumatology investigators are described. Research ethics applications from the 12 participating centres were compared. RESULTS: Although the applications were similar in their content, they differed in their formatting. All applications shared a commitment to ensuring that the study conformed to exemplary ethical standards. CONCLUSIONS: There is wide variation in the multicentre clinical study ethics application process at the institutional level. Considering the common fundamental elements required by all ethics review boards, the present study conceptualized introducing a discipline-specific uniform ethics application process acceptable to all Canadian research ethics boards. This may be a more efficient strategy that could help lessen the burden of collaborative research.

INTRODUCTION: Chaque établissement régit des applications éthiques en recherche, et doit administrer et réglementer ses propres protocoles. Des variations dans les méthodes d'analyse éthique et les attentes entre les centres imposent des entraves à la tenue d'études multicentriques. MÉTHODOLOGIE: Les auteurs décrivent les observations relatives à la préparation de documents éthiques multisites dans le cadre d'une étude menée par des chercheurs canadiens en rhumatologie pédiatrique. Ils ont comparé les applications éthiques de recherche utilisées dans les 12 centres participants. RÉSULTATS: Même si le contenu des applications était similaire, leur mise en forme était différente. Dans tous les documents, on s'engageait à s'assurer que l'étude respectait des normes éthiques exemplaires. CONCLUSIONS: On constate une importante variation dans le processus d'applications éthiques des études cliniques multicentriques des établissements. Étant donné les éléments fondamentaux communs exigés de tous les comités d'analyse éthique, la présente étude a conceptualisé l'adoption d'un processus d'applications éthiques uniforme propre à chaque discipline et acceptable au sein de tous les comités canadiens d'éthique de la recherche. Ce pourrait être une stratégie plus efficace qui pourrait rendre les recherches coopératives moins fastidieuses.

Use of monocyte/endothelial cell co-cultures (in vitro) and a subcutaneous implant mouse model (in vivo) to evaluate a degradable polar hydrophobic ionic polyurethane.

Potential benefits of co-culturing monocytes (MC) with vascular smooth muscle cells have been reported on for tissue engineering applications with a degradable, polar, hydrophobic, and ionic polyurethane (D-PHI). Since the interaction of MC and endothelial cells (EC) within the blood vessel endothelium is also a process of wound repair it was of interest to investigate their function when cultured on the synthetic D-PHI materials, prior to considering the materials' use in vascular engineering. The co-culture (MC/EC) in vitro studies were carried out on films in 96 well plates and porous scaffold disks were prepared for implant studies in an in vivo subcutaneous mouse model. After 7 days in culture, the MC/EC condition was equal to EC growth but had lower esterase activity (a measure of degradative potential), no pro-inflammatory TNF-α and a relatively high anti-inflammatory IL-10 release while the ECs maintained their functional marker CD31. After explantation of the porous scaffolds, a live/dead stain showed that the cells infiltrating the scaffolds were viable and histological stains (May-Grunwald, Trichrome) demonstrated tissue in growth and extracellular matrix synthesis. Lysates from the implant scaffolds analyzed with a cytokine antibody array showed decreased pro-inflammatory cytokines (IL-6, TNF-α, GM-CSF), increased anti-inflammatory cytokines (IL-10, IL-13, TNF-RI), and increased chemotactic cytokines (MCP-1, MCP-5, RANTES). The low foreign body response elicited by D-PHI when implanted in vivo supported the in vitro studies (EC and MC co-culture), demonstrating that D-PHI promoted EC growth along with an anti-inflammatory MC, further demonstrating its potential as a tissue engineering scaffold for vascular applications.

Vitamin D deficiency in young children with severe acute lower respiratory infection.

RATIONALE: Acute lower respiratory infection (ALRI) is one of the most common reasons for hospitalization and intensive care unit admission among children. Season related decreases in the immunomodulatory molecule, vitamin D, remain an unexplored factor that might contribute to the increased occurrence of ALRI in children. OBJECTIVE: To investigate a possible association between vitamin D deficiency and respiratory infection by comparing serum 25 hydroxyvitamin D [25(OH)D] levels in a group of young children with ALRI to an age-matched group without respiratory infection. PATIENTS AND METHODS: Participants with a diagnosis of bronchiolitis or pneumonia (n = 55 or 50, respectively), as well as control subjects without respiratory symptoms (n = 92), were recruited at the Royal University Hospital, Saskatoon, Saskatchewan, Canada from November 2007 to May 2008. 25(OH)D levels were measured in patient serum using a competitive enzyme linked immunoassay. RESULTS: The mean vitamin D level for the entire ALRI group was not significantly different from the control group (81 +/- 40 vs. 83 +/- 30 nmol/L, respectively). The mean vitamin D level for the ALRI subjects admitted to the pediatric intensive care unit (49 +/- 24 nmol/L) was significantly lower than that observed for both control (83 +/- 30 nmol/L) and ALRI subjects admitted to the general pediatrics ward (87 +/- 39 nmol/L). Vitamin D deficiency remained statistically related to pediatric intensive care unit admission in the multivariate analysis. CONCLUSION: No difference was observed in vitamin D levels between the entire ALRI group and control groups; however, significantly more children admitted to the pediatric intensive care unit with ALRI were vitamin D deficient. These findings suggest that the immunomodulatory properties of vitamin D might influence ALRI disease severity.

Effect of polyurethane chemistry and protein coating on monocyte differentiation towards a wound healing phenotype macrophage.

Tissue regeneration alternatives for peripheral vascular disease are actively being investigated; however, few studies in this area have probed the role of the wound healing monocyte-derived macrophage (MDM). Inflammatory MDMs transition to wound healing MDMs as the relative levels of tumor necrosis factor-alpha (TNF-alpha) decrease and IL-10 increase. TNF-alpha has been linked to the regulation of HMGB1 (high mobility group box 1 protein), a nuclear protein that upon macrophage stimulation can be secreted and act as a pro-inflammatory cytokine. This study investigated the influence of a degradable polar hydrophobic ionic polyurethane (D-PHI) on MDM cell expression of pro- versus anti-inflammatory markers, when the material was uncoated or pre-coated with collagen prior to cell studies. Effects were compared to similar groups on tissue culture polystyrene (TCPS). Collagen coated TCPS and D-PHI had significantly more DNA than the uncoated TCPS after 7d (p=0.001 and p=0.006 respectively); however, there was significantly less esterase activity from cells on D-PHI (+/-collagen) than for cells on TCPS after 7d (p=0.002, p=0.0003 respectively). No significant differences in esterase activity were observed between collagen coated and non-coated D-PHI surfaces. Analyses of pro-inflammatory cytokines (TNF-alpha, IL-1beta and HMGB1) secreted from differentiating monocytes adherent to D-PHI demonstrated a decrease whereas anti-inflammatory IL-10 increased over time when compared to TCPS, suggesting that D-PHI was less inflammatory than TCPS. Since D-PHI maintains cell attachment while aiding in the transition of MDM to a wound healing phenotype, this material has qualities suitable to be used in tissue engineering applications where MDM play a key role in tissue regeneration.

The effects of phorbol ester activation and reactive oxygen species scavengers on the macrophage-mediated foreign body reaction to polyurethanes.

Phorbol myristate acetate, a protein kinase C activator, inhibited monocyte-derived macrophage (MDM)-mediated degradation of aliphatic (HDI) polycarbonate-based polyurethanes but not degradation of the aromatic polycarbonate-based polyurethane (MDI). The objectives of this study were to determine if reactive oxygen species are involved in the phorbol myristate acetate effect on esterase activity and MDM-mediated polycarbonate-based polyurethane degradation and to find a good marker of material-initiated activation of MDM. The phorbol myristate acetate-dependent effects of the material chemistry on cell activation and degradation were evaluated by adding reactive oxygen species scavengers, catalase plus superoxide dismutase to MDM and assaying possible markers of MDM activation: esterase activity, acid phosphatase activity, and high molecular weight group box 1 protein (HMGB1). All treatments reduced the esterase activity in MDM on HDI but not in MDM on MDI. Acid phosphatase was inhibited in MDM to varying degrees on all surfaces by phorbol myristate acetate or catalase plus superoxide dismutase either alone or together. Secretion of HMGB1 from MDM on HDI431 was higher than MDI; however only secretion from MDM on HDI was inhibited by phorbol myristate acetate. In MDM on HDI, catalase plus superoxide dismutase reduced intracellular HMGB1 levels +/- phorbol myristate acetate; whereas, catalase, superoxide dismutase plus phorbol myristate acetate increased intracellular HMGB1 in MDM on MDI, suggesting that esterase and HMGB1 are more specific markers of activation than acid phosphatase. Manipulation of signaling pathways may provide insight surrounding the mechanism of activation for oxidative and/or hydrolytic degradative pathways in the MDM response to material surface chemistry.

Dynamic organization of COPII coat proteins at endoplasmic reticulum export sites in plant cells.

Protein export from the endoplasmic reticulum (ER) is mediated by the accumulation of COPII proteins such as Sar1, Sec23/24 and Sec13/31 at specialized ER export sites (ERES). Although the distribution of COPII components in mammalian and yeast systems is established, a unified model of ERES dynamics has yet to be presented in plants. To investigate this, we have followed the dynamics of fluorescent fusions to inner and outer components of the coat, AtSec24 and AtSec13, in three different plant model systems: tobacco and Arabidopsis leaf epidermis, as well as tobacco BY-2 suspension cells. In leaves, AtSec24 accumulated at Golgi-associated ERES, whereas AtSec13 showed higher levels of cytosolic staining compared with AtSec24. However, in BY-2 cells, both AtSec13 and AtSec24 labelled Golgi-associated ERES, along with AtSec24. To correlate the distribution of the COPII coat with the dynamics of organelle movement, quantitative live-cell imaging analyses demonstrated that AtSec24 and AtSec13 maintained a constant association with Golgi-associated ERES, irrespective of their velocity. However, recruitment of AtSec24 and AtSec13 to ERES, as well as the number of ERES marked by these proteins, was influenced by export of membrane cargo proteins from the ER to the Golgi. Additionally, the increased availability of AtSec24 affected the distribution of AtSec13, inducing recruitment of this outer COPII coat component to ERES. These results provide a model that, in plants, protein export from the ER occurs via sequential recruitment of inner and outer COPII components to form transport intermediates at mobile, Golgi-associated ERES.

Efficient mitochondrial targeting relies on co-operation of multiple protein signals in plants.

To date, the most prevalent model for transport of pre-proteins to plant mitochondria is based on the activity of an N-terminal extension serving as a targeting peptide. Whether the efficient delivery of proteins to mitochondria is based exclusively on the action of the N-terminal extension or also on that of other protein determinants has yet to be defined. A novel mechanism is reported here for the targeting of a plant protein, named MITS1, to mitochondria. It was found that MITS1 contains an N-terminal extension that is responsible for mitochondrial targeting. Functional dissection of this extension shows the existence of a cryptic signal for protein targeting to the secretory pathway. The first 11 amino acids of the N-terminal extension are necessary to overcome the activity of this signal sequence and target the protein to the mitochondria. These data suggest that co-operation of multiple determinants within the N-terminal extension of mitochondrial proteins may be necessary for efficient mitochondrial targeting. It was also established that the presence of a tryptophan residue toward the C-terminus of the protein is crucial for mitochondrial targeting, as mutation of this residue results in a redistribution of MITS1 to the endoplasmic reticulum and Golgi apparatus. These data suggest a novel targeting model whereby protein traffic to plant mitochondria is influenced by domains in the full-length protein as well as the N-terminal extension.

Epidemiologic considerations in unexplained pediatric arthralgia: the role of season, school, and stress.

OBJECTIVE: To determine demographic and epidemiologic characteristics in children with unexplained joint pain. METHODS: The study population included 730 children (< 18 yrs of age) referred between 1981 and 2007 to the Saskatchewan Pediatric Rheumatology Program, University of Saskatchewan, because of arthralgia. Parents and patients completed a questionnaire at the time of initial presentation, and a diagnosis of unexplained arthralgia was assigned based on clinical assessment. Serum vitamin D levels were measured in 73 patients diagnosed with arthralgia. RESULTS: Subjects with arthralgia were more likely to report psychosocial stresses including family discord and illness in the family, and to be cared for by a single parent as a consequence of parental separation or death. Significantly more patients reported fall and winter (30%) as the season of symptom onset compared to spring or summer (20%; p = 0.01). Significantly more survey respondents in the arthralgia group reported missing school compared to the control group (62% vs 31%; p = 0.001). Referrals from northern Saskatchewan were significantly more numerous than from southern Saskatchewan (107 vs 45 per 100,000; p < 0.001). Serum vitamin D concentrations measured in a subgroup of patients (n = 73) showed that 62 (82%) were abnormally low, 42% between 50 and 75 nmol/l (insufficient), and 40% < 50 nmol/l (deficient). CONCLUSION: Our results suggest an association between psychosocial stress, school absenteeism, vitamin D insufficiency, and unexplained arthralgia in children.

Capillary blood sampling as an alternative to venipuncture in the assessment of serum 25 hydroxyvitamin D levels.

This study compared 25-hydroxyvitamin D [25(OH)D] measurements in capillary and venous blood samples collected, respectively by fingerprick and venipuncture. Capillary blood for measuring 25(OH)D has potential advantages by reducing blood volume required (2mL versus 0.3mL for venipuncture and capillary sampling, respectively), facilitating blood collection for those populations in whom venipuncture is difficult (e.g. infants and children), improving patient convenience and reducing costs associated with phlebotomy. The results demonstrated a highly significant relationship between 25(OH)D levels in serum derived from venous and capillary blood samples (r(2)=0.901). Despite statistically higher 25(OH)D levels in fingerprick samples (108+/-9nmol/L) compared with venipuncture samples (90+/-7nmol/L), the correlation between venous and capillary samples provides support for this approach as a practical alternative to venipuncture for vitamin D determination. However, clinical application may require the incorporation of a correction factor for the assessment of insufficiency, and research studies should avoid using the two methods interchangeably. Studying vitamin D's role in health and disease requires collection techniques and measurement methods that are reliable, reproducible, easily accessible, inexpensive and minimally burdensome to the patient. The option to collect patient samples by fingerprick may facilitate the collection process.

Assessment of sample collection and storage methods for multicenter immunologic research in children.

Multicenter studies involving both large and small centers separated by significant distances pose unique challenges to biological sample collection. The objective of this study was to evaluate protocols for determining inflammatory biomarkers that are cost and manpower efficient for handling blood destined for a sample repository. Tempus (Applied Biosystems) and Paxgene (Qiagen) blood collection systems were evaluated for RNA isolation. P100 tubes (BD), containing propriety stabilizers for preservation of plasma proteins, were evaluated for protein content and compared with plasma collected in conventional tubes. Blood for plasma separation was spiked with recombinant TNF-alpha and IL-2 prior to being processed and stored under various conditions. The Tempus RNA system produced a significantly greater yield of RNA at comparable quality when stored at 4 degrees C and shipped at ambient temperature than any other condition tested. The Tempus system was 20% less expensive and required approximately 40% less processing time thereby reducing costs. The P100 system preserved recombinant TNF-alpha in blood shipped at ambient temperature significantly better than conventionally collected plasma that was shipped on dry ice. There was no significant difference in IL-2 levels between the two collection methods and shipping temperatures. The Tempus RNA blood collection tubes and the P100 protein stabilization system provide the opportunity for reliable collection and ambient temperature transport of samples in multicenter studies. This cost-effective, standardized protocol for a large multicenter trial ensures the integrity of biological samples and maximizes study participation by both large and small centers.

Plant Sar1 isoforms with near-identical protein sequences exhibit different localisations and effects on secretion.

In plants, differentiation of subdomains of the endoplasmic reticulum (ER) dedicated to protein export, the ER export sites (ERES), is influenced by the type of export-competent membrane cargo to be delivered to the Golgi. This raises a fundamental biological question: is the formation of transport intermediates at the ER for trafficking to the Golgi always regulated in the same manner? To test this, we followed the distribution and activity of two plant Sar1 isoforms. Sar1 is the small GTPase that regulates assembly of COPII (coat protein complex II) on carriers that transport secretory cargo from ER to Golgi. We show that, in contrast to a tobacco Sar1 isoform, the two Arabidopsis Sar1 GTPases were localised at ERES, independently of co-expression of Golgi-destined membrane cargo in tobacco cells. Although both isoforms labelled ERES, one was found to partition with the membrane fraction to a greater extent. The different distribution of fluorescent fusions of the two isoforms was influenced by the nature of an amino acid residue at the C-terminus of the protein, suggesting that the requirements for membrane association of the two GTPases are not equal. Furthermore, functional analyses based on the secretion of the bulk flow marker alpha-amylase indicated that over-expression of GTP-restricted mutants of the two isoforms caused different levels of ER export inhibition. These novel results indicate a functional heterogeneity among plant Sar1 isoforms.

Correct targeting of plant ARF GTPases relies on distinct protein domains.

Indispensable membrane trafficking events depend on the activity of conserved small guanosine triphosphatases (GTPases), anchored to individual organelle membranes. In plant cells, it is currently unknown how these proteins reach their correct target membranes and interact with their effectors. To address these important biological questions, we studied two members of the ADP ribosylation factor (ARF) GTPase family, ARF1 and ARFB, which are membrane anchored through the same N-terminal myristoyl group but to different target membranes. Specifically, we investigated how ARF1 is targeted to the Golgi and post-Golgi structures, whereas ARFB accumulates at the plasma membrane. While the subcellular localization of ARFB appears to depend on multiple domains including the C-terminal half of the GTPase, the correct targeting of ARF1 is dependent on two domains: an N-terminal ARF1 domain that is necessary for the targeting of the GTPase to membranes and a core domain carrying a conserved MxxE motif that influences the relative distribution of ARF1 between the Golgi and post-Golgi compartments. We also established that the N-terminal ARF1 domain alone was insufficient to maintain an interaction with membranes and that correct targeting is a protein-specific property that depends on the status of the GTP switch. Finally, an ARF1-ARFB chimera containing only the first 18 amino acids from ARF1 was shown to compete with ARF1 membrane binding loci. Although this chimera exhibited GTPase activity in vitro, it was unable to recruit coatomer, a known ARF1 effector, onto Golgi membranes. Our results suggest that the targeting of ARF GTPases to the correct membranes may not only depend on interactions with effectors but also relies on distinct protein domains and further binding partners on the Golgi surface.

Studying protein export from the endoplasmic reticulum in plants.

Understanding the mechanisms of protein sorting and targeting through the plant secretory pathway has become the focus of many research laboratories. The development of a model system whereby recombinant genes can be transiently expressed in protoplasts has facilitated the study of protein transport signals. Experimental strategies combining a protoplast expression system with endoglycosidase H, vacuole purification, and pulse-chase analyses are used to investigate aspects of specific proteins as they pass through the secretory system. This chapter provides details of protoplast preparation and electroporation as well as techniques to study protein trafficking from the endoplasmic reticulum to the Golgi apparatus or vacuolar compartments. Recommendations as to how to troubleshoot problems that can arise while following these protocols are also discussed in this chapter.

Is cell culture stressful? Effects of degradable and nondegradable culture surfaces on U937 cell function.

In vitro cell culture has become one of the most widely used techniques in biological and health sciences research, with the most common culture supports being either tissue culture grade polystyrene (TCPS) or polydimethylsiloxane (PDMS). It has previously been shown that monocyte-derived macrophages (MDMs) respond to material surface chemistry, synthesizing and releasing degradative activities that could produce products, which alter the cell's response. In this study, functional parameters of differentiated U937 macrophage-like cells were compared when cultured on nondegradable standard control surfaces versus models of biomaterials (polycarbonate-based polyurethanes) used in the manufacture of medical devices previously shown to degrade and/or elicit pathways of inflammation. Although the influence of PDMS and TCPS on cell function is often underappreciated by investigators, both surfaces elicited enzyme markers of inflammation. Cells on TCPS had the highest intracellular and released esterase activities and protein levels. Cells on PDMS had the most released acid phosphatase activity and protein (P < 0.001), as well as de novo 57- and 59-kDa released proteins. The criteria for defining an activated cell phenotype become critically important when materials such as PDMS and TCPS are used as standard control surfaces whether in experiments for research in elucidating metabolic pathways or in screening drugs and materials for therapeutic uses.

Post-Golgi protein traffic in the plant secretory pathway.

The Golgi apparatus in plants is organized as a multitude of individual stacks that are motile in the cytoplasm and in close association with the endoplasmic reticulum (ER) (Boevink et al. in Plant J 15:441-447, 1998). These stacks operate as a sorting centre for cargo molecules, providing modification and redirection to other organelles as appropriate. In the post-Golgi direction, these include vacuole and plasma membrane, and specialized transport routes to each are required to prevent mislocalization. Recent evidence in plant cells points to the existence of post-Golgi organelles that function as intermediate stations for efficient protein traffic, as well as to the influence of small GTPases such as Rabs and ARFs on post-Golgi trafficking. This review focuses on the latest findings on post-Golgi trafficking routes and on the involvement of GTPases and their effectors on the trafficking of proteins in the plant secretory pathway.

Multiple roles of ADP-ribosylation factor 1 in plant cells include spatially regulated recruitment of coatomer and elements of the Golgi matrix.

Recent evidence indicates that ADP-ribosylation factor 1 (ARF1) carries out multiple roles in plant cells that may be independent from the established effector complex COPI. To investigate potential COPI-independent functions, we have followed the dynamics of ARF1 and a novel putative effector, the plant golgin GRIP-related ARF-binding domain-containing Arabidopsis (Arabidopsis thaliana) protein 1 (GDAP1) in living plant cells. We present data that ascribe a new role to ARF1 in plant cell membrane traffic by showing that the GTPase functions to recruit GDAP1 to membranes. In addition, although ARF1 appears to be central to the recruitment of both COPI components and the golgin, we have established a different subcellular distribution of these ARF1 effectors. Live cell imaging demonstrates that GDAP1 and COPI are distributed on Golgi membranes. However, GDAP1 is also found on ARF1-labeled structures that lack coatomer, suggesting that the membrane environment, rather than ARF1 alone, influences the differential recruitment of ARF1 effectors. In support of this hypothesis, fluorescence recovery after photobleaching analyses demonstrated that GDAP1 and COPI have different kinetics on membranes during the cycle of activation and inactivation of ARF1. Therefore, our data support a model where modulation of the cellular functions of ARF1 in plant cells encompasses not only the intrinsic activities of the effectors, but also differential recruitment onto membranes that is spatially regulated.

De novo formation of plant endoplasmic reticulum export sites is membrane cargo induced and signal mediated.

The plant endoplasmic reticulum (ER) contains functionally distinct subdomains at which cargo molecules are packed into transport carriers. To study these ER export sites (ERES), we used tobacco (Nicotiana tabacum) leaf epidermis as a model system and tested whether increased cargo dosage leads to their de novo formation. We have followed the subcellular distribution of the known ERES marker based on a yellow fluorescent protein (YFP) fusion of the Sec24 COPII coat component (YFP-Sec24), which, differently from the previously described ERES marker, tobacco Sar1-YFP, is visibly recruited at ERES in both the presence and absence of overexpressed membrane cargo. This allowed us to quantify variation in the ERES number and in the recruitment of Sec24 to ERES upon expression of cargo. We show that increased synthesis of membrane cargo leads to an increase in the number of ERES and induces the recruitment of Sec24 to these ER subdomains. Soluble proteins that are passively secreted were found to leave the ER with no apparent up-regulation of either the ERES number or the COPII marker, showing that bulk flow transport has spare capacity in vivo. However, de novo ERES formation, as well as increased recruitment of Sec24 to ERES, was found to be dependent on the presence of the diacidic ER export motif in the cytosolic domain of the membrane cargo. Our data suggest that the plant ER can adapt to a sudden increase in membrane cargo-stimulated secretory activity by signal-mediated recruitment of COPII machinery onto existing ERES, accompanied by de novo generation of new ERES.