In previous studies, the PD-NINJA complex was found to control the initial shape of the leaf and inhibit CYCD3, thereby controlling leaf flatness in angiosperms [18]. Boucheron et al. [19] transferred CYCD3 to tobacco, which resulted in increased leaf growth rates and changes in the structure of the shoot apical meristem. Dewitte et al. [15] found that Arabidopsis leaves transfected with CYCD3 curled toward the medial axis. The A. thaliana AtCYCD3;1 gene was also found to wrinkle and curl leaves in transgenic tobacco [20]. CYCD3 overexpression caused delayed morphological differences in SAM and leaf senescence [14]. Additionally, CYCD3 was also found to be expressed at its highest levels during the flower bud stage in Rhododendron [10]. Taihangia unisexual male flowers downregulated the expression of CYCA1 and CYCD3, which resulted in the disturbance of the cell cycle in gynoecium primordia and subsequently determined the arrest of pistil development [21]. Currently, CYCD3 is the main focus of herbaceous plant investigations, which are mostly limited to overexpression. However, CYCD3 against woody plants is rarely reported, and the expression of inhibition after gene silencing is hardly described. The cyclin aa of woody plants varies widely, and existing studies have not been able to clarify the function of woody plant CYCD3. The transcription factor AINTEGUMENTA (ANT) and the D-type cyclin CYCD3;1 are expressed in the vascular cambium of Arabidopsis roots, respond to cytokinins and are both required for proper root secondary thickening, AIL1 can directly interact with the promoters of CYCD3:2 and CYCD6:1, AIL1 presumably participates in the regulation of cell division in meristematic tissues at the shoot apex [22–23]. However, there are few reports on the regulation of flowering by CYCD3 in woody plants, and the expression of inhibition after gene silencing is hardly recorded, and the expression of inhibition after gene silencing is hardly described. The cyclin aa of woody plants varies widely, and existing studies have not been able to clarify the function of woody plant CYCD3.
Typical animal or plant cyclins contain a conserved region called the cyclin core, which is ~ 250 aa and consists of 2 domains: Cyclin_N and Cyclin_C. Cyclins have certain structural differences, but they all have highly conserved cyclin cassette sequences. Cyclin_N is required for the binding and activation of cyclin-dependent kinase (CDK), while Cyclin_C is less conserved and may not be present in some cyclins [24–27]. Arabidopsis C-, D-, H-, T-, L-, and P-type cyclins have a Cyclin_N domain, but no Cyclin_C domain; thus, the Cyclin_C domain may not be critical for its function [28]. Intron retention is the most common alternative splicing form in plants [29]. In JnCYCD3;1, retention of the first intron leads to its coding frame in advance. However, the termination or loss of the Cyclin_C domain, whose structural differences may lead to changes in traits, requires further investigation. The expression pattern of CYCD3 in different tissues of 2 varieties of J. curcas was essentially the same, and both were the highest expressed in young leaves, followed by inflorescence buds during the differentiation process. Transcript abundance of CYCD3;1 increased more than 5-fold during the early stages of L1 flower development (the first 10 inflorescences of each tomato plant retaining the fruit of the second inflorescence) and is possibly involved in cell cycle regulation in response to mitosis signals [30]. Brassica napus BnCYCD3-1-like-2-1 and BnCYCD3-1-like-2-2 are different splicing bodies of the same gene and have the highest expression levels in leaves. Additionally, the variable cleavage of CYCD3:1 could possibly help rapeseed cope with environmental stress by coordinating the transcript levels of different splices [31].
Subcellular functional localization indicated that JnCYCD3;1 functions in the nucleus. BIFC experiments indicated that JnMYC2 interacts with JnCYCD3;1. The transgenic seedlings demonstrated that JnCYCD3;1 can considerably advance the reproductive cycle, which may promote flower formation and transformation. However, the cause of this phenomenon, as well as research on transcription, metabolism, and late phenotypes of wild-type plants, requires further investigation. Because tobacco is a hermaphroditic plant, if JnCYCD3;1 leads to the proliferation of female flower primordia, it is necessary to construct a related overexpression/RNAi vector and introduce it to medicinal J. curcas/nigroviensrugosus in future observations.