The unique disarticulation layer formed in the rachis of Aegilops longissima probably results from the spatial co-expression of Btr1 and Btr2

Abstract

Background and aims: The brittle rachis trait is a feature of many wild grasses, particularly within the tribe Triticeae. Wild Hordeum and Triticum species form a disarticulation layer above the rachis node, resulting in the production of wedge-type dispersal units. In Aegilops longissima, only one or two of the nodes in the central portion of its rachis are brittle. In Triticeae species, the formation of a disarticulation layer above the rachis node requires the co-transcription of the two dominant and complementary genes Btr1 and Btr2. This study aims to establish whether homologues of Btr1 and/or Btr2 underlie the unusual brittle rachis phenotype observed in Ae. longissima.

Methods: Scanning electron microscopy was used to examine the disarticulation surfaces. Quantitative RT-PCR and RNA in situ hybridization experiments were used to identify gene expression in the immature inflorescence.

Key results: Analysis based on scanning electron microscopy was able to demonstrate that the disarticulation surfaces formed in the Ae. longissima rachis are morphologically indistinguishable from those formed in the rachises of wild Hordeum and Triticum species. RNA in situ hybridization showed that in the immature Ae. longissima inflorescence, the intensity of Btr1 transcription varied from high at the rachis base to low at its apex, while that of Btr2 was limited to the nodes in the central to distal portion of the rachis.

Conclusions: The disarticulation pattern shown by Ae. longissima results from the limitation of Btr1 and Btr2 co-expression to nodes lying in the centre of the rachis.

Keywords: Aegilops longissima; Btr1; Btr2; Seed dispersal; disarticulation zone; rachis; spatial co-expression.

Fig. 1.

The mature inflorescence of Aegilops longissima accession AE 417. The dispersal unit is the result of either (A) a single fracture, producing a spike-type unit, or (B) two fractures, producing a wedge-type unit and a spike-type unit comprising several spikelets. After disarticulation, a segment of the rachis, bearing a small number of spikelets, remains attached to the plant. The breakage points are indicated by the white boxes. (C,D) An enlarged view of the inflorescences shown in, respectively, A and B. Arrowheads indicate the points of disarticulation above a rachis node. Scale bar: A and B, 2 cm; C and D, 2 mm.

Fig. 2.

Scanning electron micrographs of the surface of the separation layer. The surface is (C) smooth in the central portion of the rachis exhibiting brittleness, but (A,E) broken and rough where the rachis is tough at both its (A) distal and (E) proximal portions. The images shown in B, D and F are close-up views of the sections in A, C and E highlighted by a white box. Scale bars: A, C, E, 100 μm; B, D, F, 20 μm.

Fig. 3.

Alignment of putative BTR polypeptide sequences encoded by Aegilops longissima accession AE 417. (A) BTR1 homologues, (B) BTR2 homologues.

Fig. 4.

Btr1 and Btr2 transcript abundance in the immature inflorescence of Aegilops longissima accession AE 417. (A) Btr1-Lo-1, (B) Btr1-Lo-2, (C) Btr1-Lo-3, (D) Btr2-Lo-1, (E) Btr2-Lo-2. FP: floret primordium stage, WA: white anther stage, GA: green anther stage, YA: yellow anther stage, WR: water ripe stage. Values are presented in the form mean ± s.e. (n = 3).

Fig. 5.

The localization of Btr1 and Btr2 transcript in inflorescences sampled at the white anther stage. The first to the sixth rachis node part of a rachis bearing 11 nodes is shown. (A) Btr1 transcript was detected in rachis cortex cells between the first and fourth nodes. The signal was concentrated in nodal tissue but was also detected in internodal tissue. (B) Btr2 transcript, as detected by the Btr2-Lo-1 probe, visible above the fourth, fifth and sixth rachis nodes, was also present between the seventh and 11th nodes (see Supplementary Data Fig. S3c). (C) Btr2 transcript, as detected by the Btr2-Lo-2 probe. The profile is the same as that detected by the Btr2-Lo-1 probe. Arrowheads indicate the site of a hybridization signal. Scale bar: 2 mm.