Temporal and Spatial Expression of the Major Allergens in Developing and Germinating Peanut Seed

Abstract

Peanut (Arachis hypogaea) seed proteins Ara h 1, Ara h 2, and Ara h 3 are considered to be the major peanut allergens. However, little is known about their temporal and spatial expression during seed development and upon germination and seedling growth. In this study, transcript levels of the three major peanut allergen genes, ara h 1, ara h 2, and ara h 3, and their corresponding proteins were found in all cultivars. Expression patterns were heterogeneous depending on the specific peanut allergen gene and the cultivars tested. However, ara h 3 expression patterns among the cultivars were more variable than ara h 1 and ara h 2. Transcripts were tissue specific, observed in seeds, but not in leaves, flowers, or roots, and were undetectable during seed germination. In situ hybridizations and immunotissue prints revealed that both embryonic axes and cotyledons expressed the allergens. However, more ara h 1 and ara h 3 messenger RNA was detected in cotyledons relative to embryonic axes. Allergen polypeptide degradation patterns were different in embryonic axes compared with cotyledons during germination and seedling growth, with levels of Ara h 1 and Ara h 2 dramatically reduced compared to the Ara h 3 polypeptides in embryonic axes. These characterization studies of major peanut allergen genes and their corresponding seed storage proteins can provide the basic information needed for biochemical and molecular approaches to obtain a hypoallergenic peanut.

Reporter Gene Expression Patterns Regulated by an Ara h 2 Promoter Differ in Homologous Versus Heterologous Systems1

“…Qu et al (2008) found prolamin and endospermspecific motifs in all but one rice glutelin gene promoter which controlled seed specific expression. Our results for reporter gene expression in peanut under the control of the Ara h 2 promoter largely conform with the results obtained by Kang et al (2007b) who showed that Ara h 2 polypeptides were abundant in mature seeds and diminished after germination during seedling growth. An unexpected result was the lack of reporter gene expression in embryo axes, although expression in cotyledons was strong.…”

“…An unexpected result was the lack of reporter gene expression in embryo axes, although expression in cotyledons was strong. Furthermore, we did not detect Ara h 2 transcripts in either 2-wk-or 8-wk-old leaf tissues even though reporter gene transcripts could be observed as faint RT-PCR products in the 2-wk-old samples, data which also are consistent with Kang et al (2007b) who could not detect Ara h 2 transcripts in leaves by Northern blot analysis. The present study also shows that a higher copy number of the transgene did not result in a difference in the steady state level of transcripts.…”

“…Also, the expression of Ara h 2 was reduced in embryonic axes compared to cotyledons in the post germination (seedling) growth stages. The Ara h 2 transcript was absent from the vegetative parts of the plant as confirmed by Northern blots and RT-PCR analysis (Kang et al, 2007b).…”

“…The two protein isoforms mainly differ by a 12-amino acid insertion in Ara h 2.02. Ara h 2 expression was shown to begin in immature seeds (stages 1 and 2) and peak as seeds matured (stages 3-4) (Kang et al, 2007b). Also, the expression of Ara h 2 was reduced in embryonic axes compared to cotyledons in the post germination (seedling) growth stages.…”

Identification of expressed sequences in the coffee genome potentially associated with somatic embryogenesis

“…Many PR10 proteins are constitutively expressed in various development stages and in seeds, roots (Sikorski et al, 1999), flowers, and leaves (Sikorski et al, 1999). Conversely, in Arachis hypogaea, 3 genes of allergenine (ara h1, ara h2, and ara h3) are exclusively expressed during seed development (Kang et al, 2007), and 2 homologs of the major allergen in Dacus carrota were highly expressed in the initial stages of embryogenesis and cellular suspension (Sano et al, 2004). Therefore, considering that contig 13686 was expressed in silico in a non-constitutive form, in lower intensities, in the libraries of leaves, plantlets, hypocotyls, and roots (AR1, LP1, CL2, FR4, RT5, and RT8) and in higher intensities in the embryogenic callus and cellular suspension libraries (CB1, CS1, PA1, and RT8), this contig may be involved in coffee plant embryogenesis.…”