The AT-hook Motif Nuclear Localized (AHL) protein family is known for its pivotal roles in plant growth regulation, developmental patterning, and stress signal transduction. Although the gene family has been studied in various plant species, the genomic characteristics, evolutionary mechanism, and expression profiles of the AHL family in cassava (Manihot esculenta) remain unexplored. In this study, we comprehensively investigated the evolutionary features and biological response of the MeAHL gene family through genome-wide identification, phylogenetic analysis, structural characterization, and large-scale transcriptomes based on the cassava SC205 reference genome. We identified 41 putative members through genome-wide identification. Physicochemical property analysis showed that all 41 MeAHLs were hydrophilic proteins, and 40 of them were unstable proteins, with the number of amino acids generally ranging from 188 to 446 aa. Phylogenetic analysis indicated that the MeAHL family members could be divided into two clades, Clade A and Clade B. Two MeAHL gene clusters were located in the distal telomeric regions of chromosomes Chr01 and Chr02, respectively. Replication type analysis revealed that the evolution of MeAHLs was mainly driven by whole-genome duplication (WGD) and dispersed duplication (DSD), with the Ka/Ks values <1. Evolutionary mechanism analysis indicated that whole-genome duplication (WGD) primarily drove the MeAHL gene family expansion. Gene structure analysis showed that MeAHL genes were mainly composed of 1‒10 exons. Analysis of conserved domains and motifs showed that all MeAHLs had the PPC/DUF296 domain and AT-hook motif. Members of Clade A generally contained one Type-I AT-hook motif. Among members of Clade B, except for SC20508G13380 and SC20509G13950, which contained one Type-II AT-hook motif, most members contained two AT-hooks (Type-I and Type-II). Cis-acting element analysis via PlantCARE showed that the cis-acting elements related to light response were the most abundant in MeAHLs, such as Box 4, G-box, and they also contained elements responsive to hormones, biotic stresses, and abiotic stresses, such as ABRE, MBS, W-box, and TC-rich repeats. Tissue-specific expression profiling revealed distinct expression patterns between two clades of MeAHL across 11 different tissues. Stress transcriptome analysis demonstrated significant responses of specific MeAHLs to drought (ABA/PEG treatments), cassava bacterial blight (Xanthomonas axonopodis pv. manihotis), and mite infestation, showing clade-specific regulatory patterns. Protein-protein interaction (PPI) network prediction suggested some MeAHLs formed functional modules with bHLH, NAC, ARF, and NB-LRR proteins involved in plant development and stress responses. This study would provide the systematic characterization of AHL family evolution and functional diversification in cassava, offering theoretical foundations for molecular breeding applications.

In this study, Cymbopogon winterianus was used as the experimental material, and the DNA sequence of C. winterianus was sequenced using the Illumina NovaSeq 6000 sequencing platform. The sequencing data were assembled with GetOrganelle v1.7.7.0 software to construct the chloroplast genome. Referring to the known chloroplast genome of C. flexuosus, the chloroplast genome of C. winterianus was annotated, and the genomic characteristics were analyzed and a phylogenetic tree was constructed. The chloroplast genome of C. winterianus was 139 823 bp in length, with a typical circular quadripartite structure. The GC content was 38.45%, and the AT content was 61.55%. It included a large single-copy region (LSC) with a length of 82 214 bp, a pair of inverted repeat regions (IR) with a length of 21 368 bp, and a small single-copy region (SSC) of 14 873 bp. A total of 130 genes were annotated in the chloroplast genome of C. winterianus (including 85 mRNA genes, 37 tRNA genes, and 8 rRNA genes). In addition, among the annotated genes, there were 16 double-copy genes, accounting for 12.31%, including 7 tRNA genes, 4 self-replication genes, 4 rRNA genes, 2 protein genes with unknown functions, and 1 NADH dehydrogenase subunit gene. A total of 144 SSR loci were detected in the chloroplast genome of C. winterianus, with mononucleotide repeats being absolutely dominant, mainly A/T. After comparing the boundaries of the inverted repeat sequences of four Cymbopogon species, it was found that C. flexuosus, C. pospischilii, and C. winterianus exhibited extremely high homology in gene structure and species. Among them, the ndhH gene was located in the small single-copy region (SSC), and the ndhF gene was located in the boundary region between the SSC and IRb. However, C. winterianus had an additional rps3 gene in the LSC region compared with C. flexuosus and C. pospischilii. Phylogenetic tree analysis showed that C. winterianus had the closest genetic relationship with C. pospischilii and C. citratus (MK593547.1). This study completed the assembly and annotation of the complete chloroplast genome of C. winterianus, analyzed the characteristics of the chloroplast genome of C. winterianus, and preliminarily explored the phylogenetic position of C. winterianus within the genus Cymbopogon. It would lay a good foundation for the phylogenetic, genetic diversity, and genomic studies of Cymbopogon plants, as well as for the discovery and utilization of important functional genes.