Japonica Pollen Research Articles

Effect of cigarette smoke on immune response to pollen allergen in the lung

Cigarette smoke (CS) is a major risk factor for pulmonary diseases. Alveolar macrophages (AM) are known to play an important role in the lung immune system. Cryptomeria Japonica pollen (CJp) is known as pollen allergen but not cleared to immune response in the lung. Also, it is not fully understood the relationship of CS and CJp on immune cells in the lung. Therefore, we investigated the effects of CS and CJp on immune cells response in the lung. Mice were exposed to CS during 10 days. The next day, mice were inhaled 600μg of CJp.

Plant Regeneration and In Vitro Growth Performance of Male-Sterile Somatic Plantlets of Sugi (Japanese Cedar, Cryptomeria japonica) Derived from Different Embryogenic Cell Lines

With the spread of pollinosis caused by sugi (Japanese cedar, Cryptomeria japonica) pollen, the use of pollen-free somatic seedlings of sugi is expected in Japan. However, there is a lack of knowledge on the relationship between the abilities during somatic embryogenesis, initial in vitro growth traits, and subsequent growth of somatic seedlings. In the present study, we provide the first basic information on somatic embryo maturation efficiency, somatic embryo germination, and plantlet conversion frequencies, as well as on in vitro growth performance of pollen-free somatic plantlets derived from different embryogenic cell lines (ECLs). Somatic embryo maturation efficiency varied from 34 to 514 cotyledonary embryos per plate and the average for the 19 ECLs tested was 244 embryos per plate. Subsequently, the overall average rates of somatic embryo germination and conversion among ECLs were 87.8% and 85.3%, respectively. The results of in vitro growth performance of pollen-free somatic plantlets showed significant differences in growth rate among ECLs.

Multi-point analysis of airborne Japanese cedar (Cryptomeria japonica D. Don) pollen by Pollen Robo and the relationship between pollen count and the severity of symptoms

An important theme related to aerobiology is the spatial distribution of airborne pollen, which is a final goal of our research efforts. In relation to this, pursuing the severity of pollinosis symptoms and problems with airborne pollen and allergens is required. In the past, we simulated the distribution of airborne Japanese cedar (Cryptomeria japonica D. Don) pollen (Kawashima and Takahashi in Grana 34:142–150, 1995, Grana 38:316–324, 1999). The simulation results of recent years will be published in the near future. At the time, it was not possible to compare the results of actual airborne pollen data observed from multiple points at the same time. However, now we can achieve this objective if using C. japonica pollen characteristics. The pollen is dispersed in large quantities over a long distance, remains in the air for a long time, and most airborne pollen at this time is C. japonica pollen. Recently, several automatic C. japonica pollen monitoring systems have been developed and deployed nationwide. Pollen Robo is one and is the most widely installed nationwide. In this study, we addressed Pollen Robo. First, we examined whether Pollen Robo can accurately count C. japonica pollen and found that Pollen Robo was useful during the major pollen season. However, it sometimes counts non-pollen particles and other pollen rather than C. japonica pollen. Therefore, it is not useful when a small amount of pollen is dispersed such as the start day of the pollen season. Next, we examined the daily dispersion of C. japonica pollen and it was shown that it is related to the distance between sampling locations. However, factors other than distance may be involved. Another matter of concern is the relationship between the number of airborne pollen and pollinosis symptoms. Symptoms began to appear from approximately the start day of the pollen season in moderate and light cases. The most serious symptoms occurred when the pollen count was the highest. Meanwhile, in severe cases, symptoms were observed at the time the pollen was scarcely detected and plateaued when the pollen count reached 100 counts/m3 on average over 6 days. Thus, Pollen Robo was not useful when symptoms began in severe cases. This problem was solved using pollen antigen measurement. The time for the symptoms to begin in severe cases is between 2 and 4 weeks before the start day of the pollen season. The amount of Cry j 1 on the start day during some years was considerably larger than that calculated from the number of pollen. The results obtained in this study will be the basic data used for achieving our final goal.

Relationship between Pollen Allergy (Allergic Rhinitis) and Cryptomeria-Specific Immunoglobulin E during Early Pregnancy: Part of a Nationwide Birth-Cohort Study

The Japan Environment and Children’s Study (JECS) measured serum concentrations of immunoglobulin E (IgE) antibodies against Cryptomeria japonica pollen during early pregnancy. Cryptomeria-sp...

Identification of Cha o 3 homolog Cry j 4 from Cryptomeria japonica (Japanese cedar) pollen: Limitation of the present Japanese cedar–specific ASIT

BackgroundAbout one-third of the Japanese population suffers from Japanese cedar pollinosis, which is frequently accompanied by Japanese cypress pollinosis. Recently, a novel major Japanese cypress pollen allergen, Cha o 3, was discovered. However, whether a Cha o 3 homolog is present in Japanese cedar pollen remains to be determined. MethodsWestern blot analysis was performed using Cha o 3–specific antiserum. In addition, cloning of the gene encoding Cry j 4 was conducted using total cDNA from the male flower of Japanese cedar trees. Allergen potency and cross-reactivity were investigated using a T-cell proliferation assay, basophil activation test, and ImmunoCAP inhibition assay. ResultsA low amount of Cha o 3 homolog protein was detected in Japanese cedar pollen extract. The deduced amino acid sequence of Cry j 4 showed 84% identity to that of Cha o 3. Cross-reactivity between Cry j 4 and Cha o 3 was observed at the T cell and IgE levels. ConclusionsCry j 4 was discovered as a counterpart allergen of Cha o 3 in Japanese cedar pollen, with a relationship similar to that between Cry j 1–Cha o 1 and Cry j 2–Cha o 2. Our findings also suggest that allergen-specific immunotherapy (ASIT) using Japanese cedar pollen extract does not induce adequate immune tolerance to Cha o 3 due to the low amount of Cry j 4 in Japanese cedar pollen. Therefore, ASIT using Cha o 3 or cypress pollen extract coupled with Japanese cedar pollen extract is required in order to optimally control allergy symptoms during Japanese cypress pollen season.

Enrichment of airborne Japanese cedar (Cryptomeria japonica) pollen in mountain ranges when passing through a front accompanying temperate low pressure

Values obtained by the real-time pollen monitors set up at 14 measuring points in the Tohoku region indicate that a large amount of Japanese cedar pollen was dispersed along the western region and northern flank of the Ou Mountain Range when a front accompanying the temperate low pressures passed through the west sea of northern Hokkaido. We are sure that this phenomenon is related to pollen enrichment by mountain ranges. The time at which highest concentrations of pollen grains were observed shifted from the western to the eastern side, in accordance with the path of the front. No pollen was seen after the passage of the cold front.

Size distribution of allergenic Cry j 2 released from airborne Cryptomeria japonica pollen grains during the pollen scattering seasons

Japanese cedar (Cryptomeria japonica) pollinosis is one of seasonal allergic rhinitis that mainly occurs in Japan. The pollinosis is caused by two main kinds of allergenic proteins called Cry j 1 and Cry j 2 which exist in Cryptomeria japonica pollen. In our previous study, we reported that the size-segregated of airborne fine allergenic Cry j 1 and morphological change of Cry j 1 due to the contact with rainfall. However, the study on airborne allergenic Cry j 2 in different particle sizes has not been identified until now. Therefore, the main aim of this study is to investigate the size distribution and scattering behavior of allergenic Cry j 2. The Cry j 2 particles were collected and determined in different particle sizes at the urban sampling points during the most severe pollination season of 2012 in Saitama, Japan. After the size-segregated Cry j 2 allergenic particles were collected using an Andersen high-volume (AHV) atmospheric sample, the airborne Cry j 2 concentrations were determined with a surface plasmon resonance (SPR) method. At the same time, the airborne Cryptomeria japonica pollens were also counted by the Durham pollen sampler. The higher concentrations of the allergenic Cry j 2 were detected even in particle sizes equal to or less than 1.1 μm (PM1.1) than other particle sizes. The airborne particles ranges from 0.06 to 11 μm were also collected by a low-pressure impactor (LPI) atmospheric sampler. After that, the concentrations of Cry j 2 allergenic particles in fine particle sizes were measured by the SPR method either. With the help of this study, we have confirmed the existence of fine daughter allergenic particles, which clearly differ from the parent pollen grains in size, especially after the rainy days. It is possible that the daughter allergenic species will be released from the fractions of cell wall and burst pollen grains. We concluded that rainwater was one of the important factors that affects the release of pollen allergenic proteins of both Cry j 1 and Cry j 2 from the parent pollen grains.

Longitudinal Investigation on Allergenic Conifer Pollen in Japan for Successful Prevention and Treatment against Japanese Cedar Pollinosis

Longitudinal Investigation on Allergenic Conifer Pollen in Japan for Successful Prevention and Treatment against Japanese Cedar Pollinosis In Japan allergic people has been recognized increasing according to the increase of Japanese cedar, Cryptomeria japonica (JC) pollinosis since first reported in 1964. We have detected allergenic airborne pollen when Japanese government has begun to investigate the background and treatment on JC pollinosis as a serious social problem since July1986. Urgently the gravitational method with Durham’s sampler has been adopted to know huge number of JC and cypress family, Cupressaceae family (Cypress) pollen grains as causative agent. The area of national artificial forest of over 31 years old has become about 7 times from1970. We have estimated annual fluctuation of Japanese important allergenic pollen, with climatic factor during about 30 years for successful treatment of pollinosis in near future. There are institutions in fifteen locations monitoring airborne pollen by Durham’s sampler in Japan between the latitude of 30 to 40 degrees north. Airborne pollen samples were collected at each institute and were sent to our hospital on a daily basis. We counted pollen grains per cm2 through microscope, classifying and summarizing them. From 1986 to 2014 we have referred to the change in meteorological condition at the close to 9 of the pollen monitoring locations and estimated statically. JC and Cypress pollen counts have shown huge but remarkable annual fluctuation and they have been increasing gradually according to over 31 y-old conifer planting area expanding. The mean temperature in summer time, JC male flower formation, has a significant correlation with next year pollen counts. The pollination season has started earlier in north and later in central Japan gradually through a changing on mean temperature. In Japan allergenic conifer pollen counts have been increasing because of over 31 y-old conifer plantations area increase through the climate change about 30 years. Moreover, it has changed the start of pollination season.

Structural Basis for the Specific Recognition of the Major Antigenic Peptide from the Japanese Cedar Pollen Allergen Cry j 1 by HLA-DP5

The major allergen, Cry j 1, was isolated from Japanese cedar Cryptomeria japonica (Cry j) pollen and was shown to react with immunoglobulin E antibodies in the sera from pollinosis patients. We previously reported that the frequency of HLA-DP5 was significantly higher in pollinosis patients and the immunodominant peptides from Cry j 1 bound to HLA-DP5 to activate Th2 cells. In the present study, we determined the crystal structure of the HLA-DP5 heterodimer in complex with a Cry j 1-derived nine-residue peptide, at 2.4Å resolution. The peptide-binding groove recognizes the minimal peptide with 10 hydrogen bonds, including those between the negatively charged P1 pocket and the Lys side chain at the first position in the peptide sequence. We confirmed that HLA-DP5 exhibits the same Cry j 1-binding mode in solution, through pull-down experiments using structure-based mutations of Cry j 1. We also identified the characteristic residues of HLA-DP5 that are responsible for the distinct properties of the groove, by comparing the structure of HLA-DP5 and the previously reported structures of HLA-DP2 in complexes with pDRA of the self-antigen. The comparison revealed that the HLA-DP5·pCry j 1 complex forms several hydrogen bond/salt bridge networks between the receptor and the antigen that were not observed in the HLA-DP2·pDRA complex. Evolutionary considerations have led us to conclude that HLA-DP5 and HLA-DP2 represent two major groups of the HLA-DP family, in which the properties of the P1 and P4 pockets have evolved and acquired the present ranges of epitope peptide-binding specificities.

Size-segregated Allergenic Particles Released from Airborne Cryptomeria japonica Pollen Grains during the Yellow Sand Events within the Pollen Scattering Seasons

Cryptomeria japonica pollen is the most common pollen, which are scattering during each spring season in Japan. Japanese cedar (Cryptomeria japonica) pollinosis is one of seasonal allergic rhinitis that mainly occurs in Japan. In addition, long range transportation of Yellow Sand from the East Asian continent was also found during the pollen scattering seasons in Japan. Therefore, the interaction or impact between pollen and Yellow Sand should be concerned. In this study, our objective was to investigate the airborne behaviour of Cryptomeria japonica pollen grains and its size-segregated allergenic (Cry j 1) particles as the airborne tracer of Cryptomeria japonica pollen during the Yellow Sand events. Airborne Cryptomeria japonica pollen grains and its size-segregated allergenic particles were collected at roadside of urban residential zones of Saitama city during the pollination periods from February to March in two year investigation of 2009 and 2010. The overlap of Yellow Sand events and dispersal peak of pollen grains was observed. According to the Meteorological data, we found that the peaks of airborne pollen grains appeared under higher wind speed and temperature than the previous day. It was thought that Yellow Sand events and airborne pollen counts were related to wind speed. From the investigation of the airborne behavior of the size-segregated allergen particles by determining Cry j 1 with Surface Plasmon Resonance (SPR), the higher concentrations of the allergenic Cry j 1 were detected in particle size equal to or less than 1.1 μm(PM 1.1 ) than other particle sizes during Yellow Sand events, especially in the rainy day. We conclude that rainwater trapping Yellow Sand is one of the important factors that affect the release of allergenic pollen species of Cry j 1. Therefore, it is very important to clarify the relationships between Cryptomeria japonica pollen allergenic species and chemical contents of the Yellow Sand particles in further studies.

Cystatin SN Upregulation in Patients with Seasonal Allergic Rhinitis

Seasonal allergic rhinitis (SAR) to the Japanese cedar, Cryptomeria japonica (JC) pollen is an IgE-mediated type I allergy affecting nasal mucosa. However, the molecular events underlying its development remain unclear. We sought to identify SAR-associated altered gene expression in nasal epithelial cells during natural exposure to JC pollen. We recruited study participants in 2009 and 2010 and collected nasal epithelial cells between February and April, which is the period of natural pollen dispersion. Fifteen patients with SAR-JC and 13 control subjects were enrolled in 2009, and 17 SAR-JC patients, 13 sensitized asymptomatic subjects (Sensitized), and 15 control subjects were enrolled in 2010. Total RNA was extracted from nasal epithelial cells and 8 SAR-JC patients and 6 control subjects in 2009 were subjected to microarray analysis with the Illumina HumanRef-8 Expression BeadChip platform. Allergen-stimulated histamine release was examined in the peripheral blood basophils isolated from patients with SAR. We identified 32 genes with significantly altered expression during allergen exposure. One of these, CST1 encodes the cysteine protease inhibitor, cystatin SN. CST1 expression in nasal epithelial cells was significantly upregulated in both the 2009 and 2010 SAR-JC groups compared with the control groups. Immunohistochemical staining confirmed the increased expression of CST1 in the nasal epithelial cells of SAR patients. Addition of exogenous CST1 to basophils inhibited JC allergen-stimulated histamine release in vitro. We propose that CST1 may contribute to inactivation of protease allergens and help re-establish homeostasis of the nasal membranes.

Diurnal and Nocturnal Behaviour of Airborne Cryptomeria japonica Pollen Grains and the Allergenic Species in Urban Atmosphere of Saitama, Japan

Japanese cedar (Cryptomeria japonica) pollinosis is the most popular pollinosis in Japan. It has been reported that Cryptomeria japonica pollen allergenic species are suspended as fine particles in the urban atmosphere. These allergenic fine particles are responsible for inducing asthma by breaking into the lower respiratory tract. It has also been found that pollinosis symptoms on the sufferers appear mainly at night-time by the results from epidemiological studies. However, the exact reason for these phenomena is not yet clarified. In this study, the diurnal and nocturnal behaviours of Cryptomeria japonica pollen grains and their allergenic species in the urban area of Saitama city of Kanto Plain were investigated. Airborne pollen grains and allergenic Cry j 1 concentrations in total suspended particulate matter (TSP) were investigated at two sampling sites, a heavy traffic road (roadside site) and at the balcony of the <TEX>$10^{th}$</TEX> floor of the Building of Research and Project of Saitama University (general urban site). The latter sampling site where located about 300 m away from the roadside site was used as a general urban site unaffected by automobile traffic. The airborne pollen counts were measured with a real-time pollen monitor. Cry j 1 particles were collected with two high volume air samplers, and these concentrations were measured by surface plasmon resonance method with a Biacore J system. The diurnal variation of the airborne pollen counts was similar to the trends of temperature and wind speed during the day-time; whereas its tendency with wind speed trend was not observed during the night-time. Airborne pollen counts were lower with northern wind than with southern wind because the pollen comes from the mountainous areas, and the mountains in the south are closer, about half the distance to the northern mountains. It is suggested that the peaks of airborne pollen counts during night-time in the sampling site occurred by transport of pollen grains released during day-time in the mountainous forest areas, located c.a. 100 km away from the sampling site. On the roadside site the allergenic Cry j 1 concentrations were higher than at the general urban site, nevertheless pollen grains counts were lower. These results suggested that worsening of pollinosis symptoms during night-time in urban area was caused by transport of pollen grains during day-time in the mountainous forest areas. Moreover, pollen allergenic species become different morphology from pollen grain at roadside site, and the subsequent pollen grains re-suspension by automobile traffic.

Changes in quantity of Cry j1, a major cedar pollinosis allergen, in Cryptomeria japonica pollen during spring in Japan

The changes in quantity of Cry j1 in Cryptomeria japonica pollen were assessed during the course of spring in Japan. C. japonica pollen was collected from the same trees in Hiroshima and Yamanashi Prefectures, and Cry j1 was quantified by enzyme-linked immunosorbent assay. The quantities of Cry j1 were found to gradually decrease, and this trend was similar in both Hiroshima and Yamanashi Prefectures. In Yamanashi Prefecture, no significant differences in the quantities of Cry j1 between the upper and lower parts of the tree were observed in samples collected from March 1 to 14. Furthermore, the quantities of Cry j1 on March 1, 5, 10 and 14 were significantly lower than those on February 1. These results suggest that expression of C. japonica pollen levels in terms of particle numbers is not sufficient and that quantities of Cry j1 vary throughout the course of spring.

A microfluidic platform for the investigation of elongation growth in pollen tubes

Pollen tubes are an excellent model for the investigation of plant cell growth: they elongate at very high rates and are easily cultured in vitro. One major constraint in the study of pollen tube growth has been the difficulty in providing an in vitro testing environment that physically resembles the in vivo conditions. This work presents the development of a microfluidic platform for the study and manipulation of individual pollen tubes. The platform is fabricated from polydimethylsiloxane using a Silicon/SU-8 mold and makes use of microfluidics to distribute pollen grains to serially arranged microchannels into which the tubes grow to allow for individual testing. A 2D finite element fluid analysis is done to assist optimization of the architectural design. Validation of the device is carried out by growing Camellia japonica pollen. Results show that pollen tube germination and growth rate within the microfluidic network are similar to those obtained in conventional plate or batch assays. The microfluidic network allows for specific testing of a variety of structural features as demonstrated with a simple collision test, and it permits the straightforward integration of further single-cell test assays.

Inflammatory and degranulation effect of yellow sand on RBL-2H3 cells in relation to chemical and biological constituents

AbstractRecent studie pointed out that allergic diseases have increased during the Asian dust storm event (ADSE) in Japan. Daily observations and the atmospheric concentrations of yellow sand (YS) aerosol have been increasing. In this study, YS samples collected from three sites of Japan during ADSE in 2009–2010 were used. The particles were analyzed by X-ray photoelectron spectroscopy (XPS) and X-ray fluorescence—energy dispersive spectrometer (XRF-EDS). We investigate ability of YS extract on enhancing the chemical mediator release and cytokine production from rat basophilic leukemia (RBL-2H3) cells. The dust particles at Fukuoka and Tsukuba were abundant in aluminum (Al), iron (Fe), potassium (K) and titan (Ti) than those at Naha. Concentration of the trace endotoxin and Cryptomeria japonica pollen allergen (Cry j 1) were measured in YS extract. After exposure of RBL-2H3 cells to YS extract, the β-hexosaminidase (β-hex) release, tumor necrosis factor-alpha (TNF-α) production were enhanced in RBL-2H3 cells. This process depends on endotoxin, Cry j 1 and other allergen present in the YS extract. YS water extract also show a strong cytotoxic effect on the cells. This data suggest that low levels of endotoxin and Cry j 1 in YS may cause allergy during the ADSE.

IgE reactivity to a Cry j 3, an allergen of Japanese cedar (Cryptomeria japonica) pollen in dogs with canine atopic dermatitis

The present study investigated IgE reactivity to a new Cryptomeria japonica pollen allergen (Cry j 3) in dogs with atopic dermatitis by using a fluorometric ELISA. Serum samples from 15 dogs that showed IgE sensitivity to crude C. japonica pollen allergen by ELISA were tested for specific IgE to each allergen, individually. All 15 dogs had anti-Cry j 1 IgE, 6 (40%) had anti-Cry j 2 IgE, and 11 (73%) had anti-Cry j 3 IgE. Further, we found that these anti-Cry j 3 IgE reacted to Cry j 3 with immunoblotting analysis. These findings indicate that Cry j 3 may be a major allergen in dogs.

Release behavior of small sized daughter allergens from Cryptomeria japonica pollen grains during urban rainfall event

In Japan, Cryptomeria japonica pollen (with diameter ~30 μm) is scattered during each spring season. Daughter allergenic particles, which are smaller in size than their parent pollen grain and are abundant in fine particles (the particle sizes 7.0 μm) was poor. The possible reason may be pollen burst at high humidity before rainfall. Additionally, Cry j 1 contents were emitted from the so-called Ubisch body, which contains allergenic Cry j 1 abundantly when pollen was in contact with rainfall. In particular, we found that 60% of allergenic Cry j 1 contents released in air polluted rainfall contained Ca2+ ion derived from road dust and ADS. Therefore, rainfall should be a main factor to induce transition of pollen allergenic contents to fine particles. In conclusion, allergenic particles which are small sized and translated into fine particles by rainfall can be inhaled into the lower respiratory tract and contribute to the hypersensitivity of asthma.

Fullerene fine particles adhere to pollen grains and affect their autofluorescence and germination

Adhesion of commercially produced fullerene fine particles to Cryptomeria japonica, Chamaecyparis obtusa and Camellia japonica pollen grains was investigated. The autofluorescence of pollen grains was affected by the adhesion of fullerene fine particles to the pollen grains. The degree of adhesion of fullerene fine particles to the pollen grains varied depending on the type of fullerene. Furthermore, germination of Camellia japonica pollen grains was inhibited by the adhesion of fullerene fine particles.

A New Lipid Transfer Protein Homolog Identified as an IgE-Binding Antigen from Japanese Cedar Pollen

Japanese cedar (Cryptomeria japonica) pollen is a major cause of seasonal rhinitis and conjunctivitis in Japan, and an understanding of its full allergen repertoire is prerequisite for the development of future molecular diagnostics and immunotherapeutic strategies. Here we report the identification of a new C. japonica pollen IgE-binding antigen (CJP-8) homologous to lipid transfer proteins (LTPs), a class of plant cross-reactive allergens found in foods, latex, and pollen grains. The cjp-8 cDNA encodes a 165-amino acid polypeptide possessing the conserved eight cysteines characteristic of plant LTP family members. Escherichia coli-expressed recombinant CJP-8 (r-CJP-8) reacted with IgE antibody from Japanese cedar pollinosis patients at a 37.5% frequency (6/16).

Molecular Cloning and Immunochemical Characterization of a Novel Major Japanese Cedar Pollen Allergen Belonging to the Aspartic Protease Family

Background:Japanese cedar (Cryptomeria japonica) pollen is a major cause of seasonal pollinosis in Japan. Protease activity in the pollen grains may trigger pro-allergic responses but no such proteases have yet been identified as pollen allergens. Objectives:We report the molecular cloning and immunochemical characterization of a novel C. japonica pollen allergen belonging to the aspartic protease family. Methods:We focused on the C. japonica pollen allergen spot No. 63 (CPA63, 47.5% IgE binding frequency) on our 2-dimensional IgE immunoblot map. The internal amino acid sequences were determined using time-of-flight mass spectrometry. Full-length cpa63 cDNA was cloned by rapid amplification of cDNA ends (RACE)-PCR. Recombinant CPA63 (r-CPA63) was expressed using the baculovirus-insect cell culture system and its IgE binding capacity was analyzed by enzyme-linked immunosorbent assay (ELISA). The proteolytic activity of r-CPA63 was also assessed using a putative mature enzyme produced upon autolysis. Results: cpa63 cDNA encoded a 472 amino acid polypeptide showing about 40% sequence identity to members of the plant atypical aspartic protease family. ELISA showed that r-CPA63 was recognized by IgE antibodies in the serum of 58% (18/31) of Japanese cedar pollinosis patients. We also demonstrated an aspartic protease-like enzyme activity of the putative mature r-CPA63. Conclusions:We have identified the first plant aspartic protease allergen from Japanese cedar pollen. The availability of the CPA63 sequence and its recombinant allergen production system are useful not only for pharmaceutical applications but also for further examination of the role of protease activity in the pathogenesis of cedar pollinosis.