Early maturity is an important target trait in cotton genetic improvement in China, and flowering time is a key indicator of earliness. H3K9 histone lysine methyltransferases (SUVs) are involved in plant development and flowering regulation in model plants such as Arabidopsis thaliana and rice. This study aimed to investigate the role of the SUV gene family in flowering regulation in upland cotton and to identify elite allelic variations of key members. The findings provide valuable genetic resources for breeding early-maturing cotton cultivars.

Protein sequences of the SUV family from Arabidopsis were used as reference sequences. HMMER, MEME, and TBtools were employed to identify and analyze SUV family members in upland cotton at the whole-genome level. Candidate genes involved in flowering regulation were screened through integrated RNA-seq and RT-qPCR analyses. Virus-induced gene silencing (VIGS) was used to validate the functions of candidate genes in flowering regulation. Resequencing data from 419 upland cotton accessions were analyzed to identify SNPs within GhSUV1, which were further validated by Sanger sequencing. Allele frequencies and genetic differentiation were examined across different cotton ecological regions.

A total of 200 SUV homologous proteins were identified in nine higher plant species, including rice. Phylogenetic analysis divided them into SUVH and SUVR clades, indicating a relatively conserved evolutionary pattern. In upland cotton, 28 GhSUV genes were identified. Their expansion was mainly driven by polyploidization events. Most collinear gene pairs showed Ka/Ks<1, suggesting strong purifying selection. Gene structure and motif analyses revealed high similarity among members within the same clade. Promoter analysis showed that GhSUV genes are enriched in light-responsive and hormone-responsive cis-elements, indicating their involvement in multiple developmental regulatory pathways. RNA-seq and RT-qPCR analyses showed that GhSUV1 and GhSUV13 were expressed at higher levels in early-maturing materials. They were also highly expressed in leaves and certain floral organs. Functional validation using VIGS demonstrated that silencing GhSUV1 delayed squaring and flowering by 5.6 and 6.0 days, respectively. Silencing GhSUV13 delayed these stages by 5.8 and 6.8 days compared with the control. Moreover, suppression of GhSUV1 and GhSUV13 significantly reduced the expression of flowering-related genes GhSOC1, GhAP1, GhFT, and GhCAL, while significantly increasing GhSVP expression. These results indicate that both genes participate in flowering regulation. Sequence analysis identified a SNP in the exon region of GhSUV1 that was significantly associated with flowering time. The early-flowering GG allele increased in frequency with latitude. Genetic differentiation index and nucleotide diversity analyses revealed significant differences in the GhSUV1 genomic region between early- and late-maturing cultivars, suggesting that this locus has undergone breeding selection.

GhSUV1 and GhSUV13 are involved in flowering regulation in upland cotton. The elite allele of GhSUV1 is closely associated with early flowering and has been subject to breeding selection. These findings provide important genetic resources for the improvement of early-maturing cotton varieties.

The fruit branch angle is a key determinant of cotton plant architecture. Previous studies revealed differential expression of GhPDF1 in materials with extreme fruit branch angles. This study aimed to functionally characterize GhPDF1 and systematically dissect its superior allelic variations, thereby providing genetic resources and a theoretical basis for improving plant architecture and breeding machine-harvestable cotton varieties.

The sequences of PDF1 homologs from 25 species, including upland cotton (Gossypium hirsutum), were obtained through homology alignment for phylogenetic analysis. A virus-induced gene silencing (VIGS) vector targeting GhPDF1 was constructed using double digestion and transformed into upland cotton to investigate its role in regulating the fruit branch angle. Single nucleotide polymorphisms (SNPs) within GhPDF1 were identified using resequencing data from 418 upland cotton accessions, with key SNPs validated by Sanger sequencing. The distribution of superior allelic variations of GhPDF1 was analyzed across Chinese cotton varieties from different breeding eras.

A phylogenetic tree of the PDF1 gene from Gossypium hirsutum and its homologous genes from different species revealed that it is most closely related to Malvaceae, while being distantly related to Poaceae. Expression analysis showed that GhPDF1 was lowly expressed in large‑angle materials but highly expressed in small‑angle materials. Tissue‑specific expression profiling revealed high transcript levels in pistils, petals, and stems. VIGS‑mediated silencing of GhPDF1 significantly reduced the cell number at the fruit branch junction by approximately 170 and increased the fruit branch angle by 8.2° compared with empty‑vector controls, demonstrating its crucial role in regulating fruit branch angle. Phenotypic comparison between two allelic variants, GhPDF1^GG and GhPDF1^CC, indicated a significantly smaller branch angle in GhPDF1GG carriers. Moreover, the frequency of this superior allele (GhPDF1^GG) increased from 92% to 98% over successive breeding periods.

Silencing GhPDF1 in upland cotton reduces cell proliferation at the branch junction, leading to a significant increase in the fruit branch angle. The superior allelic variant GhPDF1^GG has been progressively enriched during modern cotton breeding in China.