Background: Plant genomes contain a large number of HAK/KUP/KT transporters, which play important roles in potassium uptake and translocation, osmotic potential regulation, salt tolerance, root morphogenesis and plant development. Potassium deficiency in the soil of a sugarcane planting area is serious. However, the HAK/KUP/KT gene family remains to be characterized in sugarcane (Saccharum). Results: In this study, 30 HAK/KUP/KT genes were identified in Saccharum spontaneum. Phylogenetics, duplication events, gene structures and expression patterns were analyzed. Phylogenetic analysis of the HAK/KUP/KT genes from 15 representative plants showed that this gene family is divided into four groups (clades I-IV). Both ancient whole-genome duplication (WGD) and recent gene duplication contributed to the expansion of the HAK/KUP/KT gene family. Nonsynonymous to synonymous substitution ratio (Ka/Ks) analysis showed that purifying selection was the main force driving the evolution of HAK/KUP/KT genes. The divergence time of the HAK/KUP/KT gene family was estimated to range from 134.8 to 233.7 Mya based on Ks analysis, suggesting that it is an ancient gene family in plants. Gene structure analysis showed that the HAK/KUP/KT genes were accompanied by intron gain/loss in the process of evolution. RNA-seq data analysis demonstrated that the HAK/KUP/KT genes from clades II and III were mainly constitutively expressed in various tissues, while most genes from clades I and IV had no or very low expression in the tested tissues at different developmental stages. The expression of SsHAK1 and SsHAK21 was upregulated in response to low-K⁺ stress. Yeast functional complementation analysis revealed that SsHAK1 and SsHAK21 could rescue K⁺ uptake in a yeast mutant. Conclusions: This study provided insights into the evolutionary history of HAK/KUP/KT genes. HAK7/9/18 were mainly expressed in the upper photosynthetic zone and mature zone of the stem. HAK7/9/18/25 were regulated by sunlight. SsHAK1 and SsHAK21 played important roles in mediating potassium acquisition under limited K⁺ supply. Our results provide valuable information and key candidate genes for further studies on the function of HAK/KUP/KT genes in Saccharum.

Background: B-box (BBX) zinc-finger transcription factors play crucial roles in plant growth, development, and abiotic stress responses. Nevertheless, little information is available on sugarcane (Saccharum spp.) BBX genes and their expression profiles. Results: In the present study, we characterized 25 SsBBX genes in the Saccharum spontaneum genome database. The phylogenetic relationships, gene structures, and expression patterns of these genes during plant growth and under low-nitrogen conditions were systematically analyzed. The SsBBXs were divided into five groups based on phylogenetic analysis. The evolutionary analysis further revealed that whole-genome duplications or segmental duplications were the main driving force for the expansion of the SsBBX gene family. The expression data suggested that many BBX genes (e.g., SsBBX1 and SsBBX13) may be helpful in both plant growth and low-nitrogen stress tolerance. Conclusions: The results of this study offer new evolutionary insight into the BBX family members in how sugarcane grows and responds to stress, which will facilitate their utilization in cultivated sugarcane breeding.

Plant genome sequences are presently deciphered at a staggering speed, due to the rapid advancement of high-throughput sequencing technologies. However, functional genomics significantly lag behind due to technical obstacles related to functional redundancy and mutant lethality. Artificial microRNA (amiRNA) technology is a specific, reversible, and multiplex gene silencing tool that has been frequently used in generating constitutive or conditional mutants for gene functional interrogation. The routine approach to construct amiRNA precursors involves multiple polymerase chain reactions (PCRs) that can increase both time and labor expenses, as well as the chance to introduce sequence errors. Here, we report a simplified method to clone and express amiRNAs in Arabidopsis and rice based on the engineered Arabidopsis miR319a or rice miR528 precursor, which harbor restriction sites to facilitate one-step cloning of a single PCR product. Stem-loop reverse-transcriptase quantitative PCR (RT-qPCR) and functional assays validated that amiRNAs can be accurately processed from these modified precursors and work efficiently in plant protoplasts. In addition, Arabidopsis transgenic plants overexpressing the modified miR319a precursor or its derived amiRNA could exhibit strong gene silencing phenotypes, as expected. The simplified amiRNA cloning strategy will be broadly useful for functional genomic studies in Arabidopsis and rice, and maybe other dicotyledon and monocotyledon species as well.

Plasma membrane intrinsic proteins (PIPs), as members of the aquaporin (AQPs) family, can transport not only water but also urea, CO₂, H₂O₂, metal ions, and trace elements. They are crucial for maintaining water balance, substance transport, and responding to various stresses. This article delves into the structure, function, response mechanism, molecular mechanism, and regulatory mechanism of PIPs as a result of biological and abiotic stresses. It also summarizes current research trends surrounding PIPs and highlights potential research directions for further exploration. The aim is to assist researchers in related fields in gaining a more comprehensive understanding and precise insight into the advancements in PIP research.

Background: The identification and functional analysis of genes that improve tolerance to low potassium stress in S. spontaneum is crucial for breeding sugarcane cultivars with efficient potassium utilization. Calcineurin B-like (CBL) protein is a calcium sensor that interacts with specific CBL-interacting protein kinases (CIPKs) upon plants’ exposure to various abiotic stresses. Results: In this study, nine CBL genes were identified from S. spontaneum. Phylogenetic analysis of 113 CBLs from 13 representative plants showed gene expansion and strong purifying selection in the CBL family. Analysis of CBL expression patterns revealed that SsCBL01 was the most commonly expressed gene in various tissues at different developmental stages. Expression analysis of SsCBLs under low K⁺ stress indicated that potassium deficiency moderately altered the transcription of SsCBLs. Subcellular localization showed that SsCBL01 is a plasma membrane protein and heterologous expression in yeast suggested that, while SsCBL01 alone could not absorb K⁺, it positively regulated K⁺ absorption mediated by the potassium transporter SsHAK1. Conclusions: This study provided insights into the evolution of the CBL gene family and preliminarily demonstrated that the plasma membrane protein SsCBL01 was involved in the response to low K⁺ stress in S. spontaneum.

Background: Growth regulating factors (GRFs) are transcription factors that regulate diverse biological and physiological processes in plants, including growth, development, and abiotic stress. Although GRF family genes have been studied in a variety of plant species, knowledge about the identification and expression patterns of GRFs in sugarcane (Saccharum spp.) is still lacking. Results: In the present study, a comprehensive analysis was conducted in the genome of wild sugarcane (Saccharum spontaneum) and 10 SsGRF genes were identified and characterized. The phylogenetic relationship, gene structure, and expression profiling of these genes were analyzed entirely under both regular growth and low-nitrogen stress conditions. Phylogenetic analysis suggested that the 10 SsGRF members were categorized into six clusters. Gene structure analysis indicated that the SsGRF members in the same group were greatly conserved. Expression profiling demonstrated that most SsGRF genes were extremely expressed in immature tissues, implying their critical roles in sugarcane growth and development. Expression analysis based on transcriptome data and real-time quantitative PCR verification revealed that GRF1 and GRF3 were distinctly differentially expressed in response to low-nitrogen stress, which meant that they were additional participated in sugarcane stress tolerance. Conclusion: Our study provides a scientific basis for the potential functional prediction of SsGRF and will be further scrutinized by examining their regulatory network in sugarcane development and abiotic stress response, and ultimately facilitating their application in cultivated sugarcane breeding.