Plant mutant, one of the basic phenomena in plant evolution, is highly valuable for crop breeding. With the rapid development of modern sequencing technology, we can provide support for the development of molecular breeding, and speed up the breeding process of new varieties. The fan-shaped inflorescences (fruits) in pineapple had both high ornamental value and higher fruit yields (Table 1). Therefore, clarifying the molecular mechanism of the formation of the fan-shaped inflorescence is highly valuable in plant breeding in pineapples.
No differences in the gene and signal transduction were found between the fan- and capitulum-shaped inflorescences using NGS. The comparisons across the three tissues and two pineapples provided us with 5370 DEGs; more DEGs were recognized between FIs vs. CIs and FIb vs. CIb than those between FIa vs. CIa. In addition, 5769 DEGs were identified between different tissues within each pineapple, indicating that the regulatory pathway existed in the formation of the fan-shaped inflorescence in pineapple.
Previous studies have identified genes that may be involved in the inflorescence development in plants, including some genes related to auxin and auxin processes [39, 40]; auxin has recently been reported to be associated with inflorescence development [41]. Across the three tissues and two pineapples, 206 IDGs were identified, including LAX, LOG, CKX, PAP, FLOWERING LOCUS T, LOB, and Aux / IAA, which may participate in the formation of the fan-shaped inflorescence in pineapple. Moreover, there exist 21 overlapping genes of them in all 3 comparisons within each tissue between the two pineapples; they were also crucial for regulating development of fan inflorescence in pineapple.
CLAVATA (CLV), which is related to the inflorescence development by regulating the transformation of SAM into IM, played an essential role in the transition from SAM into IM [42]. It has been proposed that LOG was involved in various induction of inflorescence formation, some transcriptional regulators and cytokinin regulation through the complex interactions [29, 30]. TERMINAL FLOWER and FLOWERING LOCUS T play an important role in the inflorescence formation [6, 43]. In our study, five differently expressed genes related to TERMINAL FLOWER (1 gene) and FLOWERING LOCUS T (4 genes) were identified within each tissue between the two pineapples (Additional file 9a). Cytokinin, auxin and ethylene are necessary for inflorescence formation [44, 45]. For example, PIN in Arabidopsis was found to control meristem activity, and was regulated in turn by three redundant PLETHORA (PLT) proteins [46].
As a plant growth hormone, auxin is considered as a necessary condition for plant growth, development [47, 48], flower bud differentiation [45], as well as transition from SAM to IM [42]. Auxin was accumulated during inflorescence development in Arabidopsis, and the endogenous auxin concentration was higher in IM tissue in developing inflorescence [37]. Auxin was also involved in the induction of flower bud differentiation in Juglans sigillata [49]. It was previously found that the free and bound auxin levels are highly regulated by inflorescence formation [45]. In pineapples, we found that the majority of genes were down-regulated across the three different tissues between the two pineapples (Fig. 8a). Therefore, we speculated that the formation of fan-shaped inflorescence may be related to the epigenetic modification (i.e., DNA methylation). Research of inflorescence development identification and characterization indicated that the auxin-related genes and proteins were related to inflorescence development in auxin-induced acquisition [37]. Thus 206 IDGs, especially the 21 overlapping IDGs, might be involved in the formation of the fan-shaped inflorescence in pineapple.
TFs have great potential value in plant growth and development. Previous studies on the inflorescence development showed that the complex transcription regulation networks played an integral part in tissue differentiation and hormone mediated signaling [47]. In this study, we obtained 271 TFs showing significantly different expression across the three different tissues between the two pineapples; 45 of them were simultaneously differentially expressed in across all three different tissue comparisons between the two pineapples. Of these 444 TFs, MYB, AP2/ERF, and MADS-box showed highly representative expression.
Cytokinin plays an integral part during regulating the inflorescence structure of different inflorescence types by regulating the activity of meristem [50, 51]. Cytokinin promoted inflorescence meristem development and affected inflorescence structure by promoting Arabidopsis meristem gene WUSCHEL (WUS) and by inhibiting meristem inhibitors CLAVATA1 (CLV1) and CLV3 [52]. The expression of WUS can be directly induced by cytokinin. WUS enhanced cytokinin signal transduction and formed a positive feedback loop [53, 54]. At the same time, the transcription of CLV3 gene can also be promoted by WUS expression. WUS is not only an important target gene of CLV negative control signal, but also a positive regulatory gene. WUS and CLV constitute a regulatory feedback loop [53, 55]. In our research, the up-regulated genes of WUS had 3, 4 and 4 genes, respectively, within each tissue comparison between the two pineapples (Additional file 7). Compared with the capitulum-shaped inflorescences, the proportion of down-regulated DEGs at FIs tissue was higher than that FIb and FIa tissues, indicating that they had a regulatory role in the formation of the fan-shaped inflorescence in pineapple.
AP1, a member of AP2/ERF domain family, shows a significant effect on inflorescence configuration regulation [56]. AP1 plays the role of transcriptional inhibition to inhibit flower development at the beginning, and can promote flower development when combined with SEP3 protein [57, 58]. LFY is a flower tissue differentiation gene, which is mainly expressed in leaf primordium in vegetative growth stage, and rapidly increased to the highest level in the flower induction stage [57]. LFY can regulate the transformation from the vegetative growth to the reproductive growth of plants, and can also regulate the special morphology of flower organs [57]. LFY and AP1 can positively promote each other's expression. LFY can bind to the promoter of AP1 gene and promote the expression of corresponding gene, while TFL1 gene can inhibit the expression of LFY and AP1 gene [59]. These interactions among the three genes play a unique role in the regulation of inflorescence configuration and inflorescence evolution. In addition, AP1 protein and its homologues CAL and FUL in Arabidopsis can inhibit TFL1 gene expression [59]. In this study, there were 6 AP2/ERF genes differentially expressed across three tissues between the two pineapples (Additional file 8a); among them, 2 genes (Aco008597,Aco013345) were expressed differently among 3 tissues within FI, but not within CI (Additional file 8a). Therefore, these AP2/ERF genes may be related to the formation of the fan-shaped inflorescence in pineapple.
The transcription regulators, e.g., SVP, SOC1 and AGL24 of MADS domain have dual effects [60]: preventing the reverse of floral meristem to inflorescence structure [61], and maintaining a moderate level of expression at a specific stage of development. On the one hand, they have redundant functions with SEP4 and can interact with AP1 to inhibit the expression of TFL1; SEP4 directly suppresses TFL1 expression without SOC1, AGL24 and SVP [60]. On the other hand, SVP, SOC1 and AGL24 also directly inhibit the expression of SEPALLATA3 (SEP3), a flower organ characteristic gene, to prevent flower meristem from early differentiation, so as to ensure the timely formation and development of flower organs in Arabidopsis [62]. Between our two studied pineapples, 8 MADS genes were differentially expressed in the three tissue comparisons; 2 genes (Aco006017,Aco018015) were expressed differently in the comparison of tissues in FI, but not in the comparison of tissues in CI (Additional file 8a).
Members of MYB are involved in growth, development and regulation of phenylpropane secondary metabolic pathways in plants [63]. They control the cell morphology and participate in the development and regulation of floral organs [64]. MYB38 and MYB84 are important regulatory factors that affect the formation of axillary meristem in Arabidopsis [65]. AtMYB38 affects the formation of auxiliary lateral branches during inflorescence development [65]. In our study, up-regulated MYB in the flower stem apex, the base of the inflorescence, and the inflorescence axis were 22, 39 and 4, respectively (Fig. 6), indicating that there was a complex regulation in the formation of the fan-shaped inflorescence in pineapple. In addition, most of MYB genes were up-regulated in the fan-shaped inflorescence tissues (Fig. 6), indicating that these family members played an important role in the formation of the fan-shaped inflorescence.
Plant inflorescence development is a complex process. In the morphogenesis of higher plants, inflorescence development is the basic step to determine the final yield and reproductive characteristics. Auxin is essential for flower development [37]. In this study, many auxin related genes and TFs were expressed differently in each comparison. Therefore, the auxin related genes and TFs regulate the formation of the fan-shaped inflorescence in pineapple. In addition to IDGs and TFs, 80 genes were involved in the GO biological process and reproduction of auxin signal in the three tissue comparisons between the two pineapples, and they were perhaps related to the formation of the fan-shaped inflorescence.