The U-box protein family of ubiquitin ligases is important in the biological processes of plant growth, development, and biotic and abiotic stress responses. Plants in the genus Zoysia are recognized as excellent warm-season turfgrass species with drought, wear and salt tolerance. In this study, we conducted the genome-wide identification of plant U-box (PUB) genes in Zoysia japonica based on U-box domain searching. In total, 71 ZjPUB genes were identified, and a protein tree was constructed of AtPUBs, OsPUBs, and ZjPUBs, clustered into five groups. The gene structures, characteristics, cis-elements and protein interaction prediction network were analyzed. There were mainly ABRE, ERE, MYB and MYC cis-elements distributed in the promoter regions of ZjPUBs. ZjPUBs were predicted to interact with PDR1 and EXO70B1, related to the abscisic acid signaling pathway. To better understand the roles of ZjPUBs under salt stress, the expression levels of 18 ZjPUBs under salt stress were detected using transcriptome data and qRT-PCR analysis, revealing that 16 ZjPUBs were upregulated in the roots under salt treatment. This indicates that ZjPUBs might participate in the Z. japonica salt stress response. This research provides insight into the Z. japonica PUB gene family and may support the genetic improvement in the molecular breeding of salt-tolerant zoysiagrass varieties.

Stenotaphrum secundatum is an excellent shade-tolerant warm-season turfgrass. Its poor cold resistance severely limits its promotion and application in temperate regions. Mining cold resistance genes is highly important for the cultivation of cold-resistant Stenotaphrum secundatum. Although there have been many reports on the role of the Shaker potassium channel family under abiotic stress, such as drought and salt stress, there is still a lack of research on their role in cold resistance. In this study, the transcriptome database of Stenotaphrum secundatum was aligned with the whole genome of Setaria italica, and eight members of the Shaker potassium channel family in Stenotaphrum secundatum were identified and named SsKAT1.1, SsKAT1.2, SsKAT2.1, SsKAT2.2, SsAKT1.1, SsAKT2.1, SsAKT2.2, and SsKOR1. The KAT3-like gene, KOR2 homologous gene, and part of the AKT-type weakly inwardly rectifying channel have not been identified in the Stenotaphrum secundatum transcriptome database. A bioinformatics analysis revealed that the potassium channels of Stenotaphrum secundatum are highly conserved in terms of protein structure but have more homologous members in the same group than those of other species. Among the three species of Oryza sativa, Arabidopsis thaliana, and Setaria italica, the potassium channel of Stenotaphrum secundatum is more closely related to the potassium channel of Setaria italica, which is consistent with the taxonomic results of these species belonging to Paniceae. Subcellular location experiments demonstrate that SsKAT1.1 is a plasma membrane protein. The expression of SsKAT1.1 reversed the growth defect of the potassium absorption-deficient yeast strain R5421 under a low potassium supply, indicating that SsKAT1.1 is a functional potassium channel. The transformation of SsKAT1.1 into the cold-sensitive yeast strain INVSC1 increased the cold resistance of the yeast, indicating that SsKAT1.1 confers cold resistance. The transformation of SsKAT1.1 into the salt-sensitive yeast strain G19 increased the resistance of yeast to salt, indicating that SsKAT1.1 is involved in salt tolerance. These results suggest that the manipulation of SsKAT1.1 will improve the cold and salt stress resistance of Stenotaphrum secundatum.

Owing to the poor cold tolerance of Stenotaphrum Trin and the urgent need for shade-tolerant grass species in temperate regions of East China, this study evaluated the cold tolerance of 55 Stenotaphrum accessions, aiming to provide shade-tolerant materials for temperate regions. A fine cold-tolerant turfgrass should have both the advantages of delayed fall dormancy and early spring green-up. However, previous research on the cold resistance of turfgrass has mainly focused on the performance of the spring green-up, with less attention paid to the fall dormancy, which has affected the ornamental and application value of turfgrass. This study first dynamically investigated the leaf colour of each accession during the fall dormancy and the coverage during the spring green-up and evaluated the cold resistance of the accession through membership functions and cluster analysis. Significant differences in the cold resistance were found with the assignment of breeding lines to four categories. The weak correlation (R² = 0.1682) between leaf colour during the fall dormancy and coverage during the spring green-up indicates that using the performance of a single period to represent the cold resistance of accessions is not appropriate. To test whether using the laboratory-based LT50 and stolon regrowth rating analysis can replace the above-improved method, we conducted a related analysis and found that the fit between these two methods is very poor. This phenomenon is attributed to the poor correlation between the laboratory-based parameters and the pot-investigated data. Therefore, this study presents a cold resistance evaluation method for Stenotaphrum that integrates performance in both the fall dormancy and spring green-up periods. This improved evaluation method cannot be simplified by the growth performance of a single period or replaced by using laboratory-based LT50 and stolon regrowth tests. With the help of this improved method, several excellent cold tolerance accessions (ST003, S13, and S12) were identified for temperate regions of East China.