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Carthamus tinctorius L., commonly known as safflower, is an important oilseed crop containing oil bodies. Oil bodies are intracellular organelles in plant cells for storing triacylglycerols (TAGs) and sterol esters. Oleosins are the most important surface proteins of the oil bodies. We predicted and retrieved the sequences of eight putative C. tinctorius oleosin (Ctoleosin) genes from the genome database of safflower. The bioinformatics analyses revealed the size of their open reading frames ranging from 414 to 675 bp, encoding 137 to 224 aa polypeptides with predicted molecular weights of 14.812 to 22.155 kDa, all containing the typical “proline knot” motif. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) determined the spatiotemporal expression pattern of Ctoleosin genes, which gradually increased and peaked during flowering and seed ripening, and decreased thereafter. To validate their role in plant development, we transformed and overexpressed these eight putative Ctoleosin genes in Arabidopsis. Overexpressing Ctoleosins did not affect leaf size, although silique length was altered. Arabidopsis transformed with Ctoleosin3, 4, and 5 grew longer siliques than did the wild-type plants, without altering seed quantity. The 100-grain weight of the transgenic Arabidopsis seeds was slightly more than that of the wild-type seeds. The seed germination rates of the plants overexpressing Ctoleosin4 and 6 were slightly lower as compared with that of the wild-type Arabidopsis, whereas that in the other transgenic lines were higher than that in the wild-type plants. The overexpression of Ctoleosin genes elevated the oil content in the seeds of transgenic Arabidopsis. Our findings not only provide an approach for increasing the oil content, but also for elucidating the intricate mechanisms of oil body synthesis.

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BackgroundEpidermal growth factor (EGF) can promote cell proliferation as well as migration, which is feasible in tissue wound healing. Oil bodies have been exploited as an important platform to produce exogenous proteins. The exogenous proteins were expressed in oil bodies from plant seeds. The process can reduce purification steps, thereby significantly reducing the purification cost. Mostly, the diameter of oil body particle ranges between 1.0 and 1.5 µm in the safflower seeds, however, it reduces to 700–1000 nm in the transgenic safflower seeds. The significant reduction of particle size in transgenic seeds is extremely beneficial to skin absorption.ResultsThe diameter of oil body in the transgenic safflower seeds was recorded in the range of 700–1000 nm. The smaller particle size improved their skin absorption. The expression level of oleosin-hEGF-hEGF in T3 transgenic seeds was highest at 69.32 mg/g of seeds. The oil body expressing oleosin-hEGF-hEGF had significant proliferative activity on NIH/3T3 cells and improved skin regeneration thereby accelerating wound healing in rats. The wound coverage rate exceeded 98% after treatment for 14 days with oil body expressing oleosin-hEGF-hEGF, while the saline without EGF group and wild type oil body group both showed less than 80%. The neonatal fibroblast and collagen were found to be increased in the safflower oil body expressing oleosin-hEGF-hEGF treatment group. TGF-β1, bFGF and VEGF were noted as important growth factors in the repair of cutaneous wounds. Their expression level increased after 4 and 7 day treatment, but decreased after 14 days. Therefore, it can promote skin regeneration to accelerate wounds healing.ConclusionsThe expression of oleosin-hEGF-hEGF in T3 transgenic seeds was 80.43 ng/μL oil body. It had significant proliferative activity on NIH/3T3 cells and improved skin regeneration to accelerate wound healing in rats. The expression process of TGF-β1, bFGF and VEGF increased at first and then gradually declined.Electronic supplementary materialThe online version of this article (10.1186/s12951-018-0387-5) contains supplementary material, which is available to authorized users.

We set out to assess the NIH/3T3 cell proliferation activity of Arabidopsis oil body-expressed recombinant oleosin–hEGF–hEGF protein. Normally, human epidermal growth factor (hEGF) is purified through complex process, however, oleosin fusion technology provides an inexpensive and scalable platform for its purification. Under a phaseolin promoter, we concatenated oleosin gene to double hEGF (hEGF–hEGF) with plant-preferred codons in the expression vectors and the construct was transformed into Arabidopsis thaliana (Arabidopsis). The transgenic Arabidopsis was validated by RT–PCR and the content of recombinant protein oleosin–hEGF–hEGF was quantified by western blot. Subsequently, the proliferation assay and transdermal absorption were determined by MTT method and immunohistochemical staining, respectively. First, the expression level of hEGF was recorded to be 14.83-ng/μL oil body and due to smaller size transgenic oil bodies expressing the recombinant oleosin–hEGF–hEGF, they were more skin permeable than those of control. Second, via the staining intensity of transgenic oil bodies was greater than EGF at all time points via immunohistochemical staining in transdermal absorption process. Lastly, activity assays of oil bodies expressed oleosin–hEGF–hEGF indicated that they stimulated the NIH/3T3 cell proliferation activity. Our results revealed oil-body-expressed oleosin–hEGF–hEGF was potential new material having implications in the field of medicine.