MiR-217 regulates autophagy through OPG/RANKL/RANK in giant cell tumors.

BACKGROUND: Increasing evidence suggests that microRNAs (miRNAs) play a crucial role in cancer development and progression. Our previous study showed remarkably lower levels of miR-217 in GCT cells and tissues, and miR-217 re-expression inhibited the occurrence and development of GCT in vitro; however, the associated mechanisms remain unknown. Thus, this study aimed to explore the mechanisms underlying the proliferation inhibitory effect of miR-217 in GCT cells. METHODS: The proliferative potential of the GCT cells was measured with an MTT assay and BrdU straining. Changes in GCT cell migration and invasion was assessed by a transwell assay. Finally, Western blot and RT-PCR assays were employed to evaluate OPG/RANKL/RANK signaling pathway-related protein expression. RESULTS: The excessive upregulation of miR-217 markedly suppressed GCT cell proliferation and tumorigenesis both in vitro and in vivo. miR-217 overexpression could inhibit the OPG/RANKL/RANK signaling pathway in vitro and in vivo. Furthermore, ALP activity was significantly decreased in GCT cells following miR-217 treatment. Importantly, miR-217 could inhibit autophagy-related protein expression and autophagosome/autolysosome formation in GCT cells and tissues. CONCLUSION: These results suggest that miR-217 upregulation could inhibit the occurrence and development of GCT by blocking autophagy. These findings offer an effective therapeutic target to improve the survival rates of patients with CGT in the future.

Effects of Polyploidization on Morphology, Photosynthetic Parameters and Sucrose Metabolism in Lily.

Polyploidization is widely used in ornamental plant breeding. The polyploids usually produce greater amounts of biomass. However, the alternations to sucrose metabolism that occur in lily during development after polyploidization induced using colchicine are poorly understood. In this study, compared with their allodiploid counterparts, allotetraploid lilies presented a larger total leaf area per plant and slightly delayed flowering time. Moreover, photosynthetic parameter measurements revealed a higher net photosynthetic rate (Pn), stomatal conductance (Gs), transpiration rate (Tr), and maximum Pn for allotetraploids than for allodiploids. Compared with allodiploids, allotetraploids also showed higher nonstructural carbohydrate (NSC) contents during development according to HILIC-CAD results. The expression levels of sucrose metabolism-related genes were higher in allotetraploids than in allodiploids at the same time points. The expression profiles of several target genes in allotetraploids were distinctly different from those in allodiploids. Susy2/3 exhibited opposite expression profiles in allotetraploids and allodiploids, and the expression profiles of SPS3 and Susy2 were significantly correlated with sucrose content change trends in allodiploids and allotetraploids, respectively.

Expression Patterns of Key Genes in the Photoperiod and Vernalization Flowering Pathways in Lilium longiflorum with Different Bulb Sizes.

Lilium longiflorum is a wild Lilium, and its flowering transition requires a long period of cold exposure to meet the demand of vernalization. The responses of different sized bulbs to cold exposure and photoperiod are different, and the floral transition pathways of small and large bulbs are different. In this study, small and large bulbs were placed in cold storage for different weeks and then cultured at a constant ambient temperature of 25 °C under long day (LD) and short day (SD) conditions. Then, the flowering characteristics and expression patterns of key genes related to the vernalization and photoperiod pathways in different groups were calculated and analyzed. The results showed that the floral transition of Lilium longiflorum was influenced by both vernalization and photoperiod, that vernalization and LD conditions can significantly improve the flowering rate of Lilium longiflorum, and that the time from planting to visible flowering buds' appearance was decreased. The flowering time and rate of large bulbs were greatly influenced by cold exposure, and the vernalization pathway acted more actively at the floral transition stage. The floral transition of small bulbs was affected more by the photoperiod pathway. Moreover, it was speculated that cold exposure may promote greater sensitivity of the small bulbs to LD conditions. In addition, the expression of LlVRN1, LlFKF1, LlGI, LlCO5, LlCO7, LlCO16, LlFT1, LlFT3 and LlSOC1 was high during the process of floral transition, and LlCO13, LlCO14 and LlCO15 were highly expressed in the vegetative stage. The expression of LlCO13 and LlCO14 was different under different lighting conditions, and the flowering induction function of LlCO9 and LlFT3 was related to vernalization. Moreover, LlFKF1, LlGI, LlCO5, LlCO16, LlSOC1 and LlFT2 were involved in the entire growth process of plants, while LlCO6, LlCO16 and LlFT1 are involved in the differentiation and formation of small bulblets of plants after the inflorescence stage, and this process is also closely related to LD conditions. This study has great significance for understanding the molecular mechanisms of the vernalization and photoperiod flowering pathways of Lilium longiflorum.

Stomata and ROS changes during Botrytis elliptica infection in diploid and tetraploid Lilium rosthornii Diels.

Polyploid plants often show improved resistance against many diseases, but whether they show increased resistance to grey mould, a devastating disease caused by Botrytis spp. fungi, is seldom reported. Stomata and reactive oxygen species (ROS) play dual roles in defence against grey mould, and it is unclear how their roles change after polyploidization. We addressed these questions in diploid and colchicine-induced Lilium rosthornii after B. elliptica infection. Tetraploids were less susceptible to grey mould, with lower morality rates in naturally infected plants. Before the stomata closed in artificially infected leaves, tetraploids, with larger stomatal apertures, were more easily invaded by the pathogen than diploids. However, the lesion area increased more slowly in tetraploids than in diploids, which may be explained by three causes based on histological and physiological characteristics. First, the pathogen required more time to penetrate the epidermis and closed stomata in tetraploids than in diploids. Second, the pathogen penetrated the reopened stomata more easily than the epidermis, and stomatal density was lower in tetraploids than in diploids. Third, tetraploids showed faster ROS accumulation, a more effective ROS-scavenging system and less malondialdehyde (MDA) accumulation than diploids. Stomatal starch and abnormal guard cell nuclei were present in the infected leaves. This phenomenon may be caused by oxalic acid, a pathogenic factor for many pathogens that promotes stomatal starch degradation and stomatal reopening in Sclerotinia spp., a pathogen closely related to Botrytis spp. This suggestion was primarily confirmed by immersing healthy leaves in oxalic acid solution.

Functional analysis and expression patterns of members of the FLOWERING LOCUS T (FT) gene family in Lilium.

Lilium is an important commercial flowering species, and there are many varieties and more than 100 species of wild Lilium. Lilium × formolongi is usually propagated from seedlings, and the flowering of these plants is driven mainly by the photoperiodic pathway. Most of the other lily plants are propagated via bulblets and need to be vernalized; these plants can be simply divided into pretransplantation types and posttransplantation types according to the time at which the floral transition occurs. We identified three Lilium FLOWERING LOCUS T (LFT) family members in 7 Lilium varieties, and for each gene, the coding sequence of the different varieties was identical. Among these genes, the LFT1 gene of Lilium was most homologous to the AtFT gene, which promotes flowering in Arabidopsis. We analyzed the expression patterns of LFT genes in Lilium × formolongi seedlings and in different Lilium varieties, and the results showed that LFT1 and LFT3 may promote floral induction. Compared with LFT3, LFT1 may have a greater effect on floral induction in Lilium, which is photoperiod sensitive, while LFT3 may play a more important role in the floral transition of lily plants, which have a high requirement for vernalization. LFT2 may be involved in the differentiation of bulblets, which was verified by tissue culture experiments, and LFT1 may have other functions involved in promoting bulblet growth. The functions of LFT genes were verified by the use of transgenic Arabidopsis thaliana plants, which showed that both the LFT1 and LFT3 genes can promote early flowering in Arabidopsis. Compared with LFT3, LFT1 promoted flowering more obviously, and thus, this gene could be an important promoter of floral induction in Lilium.

Effects of ploidy level on the cellular, photochemical and photosynthetic characteristics in Lilium FO hybrids.

Polyploidy plays an important role in plant breeding, inducing a wide range of effects on photosynthetic performance, cellular and photochemical aspects of photosynthesis. Although the impacts of polyploidization on photosynthesis have been examined in several genera, comparatively little is known about the photosynthetic responses to polyploidy in Lilium. In this study, diploid and colchicine-induced tetraploid Lilium FO hybrids were used to study the effects of polyploidization on cellular, photochemical and photosynthetic characteristics in Lilium. Polyploidization caused a significant change in leaf anatomical structure, chloroplast ultrastructure and photosynthetic pigment contents. Thicker epidermal and spongy tissue, more and thicker thylakoid lamellae and higher chlorophyll and carotenoid contents were observed in the tetraploid than in the diploid. More regularly and tightly arranged thylakoids were closely associated with more efficient light-harvesting machinery. And the tetraploid plants showed a higher net photosynthetic rate (Pn) and maximum net photosynthetic rate (Pmax) than the diploid plants, under both natural conditions and gradients of light intensity/CO2 concentration. In addition, the tetraploid had a significantly higher light saturation point (LSP), but a lower light compensation point (LCP) than the diploid, indicating that tetraploid plants acquired light energy more efficiently. These results suggested that cellular and photochemical alterations caused by polyploidization improve the capacity for light absorption and conversion and further enhanced the photosynthetic performance in Lilium FO hybrids.