HighlightSeed coat mucilage composition and heteroxylan structure varies between different Plantago species and is accompanied by differences in glycosyltransferase family 61 (GT61) copy number and transcript abundance.

Upon exposure to aqueous environments, polysaccharides present in the seed coats of myxospermous species swell and extrude forming a halo of mucilage that completely envelops the seed. The extruded mucilage is usually a composite of pectic, non‐cellulosic, and cellulosic polysaccharides, and because of its accessibility the seed mucilage system has been used extensively in the laboratory to uncover molecular mechanisms associated with plant cell wall (PCW) polysaccharide biosynthesis. The mucilage of Arabidopsis , which is predominantly composed of pectic polysaccharides, is the most extensively characterised. Interrogation of this system has revealed intricate molecular and chemical details about polysaccharide biosynthesis and seed coat development, clearly demonstrating the effectiveness of using seed mucilage systems as a proxy to study PCW polysaccharide biosynthesis. Significant advances have been made in our understanding and identification of the genes and enzymes involved in pectin, cellulose, and xylan biosynthesis. Recent results regarding the composition, combined with the definition of the architectural properties, of extruded seed mucilage are synergistic with our current models of how polysaccharides interact in the PCW. In addition to the desirable characteristics of myxospermy for the study of PCW polysaccharide biosynthesis, industrial applications of seed mucilages are extensive. Strong interest from the food manufacturing and processing, pharmaceutical, cosmetic, and waste management industries has also encouraged researchers to investigate the physiochemical properties of seed mucilages and their possible applications. This article highlights the recent advances made in our understanding of the molecular mechanisms and the architectural properties of mucilage biosynthesis across diverse species.

Seeds from the myxospermous species Plantago ovata release a polysaccharide-rich mucilage upon contact with water. This seed coat derived mucilage is composed predominantly of heteroxylan (HX) and is utilized as a gluten-free dietary fiber supplement to promote human colorectal health. In this study, a gamma-irradiated P. ovata population was generated and screened using histological stains and Fourier Transform Mid Infrared (FTMIR) spectroscopy to identify putative mutants showing defects in seed coat mucilage HX composition and/or structure. FTMIR analysis of dry seed revealed variation in regions of the IR spectra previously linked to xylan structure in Secale cereale (rye). Subsequent absorbance ratio and PCA multivariate analysis identified 22 putative mutant families with differences in the HX IR fingerprint region. Many of these showed distinct changes in the amount and subtle changes in structure of HX after mucilage extrusion, while 20% of the putative HX mutants identified by FTMIR showed no difference in staining patterns of extruded mucilage compared to wild-type. Transcriptional screening analysis of two putative reduced xylan in mucilage (rxm) mutants, rxm1 and rxm3, revealed that changes in HX levels in rxm1 correlate with reduced transcription of known and novel genes associated with xylan synthesis, possibly indicative of specific co-regulatory units within the xylan biosynthetic pathway. These results confirm that FTMIR is a suitable method for identifying putative mutants with altered mucilage HX composition in P. ovata, and therefore forms a resource to identify novel genes involved in xylan biosynthesis.

Seed mucilage polysaccharide production, storage and release in Plantago ovata is strikingly different to that of the model plant Arabidopsis. We have used microscopy techniques to track the development of mucilage secretory cells and demonstrate that mature P. ovata seeds do not have an outer intact cell layer within which the polysaccharides surround internal columellae. instead, dehydrated mucilage is spread in a thin homogenous layer over the entire seed surface and upon wetting expands directly outwards, away from the seed. observing mucilage expansion in real time combined with compositional analysis allowed mucilage layer definition and the roles they play in mucilage release and architecture upon hydration to be explored. The first emergent layer of hydrated mucilage is rich in pectin, extremely hydrophilic, and forms an expansion front that functions to 'jumpstart' hydration and swelling of the second layer. this next layer, comprising the bulk of the expanded seed mucilage, is predominantly composed of heteroxylan and appears to provide much of the structural integrity. Our results indicate that the synthesis, deposition, desiccation, and final storage position of mucilage polysaccharides must be carefully orchestrated, although many of these processes are not yet fully defined and vary widely between myxospermous plant species. Abbreviations DPA Days post-anthesis ML Mucilage layer MSC Mucilage secretory cell SEM Scanning electron microscopy Upon exposure to aqueous environments, seeds from myxospermous species extrude a polysaccharide-rich gel from their seed surface, often called mucilage. Numerous species display myxospermy and there are a range of possible evolutionary advantages of synthesising such a carbon-rich and energy-expensive substance 1. Of all myxospermous species, the seed mucilage system of Arabidopsis is the best characterised. Arabidopsis seed mucilage has been used extensively as a proxy for the study of plant cell wall polysaccharide biosynthesis, enabling increased molecular characterisation of pectin biosynthesis, its main polysaccharide component 2 , as well as the biosynthesis of cellulose 3,4 and several hemicelluloses 5-8 , which are minor but integral components. Mucilage from other species can be highly diverse 1 and while P. ovata mucilage is also a complex mixture of polymers, it is predominantly heteroxylan with only a minor pectin component. While the pectin component is a near-linear rhamnogalacturonan 9-11 , the P. ovata heteroxylan (accounting for around 90% of the mucilage polysaccharides) is highly complex with the current scientific consensus defining P. ovata heteroxylan comprising a β-(1,4)-linked-d-xylopyranose backbone, heavily substituted at O-2 and/or O-3 positions with various mono-, di-and oligosaccharide substitutions of α-l-arabinofuranose and β-d-xylopyranose 9,11,12. It is likely that, as with other eudicots, the β-(1,4)-linked-d-xylopyranose backbone is synthesised by several members of

Plantago ovata is cultivated for production of its seed husk (psyllium). When wet, the husk transforms into a mucilage with properties suitable for pharmaceutical industries, utilised in supplements for controlling blood cholesterol levels, and food industries for making gluten-free products. There has been limited success in improving husk quantity and quality through breeding approaches, partly due to the lack of a reference genome. Here we constructed the first chromosome-scale reference assembly of P. ovata using a combination of 5.98 million PacBio and 636.5 million Hi-C reads. We also used corrected PacBio reads to estimate genome size and transcripts to generate gene models. The final assembly covers ~ 500 Mb with 99.3% gene set completeness. A total of 97% of the sequences are anchored to four chromosomes with an N50 of ~ 128.87 Mb. The P. ovata genome contains 61.90% repeats, where 40.04% are long terminal repeats. We identified 41,820 protein-coding genes, 411 non-coding RNAs, 108 ribosomal RNAs, and 1295 transfer RNAs. This genome will provide a resource for plant breeding programs to, for example, reduce agronomic constraints such as seed shattering, increase psyllium yield and quality, and overcome crop disease susceptibility.