Antagonistic RALF peptides control an intergeneric hybridization barrier on Brassicaceae stigmas.

Pollen-pistil interactions establish interspecific/intergeneric pre-zygotic hybridization barriers in plants. The rejection of undesired pollen at the stigma is crucial to avoid outcrossing but can be overcome with the support of mentor pollen. The mechanisms underlying this hybridization barrier are largely unknown. Here, in Arabidopsis, we demonstrate that receptor-like kinases FERONIA/CURVY1/ANJEA/HERCULES RECEPTOR KINASE 1 and cell wall proteins LRX3/4/5 interact on papilla cell surfaces with autocrine stigmatic RALF1/22/23/33 peptide ligands (sRALFs) to establish a lock that blocks the penetration of undesired pollen tubes. Compatible pollen-derived RALF10/11/12/13/25/26/30 peptides (pRALFs) act as a key, outcompeting sRALFs and enabling pollen tube penetration. By treating Arabidopsis stigmas with synthetic pRALFs, we unlock the barrier, facilitating pollen tube penetration from distantly related Brassicaceae species and resulting in interspecific/intergeneric hybrid embryo formation. Therefore, we uncover a "lock-and-key" system governing the hybridization breadth of interspecific/intergeneric crosses in Brassicaceae. Manipulating this system holds promise for facilitating broad hybridization in crops.

Central-cell-produced attractants control fertilization recovery.

Animal fertilization relies on hundreds of sperm racing toward the egg, whereas, in angiosperms, only two sperm cells are delivered by a pollen tube to the female gametes (egg cell and central cell) for double fertilization. However, unsuccessful fertilization under this one-pollen-tube design can be detrimental to seed production and plant survival. To mitigate this risk, unfertilized-gamete-controlled extra pollen tube entry has been evolved to bring more sperm cells and salvage fertilization. Despite its importance, the underlying molecular mechanism of this phenomenon remains unclear. In this study, we report that, in Arabidopsis, the central cell secretes peptides SALVAGER1 and SALVAGER2 in a directional manner to attract pollen tubes when the synergid-dependent attraction fails or is terminated by pollen tubes carrying infertile sperm cells. Moreover, loss of SALs impairs the fertilization recovery capacity of the ovules. Therefore, this research uncovers a female gamete-attraction system that salvages seed production for reproductive assurance.

Pollen Cell Wall Patterns Form from Modulated Phases.

The ornately geometric walls of pollen grains have inspired scientists for decades. We show that the evolved diversity of these patterns is entirely recapitulated by a biophysical model in which an initially uniform polysaccharide layer in the extracellular space, mechanically coupled to the cell membrane, phase separates to a spatially modulated state. Experiments reveal this process occurring in living cells. We observe that in ∼10% of extant species, this phase separation reaches equilibrium during development such that individual pollen grains are identical and perfectly reproducible. About 90% of species undergo an arrest of this process prior to equilibrium such that individual grains are similar but inexact copies. Equilibrium patterns have appeared multiple times during the evolution of seed plants, but selection does not favor these states. This framework for pattern development provides a route to rationalizing the surface textures of other secreted structures, such as cell walls and insect cuticle.

Regulatory T Cell Specificity Directs Tolerance versus Allergy against Aeroantigens in Humans.

FOXP3+ regulatory T cells (Tregs) maintain tolerance against self-antigens and innocuous environmental antigens. However, it is still unknown whether Treg-mediated tolerance is antigen specific and how Treg specificity contributes to the selective loss of tolerance, as observed in human immunopathologies such as allergies. Here, we used antigen-reactive T cell enrichment to identify antigen-specific human Tregs. We demonstrate dominant Treg-mediated tolerance against particulate aeroallergens, such as pollen, house dust mites, and fungal spores. Surprisingly, we found no evidence of functional impairment of Treg responses in allergic donors. Rather, major allergenic proteins, known to rapidly dissociate from inhaled allergenic particles, have a generally reduced capability to generate Treg responses. Most strikingly, in individual allergic donors, Th2 cells and Tregs always target disparate proteins. Thus, our data highlight the importance of Treg antigen-specificity for tolerance in humans and identify antigen-specific escape from Treg control as an important mechanism enabling antigen-specific loss of tolerance in human allergy.

Flavonols improve tomato pollen thermotolerance during germination and tube elongation by maintaining ROS homeostasis.

Elevated temperatures impair pollen performance and reproductive success, resulting in lower crop yields. The tomato (Solanum lycopersicum) anthocyanin reduced (are) mutant harbors a mutation in FLAVANONE 3-HYDROXYLASE (F3H), resulting in impaired flavonol antioxidant biosynthesis. The are mutant has reduced pollen performance and seed set relative to the VF36 parental line, phenotypes that are accentuated at elevated temperatures. Transformation of are with the wild-type F3H gene, or chemical complementation with flavonols, prevented temperature-dependent reactive oxygen species (ROS) accumulation in pollen and restored the reduced viability, germination, and tube elongation of are to VF36 levels. Overexpression of F3H in VF36 prevented temperature-driven ROS increases and impaired pollen performance, revealing that flavonol biosynthesis promotes thermotolerance. Although stigmas of are had reduced flavonol and elevated ROS levels, the growth of are pollen tubes was similarly impaired in both are and VF36 pistils. RNA-seq was performed at optimal and stress temperatures in are, VF36, and the F3H overexpression line at multiple timepoints across pollen tube elongation. The number of differentially expressed genes increased over time under elevated temperatures in all genotypes, with the greatest number in are. These findings suggest potential agricultural interventions to combat the negative effects of heat-induced ROS in pollen that lead to reproductive failure.

Mechanisms of climate change and related air pollution on the immune system leading to allergic disease and asthma.

Climate change is considered the greatest threat to global health. Greenhouse gases as well as global surface temperatures have increased causing more frequent and intense heat and cold waves, wildfires, floods, drought, altered rainfall patterns, hurricanes, thunderstorms, air pollution, and windstorms. These extreme weather events have direct and indirect effects on the immune system, leading to allergic disease due to exposure to pollen, molds, and other environmental pollutants. In this review, we will focus on immune mechanisms associated with allergy and asthma-related health risks induced by climate change events. We will review current understanding of the molecular and cellular mechanisms by which the changing environment mediates these effects.

Deep imaging reveals dynamics and signaling in one-to-one pollen tube guidance.

In the pistil of flowering plants, each ovule usually associates with a single pollen tube for fertilization. This one-to-one pollen tube guidance, which contributes to polyspermy blocking and efficient seed production, is largely different from animal chemotaxis of many sperms to one egg. However, the functional mechanisms underlying the directional cues and polytubey blocks in the depths of the pistil remain unknown. Here, we develop a two-photon live imaging method to directly observe pollen tube guidance in the pistil of Arabidopsis thaliana, clarifying signaling and cellular behaviors in the one-to-one guidance. Ovules are suggested to emit multiple signals for pollen tubes, including an integument-dependent directional signal that reaches the inner surface of the septum and adhesion signals for emerged pollen tubes on the septum. Not only FERONIA in the septum but ovular gametophytic FERONIA and LORELEI, as well as FERONIA- and LORELEI-independent repulsion signal, are involved in polytubey blocks on the ovular funiculus. However, these funicular blocks are not strictly maintained in the first 45 min, explaining previous reports of polyspermy in flowering plants.

SPA, a Stigma-style-transmitting tract Physical microenvironment Assay for investigating mechano-signaling in pollen tubes.

Accurate sensing and responding to physical microenvironment are crucial for cell function and survival, but the underlying molecular mechanisms remain elusive. Pollen tube (PT) provides a perfect single-cell model for studying mechanobiology since it's naturally subjected to complex mechanical instructions from the pistil during invasive growth. Recent reports have revealed discrepant PT behaviors between in vivo and flat, two-dimensional in vitro cultures. Here, we established the Stigma-style-transmitting tract (TT) Physical microenvironment Assay (SPA) to recapitulate pressure changes in the pistil. This biomimetic assay has enabled us to swiftly identify highly redundant genes, GEF8/9/11/12/13, as new regulators for maintaining PTs integrity during style-to-TT emergence. In contrast to normal growth on solid medium, SPA successfully phenocopied gef8/9/11/12/13 PT in vivo growth-arrest deficiency. Our results suggest the existence of distinct signaling pathways regulating in vivo and in vitro PT integrity maintenance, underscoring the necessity of faithfully mimicking the physical microenvironment for studying plant cell biology.

Arabidopsis VILLIN5 bundles actin filaments using a novel mechanism.

Being a bona fide actin bundler, Arabidopsis villin5 (VLN5) plays a crucial role in regulating actin stability and organization within pollen tubes. Despite its significance, the precise mechanism through which VLN5 bundles actin filaments has remained elusive. Through meticulous deletion analysis, we have unveiled that the link between gelsolin repeat 6 (G6) and the headpiece domain (VHP), rather than VHP itself, is indispensable for VLN5-mediated actin bundling. Further refinement of this region has pinpointed a critical sequence spanning from Val763 to Ser823, essential for VLN5's actin-bundling activity. Notably, the absence of Val763-Ser823 in VLN5 results in decreased filamentous decoration within pollen tubes and a diminished ability to rescue actin bundling defects in vln2vln5 mutant pollen tubes compared to intact VLN5. Moreover, our findings highlight that the Val763-Ser823 sequence harbors a binding site for F-actin, suggesting that this linker-based F-actin binding site, in conjunction with the F-actin binding site localized in G1-G6, enables a single VLN5 to concurrently bind to two adjacent actin filaments. Therefore, our study unveils a novel mechanism by which VLN5 bundles actin filaments.

MORN motif-containing protein OsMORN1 and OsMORN2 are crucial for rice pollen viability and cold tolerance.

The pollen viability directly affects the pollination process and the ultimate grain yield of rice. Here, we identified that the MORN motif-containing proteins, OsMORN1 and OsMORN2, had a crucial role in maintaining pollen fertility. Compared with the wild type (WT), the pollen viability of the osmorn1 and osmorn2 mutants was reduced, and pollen germination was abnormal, resulting in significantly lower spikelet fertility, seed-setting rate, and grain yield per plant. Further investigation revealed that OsMORN1 was localized to the Golgi apparatus and lipid droplets. Lipids associated with pollen viability underwent alterations in osmorn mutants, such as the diacylglyceride (18:3_18:3) was 5.1-fold higher and digalactosyldiacylglycerol (18:2_18:2) was 5.2-fold lower in osmorn1, while the triacylglycerol (TG) (16:0_18:2_18:3) was 8.3-fold higher and TG (16:0_18:1_18:3) was 8.5-fold lower in osmorn2 than those in WT. Furthermore, the OsMORN1/2 was found to be associated with rice cold tolerance, as osmorn1 and osmorn2 mutants were more sensitive to chilling stress than WT. The mutants displayed increased hydrogen peroxide accumulation, reduced antioxidant enzyme activities, elevated malondialdehyde content, and a significantly decreased seedling survival rate. Lipidomics analysis revealed distinct alterations in lipids under low temperature, highlighting significant changes in TG (18:2_18:3_18:3) and TG (18:4_18:2_18:2) in osmorn1, TG (16:0_18:2_18:2) and PI (17:2_18:3) in osmorn2 compared to the WT. Therefore, it suggested that OsMORN1 and OsMORN2 regulate both pollen viability and cold tolerance through maintaining lipid homeostasis.

Redundant function of the Arabidopsis phosphatidylinositol 4-phosphate 5-kinase genes PIP5K4-6 is essential for pollen germination.

Phosphatidylinositol 4-phosphate 5-kinase (PIP5K) is a key enzyme producing the signaling lipid phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2 ] in eukaryotes. Although PIP5K genes are reported to be involved in pollen tube germination and growth, the essential roles of PIP5K in these processes remain unclear. Here, we performed a comprehensive genetic analysis of the Arabidopsis thaliana PIP5K4, PIP5K5, and PIP5K6 genes and revealed that their redundant function is essential for pollen germination. Pollen with the pip5k4pip5k5pip5k6 triple mutation was sterile, while pollen germination efficiency and pollen tube growth were reduced in the pip5k6 single mutant and further reduced in the pip5k4pip5k6 and pip5k5pip5k6 double mutants. YFP-fusion proteins, PIP5K4-YFP, PIP5K5-YFP, and PIP5K6-YFP, which could rescue the sterility of the triple mutant pollen, preferentially localized to the tricolpate aperture area and the future germination site on the plasma membrane prior to germination. Triple mutant pollen grains under the germination condition, in which spatiotemporal localization of the PtdIns(4,5)P2 fluorescent marker protein 2xmCHERRY-2xPHPLC as seen in the wild type was abolished, exhibited swelling and rupture of the pollen wall, but neither the conspicuous protruding site nor site-specific deposition of cell wall materials for germination. These data indicate that PIP5K4-6 and their product PtdIns(4,5)P2 are essential for pollen germination, possibly through the establishment of the germination polarity in a pollen grain.

In planta imaging of pyridine nucleotides using second-generation fluorescent protein biosensors.

Redox changes of pyridine nucleotides in cellular compartments are highly dynamic and their equilibria are under the influence of various reducing and oxidizing reactions. To obtain spatiotemporal data on pyridine nucleotides in living plant cells, typical biochemical approaches require cell destruction. To date, genetically encoded fluorescent biosensors are considered to be the best option to bridge the existing technology gap, as they provide a fast, accurate, and real-time readout. However, the existing pyridine nucleotides genetically encoded fluorescent biosensors are either sensitive to pH change or slow in dissociation rate. Herein, we employed the biosensors which generate readouts that are pH stable for in planta measurement of NADH/NAD+ ratio and NADPH level. We generated transgenic Arabidopsis lines that express these biosensors in plastid stroma and cytosol of whole plants and pollen tubes under the control of CaMV 35S and LAT52 promoters, respectively. These transgenic biosensor lines allow us to monitor real-time dynamic changes in NADH/NAD+ ratio and NADPH level in the plastids and cytosol of various plant tissues, including pollen tubes, root hairs, and mesophyll cells, using a variety of fluorescent instruments. We anticipate that these valuable transgenic lines may allow improvements in plant redox biology studies.

Rice myo-inositol-3-phosphate synthase 2 (RINO2) alleviates heat injury-induced impairment in pollen germination and tube growth by modulating Ca2+ signaling and actin filament cytoskeleton.

Low phytic acid (lpa) crop is considered as an effective strategy to improve crop nutritional quality, but a substantial decrease in phytic acid (PA) usually has negative effect on agronomic performance and its response to environment adversities. Myo-inositol-3-phosphate synthase (MIPS) is the rate-limiting enzyme in PA biosynthesis pathway, and regarded as the prime target for engineering lpa crop. In this paper, the rice MIPS gene (RINO2) knockout mutants and its wild type were performed to investigate the genotype-dependent alteration in the heat injury-induced spikelet fertility and its underlying mechanism for rice plants being imposed to heat stress at anthesis. Results indicated that RINO2 knockout significantly enhanced the susceptibility of rice spikelet fertility to heat injury, due to the severely exacerbated obstacles in pollen germination and pollen tube growth in pistil for RINO2 knockout under high temperature (HT) at anthesis. The loss of RINO2 function caused a marked reduction in inositol and phosphatidylinositol derivative concentrations in the HT-stressed pollen grains, which resulted in the strikingly lower content of phosphatidylinositol 4,5-diphosphate (PI (4,5) P2) in germinating pollen grain and pollen tube. The insufficient supply of PI (4,5) P2 in the HT-stressed pollen grains disrupted normal Ca2+ gradient in the apical region of pollen tubes and actin filament cytoskeleton in growing pollen tubes. The severely repressed biosynthesis of PI (4,5) P2 was among the regulatory switch steps leading to the impaired pollen germination and deformed pollen tube growth for the HT-stressed pollens of RINO2 knockout mutants.

The Arabidopsis ARID-HMG DNA-BINDING PROTEIN 15 modulates JA signaling by regulating MYC2 during pollen development.

The intricate process of male gametophyte development in flowering plants is regulated by jasmonic acid (JA) signaling. JA signaling initiates with the activation of the basic-helix-loop-helix (bHLH) transcription factor (TF), MYC2, leading to the expression of numerous JA-responsive genes during stamen development and pollen maturation. However, the regulation of JA signaling during different stages of male gametophyte development remains less understood. This study focuses on the characterization of the plant ARID-HMG DNA-BINDING PROTEIN 15 (AtHMGB15), and its role in pollen development in Arabidopsis (Arabidopsis thaliana). Phenotypic characterization of a T-DNA insertion line (athmgb15-4) revealed delayed bolting, shorter siliques, and reduced seed set in mutant plants compared to the wildtype. Additionally, AtHMGB15 deletion resulted in defective pollen morphology, delayed pollen germination, aberrant pollen tube growth, and a higher percentage of non-viable pollen grains. Molecular analysis indicated the down-regulation of JA biosynthesis and signaling genes in the athmgb15-4 mutant. Quantitative analysis demonstrated that jasmonic acid and its derivatives were approximately tenfold lower in athmgb15-4 flowers. Exogenous application of methyl jasmonate could restore pollen morphology and germination, suggesting that the low JA content in athmgb15-4 impaired JA signaling during pollen development. Furthermore, our study revealed that AtHMGB15 physically interacts with MYC2 to form a transcription activation complex. This complex promotes the transcription of key JA signaling genes, the R2R3-MYB TFs MYB21 and MYB24, during stamen and pollen development. Collectively, our findings highlight the role of AtHMGB15 as a positive regulator of the JA pathway, controlling the spatiotemporal expression of key regulators involved in Arabidopsis stamen and pollen development.

Myo-inositol oxygenase CgMIOX3 alleviates S-RNase-induced inhibition of incompatible pollen tubes in pummelo.

Pummelo (Citrus grandis L. Osbeck) exhibits S-RNase-based self-incompatibility (SI), during which S-RNase cytotoxicity inhibits pollen tubes in an S-haplotype specific manner. The entry of S-RNase into self-pollen tubes triggers a series of reactions. However, these reactions are still poorly understood in pummelo. In the present study, we used S-RNases as baits to screen a pummelo pollen cDNA library and characterized a myo-inositol oxygenase (CgMIOX3) that physically interacts with S-RNases. CgMIOX3 is highly expressed in pummelo pollen tubes and its down-regulation leads to a reduction in pollen tube growth. Upon entering pollen tubes, S-RNases increase the expression of CgMIOX3 and enhance its activity by directly binding to it in an S-haplotype-independent manner. CgMIOX3 improves pollen tube growth under oxidative stress through ascorbic acid accumulation and increases the length of self-pollen tubes. Furthermore, over-expression of CgMIOX3 increases the relative length of self-pollen tubes growing in the style of petunia (Petunia hybrida). This study provides intriguing insights into the pumelo SI system, revealing a regulatory mechanism mediated by CgMIOX3 that plays an important role in the resistance of pollen tubes to S-RNase cytotoxicity.

A peptide-mediated, multilateral molecular dialogue for the coordination of pollen wall formation.

The surface of pollen grains is reinforced by pollen wall components produced noncell autonomously by tapetum cells that surround developing pollen within the male floral organ, the anther. Here, we show that tapetum activity is regulated by the GASSHO (GSO) receptor-like kinase pathway, controlled by two sulfated peptides, CASPARIAN STRIP INTEGRITY FACTOR 3 (CIF3) and CIF4, the precursors of which are expressed in the tapetum itself. Coordination of tapetum activity with pollen grain development depends on the action of subtilases, including AtSBT5.4, which are produced stage specifically by developing pollen grains. Tapetum-derived CIF precursors are processed by subtilases, triggering GSO-dependent tapetum activation. We show that the GSO receptors act from the middle layer, a tissue surrounding the tapetum and developing pollen. Three concentrically organized cell types, therefore, cooperate to coordinate pollen wall deposition through a multilateral molecular dialogue.

Phl p 5 levels more strongly associated than grass pollen counts with allergic respiratory health.

BACKGROUND: Studies have linked daily pollen counts to respiratory allergic health outcomes, but few have considered allergen levels. OBJECTIVE: We sought to assess associations of grass pollen counts and grass allergen levels (Phl p 5) with respiratory allergic health symptoms in a panel of 93 adults with moderate-severe allergic rhinitis and daily asthma hospital admissions in London, United Kingdom. METHODS: Daily symptom and medication scores were collected from adult participants in an allergy clinical trial. Daily counts of asthma hospital admissions in the London general population were obtained from Hospital Episode Statistics data. Daily grass pollen counts were measured using a volumetric air sampler, and novel Phl p 5 levels were measured using a ChemVol High Volume Cascade Impactor and ELISA analyses (May through August). Associations between the 2 pollen variables and daily health scores (dichotomized based on within-person 75th percentiles) were assessed using generalized estimating equation logistic models and with asthma hospital admissions using Poisson regression models. RESULTS: Daily pollen counts and Phl p 5 levels were each positively associated with reporting a high combined symptom and medication health score in separate models. However, in mutually adjusted models including terms for both pollen counts and Phl p 5 levels, associations remained for Phl p 5 levels (odds ratio [95% CI]: 1.18 [1.12, 1.24]), but were heavily attenuated for pollen counts (odds ratio [95% CI]: 1.00 [0.93, 1.07]). Similar trends were not observed for asthma hospital admissions in London. CONCLUSIONS: Grass allergen (Phl p 5) levels are more consistently associated with allergic respiratory symptoms than grass pollen counts.

New light on an old syndrome: Role of Api g 7 in mugwort pollen-related celery allergy.

BACKGROUND: Celery root is known to cause severe allergic reactions in patients sensitized to mugwort pollen. OBJECTIVE: We studied clinically well-characterized patients with celery allergy by IgE testing with a comprehensive panel of celery allergens to disentangle the molecular basis of what is known as the celery-mugwort syndrome. METHODS: Patients with suspected food allergy to celery underwent a standardized interview. Main inclusion criteria were a positive food challenge with celery or an unambiguous case history of severe anaphylaxis. IgE to celery allergens (rApi g 1.01, rApi g 1.02, rApi g 2, rApi g 4, nApi g 5, rApi g 6, rApi g 7) and to mugwort allergens (rArt v 1, rArt v 3, rArt v 4) were determined. IgE levels ≥0.35 kUA/L were regarded positive. RESULTS: Seventy-nine patients with allergy to celery were included. Thirty patients had mild oral or rhinoconjunctival symptoms, and 49 had systemic reactions. Sixty-eight percent had IgE to celery extract, 80% to birch pollen, and 77% to mugwort pollen. A combination of Api g 1.01, 1.02, 4, 5, and 7 increased the diagnostic sensitivity for celery allergy to 92%. The lipid transfer proteins Api g 2 and Api g 6 were not relevant in our celery-allergic population. IgE to Api g 7, detected in 52% of patients, correlated closely (r = 0.86) to Art v 1 from mugwort pollen. Eleven of 12 patients with monosensitization to Api g 7 were IgE negative to celery extract. The odds ratio for developing a severe anaphylactic reaction rather than only mild oral symptoms was about 6 times greater (odds ratio, 5.87; 95% confidence interval, 1.08-32.0; P = .0410) for Api g 7-sensitized versus -nonsensitized subjects. CONCLUSION: There is an urgent need for routine diagnostic tests to assess sensitization to Api g 7, not only to increase test sensitivity but also to identify patients at risk of a severe allergic reaction to celery.

IgE to cyclophilins in pollen-allergic children: Epidemiologic, clinical, and diagnostic relevance of a neglected panallergen.

BACKGROUND: Cyclophilins are ubiquitous panallergens whose epidemiologic, diagnostic, and clinical relevance is largely unknown and whose sensitization is rarely examined in routine allergy practice. OBJECTIVE: We investigated the epidemiologic, diagnostic, and clinical relevance of cyclophilins in seasonal allergic rhinitis and its comorbidities. METHODS: We examined a random sample of 253 (25%) of 1263 Italian children with seasonal allergic rhinitis from the Panallergens in Pediatrics (PAN-PED) cohort with characterized disease phenotypes. Nested studies of sensitization prevalence, correlation, and allergen extract inhibition were performed in patients sensitized to birch pollen extract but lacking IgE to Bet v 1/2/4 (74/1263) or with highest serum level of IgE to Bet v 1 (26/1263); and in patients with sensitization to various extracts (ragweed, mugwort, pellitory, Plantago, and plane tree), but not to their respective major allergenic molecule, profilins, and polcalcins. IgE to cyclophilin was detected with recombinant Bet v 7, and extract inhibition tests were performed with the same rBet v 7. RESULTS: IgE to rBet v 7 was detected in 43 (17%) of 253 patients. It was associated with asthma (P < .028) and oral allergy syndrome (P < .017) in univariate but not multivariate analysis adjusted for IgE to profilins (Phl p 12), PR-10s (Bet v 1), and lipid transfer proteins (Pru p 3). IgE to rBet v 7 was also highly prevalent (47/74, 63%) among patients with unexplained sensitization to birch pollen extract. In patients with unexplained sensitization to ragweed, mugwort, pellitory, Plantago and plane tree pollen, the levels of IgE to those extracts correlated with the levels of IgE to rBet v 7, and they were also significantly inhibited by rBet v 7 (inhibition range 45%-74%). CONCLUSIONS: IgE sensitization to cyclophilin is frequent in pollen-allergic patients living in temperate areas and can produce "false" positive outcomes in skin prick and IgE tests to pollen extracts. Molecular diagnostic guidelines should include this panallergen family.

Full-length EFOP3 and EFOP4 proteins are essential for pollen intine development in Arabidopsis thaliana.

Pollen development is critical to plant reproduction, but the underlying regulatory molecular mechanisms have not been fully elucidated. The Arabidopsis (Arabidopsis thaliana) EFR3 OF PLANT 3 (EFOP3) and EFR3 OF PLANT 4 (EFOP4) genes encode members of the Armadillo (ARM) repeat superfamily that play key roles in pollen development. Herein, we demonstrate that EFOP3 and EFOP4 are co-expressed in pollen at anther stages 10-12, but loss-of-function of both EFOP3 and EFOP4 leads to male gametophyte sterility, irregular intine, and shriveled pollen grains at anther stage 12. We further established that full-length EFOP3 and EFOP4 specifically localize to the plasma membrane, and the integrity of these proteins is essential for pollen development. We observed uneven intine, less organized cellulose and reduced pectin content in mutant pollen compared with the wild-type. These, together with the misexpression of several genes related to cell wall metabolism in efop3-/- efop4+/- mutants, suggest that EFOP3 and EFOP4 may indirectly regulate the expression of these genes to affect intine formation, thus controlling Arabidopsis pollen fertility in a functionally redundant manner. Moreover, transcriptome analysis showed that the absence of EFOP3 and EFOP4 function affects multiple pollen development pathways. These results enhance our understanding of EFOPs proteins and their role in pollen development.