Brassinosteroid-insensitive dwarf mutants of Arabidopsis accumulate brassinosteroids

Abstract

Seven dwarf mutants resembling brassinosteroid (BR)-biosynthetic dwarfs were isolated that did not respond significantly to the application of exogenous BRs. Genetic and molecular analyses revealed that these were novel alleles of BRI1 (Brassinosteroid-Insensitive 1), which encodes a receptor kinase that may act as a receptor for BRs or be involved in downstream signaling. The results of morphological and molecular analyses indicated that these represent a range of alleles from weak to null. The endogenous BRs were examined from 5-week-old plants of a null allele (bri1-4) and two weak alleles (bri1-5 and bri1-6). Previous analysis of endogenous BRs in several BR-biosynthetic dwarf mutants revealed that active BRs are deficient in these mutants. However, bri1-4 plants accumulated very high levels of brassinolide, castasterone, and typhasterol (57-, 128-, and 33-fold higher, respectively, than those of wild-type plants). Weaker alleles (bri1-5 and bri1-6) also accumulated considerable levels of brassinolide, castasterone, and typhasterol, but less than the null allele (bri1-4). The levels of 6-deoxoBRs in bri1 mutants were comparable to that of wild type. The accumulation of biologically active BRs may result from the inability to utilize these active BRs, the inability to regulate BR biosynthesis in bri1 mutants, or both. Therefore, BRI1 is required for the homeostasis of endogenous BR levels.

Figure 1

Wild type and four representative alleles of bri1 at 5 weeks of age. A, Wild type; B, bri1-3; C, bri1-4; D, bri1-5; and E, bri1-6.

Figure 2

Schematic of the BRI1 locus including the positions of the bri1 mutations. aa, Amino acid.

Figure 3

The proposed brassinolide biosynthetic pathway and the quantification of endogenous sterols and BRs from bri1-4 (a null allele), bri1-5 (a weaker allele), and wild type (Ws-2). Values in top, middle, and bottom represent endogenous levels (per gram fresh weight) in bri1-4, bri1-5, and the wild type, respectively. Most of the data for the wild type have been already published (Choe et al., 1999b). Data quantifying teasterone were not available in our previous study because of low recovery. In this study, we repeated the analysis using the same plant materials. Endogenous teasterone was not detected, while recovery of the internal standard ([²H₆]teasterone) was very good.