London plane tree is widely grown as a landscaping and street tree, but the release of its trichomes creates a serious air-borne pollution problem. Identifying the key genes that regulate the development of trichomes is, therefore, an important tool for the molecular breeding of Platanus acerifolia. In this study, a sequence homologous with the Arabidopsis Class II TCP subfamily was identified from London plane, and named PaTCP4. The expression of PaTCP4 was detected in various organs of London plane trees, significantly in the trichomes. Overexpression of PaTCP4 in Arabidopsis reduced the trichome density on the first pair of true leaves, and atypical 5-branched trichomes were also detected on those leaves. The expression of endogenous AtCPC and AtTCL2 was significantly increased in PaTCP4 transgenic lines, and was associated with a decrease in the expression of endogenous AtGL2. Furthermore, the expression of endogenous AtGL3 was significantly increased. In addition, the protein product of PaTCP4 was shown to directly activate AtCPC, AtTCL2, AtGL3, AtGIS, PaGIS, and PaGL3 in yeast one-hybrid assays and in the dual-luciferase reporter system. Taken together, these results identify a role for PaTCP4 in trichome initiation and branching in Arabidopsis. Thus, PaTCP4 represents a strong candidate gene for regulating the development of trichomes in London plane trees.

Flowering and senescence are essential developmental stages of green plants, which are governed by complex molecular regulatory networks. However, the connection between flowering regulation and senescence regulation in London plane tree (Platanus acerifolia) remains unknown. In this study, we identified a gene PaNAC089 from London plane tree, which encodes a membrane-tethered transcription factor (MTTF) belonging to the NAC (NAM, ATAF1/2, CUC2) transcription factor family. We investigated the functions of PaNAC089 in the regulation of flowering and senescence through the analysis of expression profiles and transgenic phenotypes. Heterologous overexpression of ΔPaNAC089 delayed flowering and inhibited chlorophyll breakdown to produce dark green rosette leaves in Arabidopsis. In addition, the trichome density of rosette leaves was decreased in transgenic lines. In ΔPaNAC089 overexpression plants, a series of functional genes with inhibited expression were identified by quantitative real-time polymerase chain reaction (qRT-PCR), including genes that regulate flowering, chlorophyll decomposition, and trichome initiation. Furthermore, ΔPaNAC089 directly binds to the promoter of CONSTANS (CO) and NON-YELLOWING2 (NYE2) in the yeast one-hybrid assay. Consistent with this, luciferase (LUC) transient expression assays also showed that ΔPaNAC089 could inhibit the activity of NYE2. To summarise, our data suggests that PaNAC089 is an MTTF that modulates flowering, chlorophyll breakdown and trichome initiation.

Age-dependent flowering is a prominent feature of perennial orchids, yet its physiological and small RNA basis remains poorly understood in Cymbidium goeringii. Here, floral bud formation, carbohydrate contents, and endogenous hormones were examined across successive shoot age stages, and four representative stages were selected for small RNA sequencing. Floral bud formation showed a clear age-dependent rise-and-fall pattern, accompanied by distinct changes in carbohydrate and hormone levels. Small RNA sequencing identified 810 miRNAs, including 356 known and 454 putative novel miRNAs, of which 311 were differentially expressed. Temporal expression analysis grouped 210 DEMIs into four significant profiles, all showing reduced expression in the oldest shoots. Functional analysis of predicted targets highlighted starch and sucrose metabolism and plant hormone signal transduction, and further pathway analysis identified multiple predicted miRNA–target associations related to carbohydrate metabolism and hormone signaling. qRT-PCR analysis generally supported the sequencing-based expression patterns. These findings reveal coordinated age-related changes in flowering competence, physiological status, and miRNA expression, and identify candidate miRNAs and functional pathways potentially associated with age-dependent flowering in C. goeringii.