Influence of nitrogen nutrition on the amount of hordein, protein Z and β-amylase messenger RNA in developing endosperms of barley

Abstract

Keywords: H o r d e u m vulgare L. cv. Bomi, m u t a n t Riso 56, c D N A , storage proteins, molecular hybridization, liquid culture of spikes, p r o l a m i n , grain filling, high-lysine m u t a n t , a l b u m i n , lysine-rich proteins, i m m u n o a f f i n i t y isolation, in vitro protein synthesisThe amount ofmRNA encoding hordein, protein Z and !3-amylase was analyzed in developing endosperms of the barley variety Bomi and the high-lysine mutant Riso 56. Spikes were cultured in liquid media containing different amounts of ammonium nitrate and endosperms harvested between 15 and 25 days after anthesis. Levels of mRNA were determined by in vitro translations. In addition, B hordein and protein Z mRNA species were titrated by molecular hybridization to cDNAs encoding B hordein and protein Z polypeptides, respectively. Increasing amounts ofmRNAs were found in response to increasing nitrogen application. The amount of mRNA for C hordein and protein Z increased linearly with increasing nitrogen concentrations in the medium, whereas the rise in IS-amylase mRNA was slight. A concentration of 0.5 gxl -~ nitrogen stimulated the transcription ofB hordein mRNA, but further increases of nitrogen supply up to 2 g• -~ resulted in little additional B hordein mRNA production. At high nitrogen levels C hordein and protein Z mRNA are preferentially transcribed whereby these proteins serve as major nitrogen sinks. Hordein and protein Z mRNA are effectively augmented by application of nitrogen after the onset of grain filling and storage protein synthesis. Removal of nitrogen in the medium during grain filling led to a short term decrease in the amounts of these mRNAs. Thus it is possible to modulate the mRNA levels by changing the nitrogen concentration in the medium. The absence of mRNA for B hordein polypeptides in the high-lysine mutant Riso 56, (hor 2 ca ), does not influence the effect of nitrogen on the expression of the C hordein, protein Z and [5-amylase genes.Abbreviations: cDNA = DNA complementary to mRNA; SDS-PAGE = sodium dodecyl sulphate polyacrylamide gel electrophoresis;Tm = melting temperature of the DNA Springer-Verlag

Monoclonal antibodies to hordein polypeptides

“…Horl with 3-6 members and Hor3 with only a few (5,14,19,37). The B-hordein polypeptides are by far the most abundant and may constitute 70% of total hordein, but the proportion of the three groups in the endosperm is highly influenced by nitrogen fertilization (21). The C-hordein polypeptides are preferentially synthesized when the level of nitrogen in the soil is high.…”

Hordein Gene Expression in a Low Protein Barley Cultivar

“…Since the Horl and Hor2 loci are multigene families and are separated by 8 cM (12), it is unlikely that an alteration in one of the structural genes could cause the observed reduction. Thus, the low hordein characteristic of Karl (2,10,14,15,20,21,24,25). In most ofthese studies, transcriptional control was proposed as the cause of low mRNA levels.…”

“…In most ofthese studies, transcriptional control was proposed as the cause of low mRNA levels. The abundance of hordein mRNAs has been shown to be influenced by the nutritional status of the plant (10,21). However, deficiencies of N or S have resulted in differential effects on the B and C hordein mRNAs.…”

“…These proteins are classified into three groups of polypeptides called B, C, and D hordeins, and each group is synthesized from a family of structural genes, Hor2, Horl, and Hor3, respectively (23). The B, C, and D hordeins account for approximately 80 to 90%, 10 to 20%, and <5% of the total hordein fraction, respectively (14). The B and D hordeins are relatively high in sulfur-containing amino acids compared to the C hordeins (23).…”

Mapping of theHor2 locus in barley by pulsed field gel electrophoresis

“…Furthermore, hordein mRNA levels respond differentially to changed levels of nutrients, C-hordein being preferentially expressed relative to Bhordein with increasing concentrations of nitrogen (19). A possible connection between this differential regulation of expression and the clustering of the hordein genes in multigenic loci has yet to be proven.…”