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Xylem vessel pits are tiny openings on the cell wall of water-conducting cells—with pit geometry influencing crop yield through its effect on plant hydraulics and nitrogen transport.

Researchers from the Institute of Genetics and Developmental Biology (IGDB) of the Chinese Academy of Sciences have now revealed the complete three-dimensional (3D) ultrastructure of xylem vessel pits in rice and identified a critical regulatory module, MYB61-PS1, that shapes the 3D structure of pits in rice.

The findings, in Cell on October 14, demonstrate that the 3D geometry of vessel pits is precisely controlled through xylan deacetylation around pit borders, which is mediated by the molecular module MYB61-PS1. In turn, the optimized geometry improves yield by sustaining vessel hydraulics and facilitating nitrogen transport.

Using scanning electron microscopy, the researchers, led by Profs. Zhang Baocai and Zhou Yihua, investigated vessel pit variations in rice core accessions, discovering a wide range of pit-size variation. Whole-genome-wide association analysis identified PS1 as the major quantitative trait locus (QTL) gene controlling pit size. PS1 encodes a xylan deacetylase, and its role in controlling pit size and xylem hydraulics has been genetically validated.

To clarify the structural details of pits shaped by PS1 alleles, the researchers applied focused ion beam–scanning electron microscopy to reconstruct the complete 3D pit structure at the nanoscale, revealing complex curvature of the pit cavity structure. Furthermore, the narrowest part (pit aperture) was identified as a determinant of xylem hydraulics. Notably, they discovered that PS1 Hap2 (haplotype 2) generates smaller pit apertures compared with PS1Hap1 (haplotype 1), leading to higher xylem transport efficiency.

To elucidate the biological function of PS1, the researchers developed a rapid screening system to obtain the antibodies capable of distinguishing different xylan acetylation states in situ and discovered that the xylans clustered at the pit borders are low-acetylated, which was corroborated by super-resolution microscopy and label-free in situ analysis of high-resolution stimulated Raman microscopy.

Serial assays showed that PS1 Hap2 exhibits stronger deacetylase activity than PS1 Hap1 due to an amino acid variation (V163A). PS1 Hap2 proteins produce xylans with lower acetylation level, which bind cellulose with higher affinity, thus facilitating the packing of cellulosic nanofibers at the pit borders to form pits with smaller apertures.

Further nitrogen treatment assays revealed that rice plants harboring PS1 Hap2 displayed significantly improved nitrogen transport efficiency, leading to increased grain yield under different nitrogen conditions. PS1 Hap2 is thus an elite haplotype. Haplotype network analysis revealed an obvious divergence in PS1 between the two major rice subspecies indica and japonica. Hap2 has been increasingly utilized in modern indica varieties but rarely used in the breeding of japonica cultivars.

In addition, the genetic introduction of PS1 Hap2 into the two japonica varieties, Wuyujing 3 and Wumijing, resulted in a significant increase in grain yield, indicating its great potential in breeding.

Unexpectedly, the researchers found that nitrogen regulates pit size through PS1 under the control of MYB61, a QTL gene of nitrogen utilization efficiency. Pyramiding both elite alleles can further optimize pit geometry and thereby significantly increase grain yield in rice.

This study has identified PS1 as the first QTL gene controlling pit size in xylem vessels and revealed the molecular mechanism by which PS1 deacetylates xylans destined for pit borders to orchestrate cellulose-xylan crosslinking and nanofiber assembly, thereby shaping the pits and consequently regulating xylem transport and grain yield in rice. It also inspires new thinking on how signaling molecules regulate the plasticity of vessel cell structure.

The findings provide a multiscale research strategy, covering molecular levels from genes to polysaccharide macromolecules, from subcellular ultrastructure to cells and tissues, and ultimately entire plants, thereby facilitating technological innovation for sustainable agriculture.