GALACTINOL SYNTHASE is the first committed enzyme in the raffinose biosynthetic pathway. We have previously characterized the maize (Zea mays) GALACTINOL SYNTHASE2 gene (ZmGOLS2) as abiotic stress induced. To further investigate the regulation of ZmGOLS2 gene expression, individual luciferase expression vectors,in which the luciferase gene was controlled by different lengths of the ZmGOLS2 promoter, were co-transfected into maize protoplasts with either a ZmDREB2A- or a GFP-expression vector. Over-expression of ZmDREB2A up-regulated both the expression of the luciferase gene controlled by the ZmGOLS2 promoter and the endogenous ZmGOLS2 gene in protoplasts. Only one of the two DRE elements in the ZmGOLS2 promoter was identified as necessary for this up-regulation. Expression vectors of GFP, ZmGOLS2 or ZmDREB2A were stably transformed into Arabidopsis. Expression of ZmDREB2A up-regulated the AtGOLS3 gene but only over-expression of ZmGOLS2 resulted in hyper-accumulation of galactinol and raffinose. Regardless, under drought-, heat shock-, high osmotic- or salinity-stress conditions, both the ZmGOLS2- and the ZmDREB2A- expressing plants had greater germination percentages, greater percentages of seedlings becoming autotropic, and/or greater survival percentages during/after stress than the control plants. Under normal growing conditions, transgenic Arabidopsis plants expressing the ZmGOLS2 gene had similar growth to that of untransformed wild type or GFP-expressing control plants, whereas ZmDREB2A over-expressing plants exhibited retarded growth relative to either of the controls. These data suggest that over-expression of ZmGOLS2, rather than the transcription factor ZmDREB2A, is a more practical target for generation of abiotic-stress tolerant crops.

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Summary Raffinose is thought to play an important role in plant tolerance of abiotic stress. We report here that maize HEAT SHOCK FACTOR A2 (ZmHSFA2) and HEAT SHOCK BINDING PROTEIN 2 (ZmHSBP2) physically interact with each other and antagonistically modulate expression of GALACTINOL SYNTHASE2 (ZmGOLS2) and raffinose biosynthesis in transformed maize protoplasts and Arabidopsis plants. Overexpression of ZmHSFA2 in Arabidopsis increased the expression of Arabidopsis AtGOLS1, AtGOLS2 and AtRS5 (RAFFINOSE SYNTHASE), increased the raffinose content in leaves and enhanced plant heat stress tolerance. Contrary to ZmHSFA2, overexpression of ZmHSBP2 in Arabidopsis decreased expression of AtGOLS1, AtGOLS2 and AtRS5, decreased the raffinose content in leaves and reduced plant heat stress tolerance. ZmHSFA2 and ZmHSBP2 also interact with their Arabidopsis counterparts AtHSBP and AtHSFA2 as determined using bimolecular fluorescence complementation assays. Furthermore, endogenous ZmHSBP2 and Rluc, controlled by the ZmHSBP2 promoter, are transcriptionally activated by ZmHSFA2 and inhibited by ZmHSBP2 in maize protoplasts. These findings provide insights into the transcriptional regulation of raffinose biosynthetic genes, and the tolerance their product confers to plant heat stress.

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BackgroundThe outcomes for open tibial fractures with severe soft tissue injury are still a great challenge for all the trauma surgeons in the treatment. However, most of the existing open tibial fracture models can only provide minimal soft tissue injury which cannot meet the requirement of severe trauma research. Our goal is to investigate a novel tibial fracture model providing different fractures combined with soft tissue injury for better application in trauma research.MethodsA total of 144 Sprague-Dawley rats were randomly divided into 4 groups. With group 1 as control, the other groups sustained different right tibial fractures by the apparatus with buffer disc settings either 3 mm, 10 mm, or 15 mm. X-ray and computed tomography angiography (CTA) were performed at 6 h to evaluate the fracture patterns and vascular injuries. Peripheral blood and tibialis anterior muscle were harvested at 6 h, 1 day, 3 days, 7 days, 14 days, and 28 days for ELISA and histological analysis.ResultsX-ray and μCT results indicated that different fractures combined with soft tissue injuries could be successfully provided in this model. According to OTA and Gustilo classification, the fractures and soft tissue injuries were evaluated and defined: 36 type I in group 2, 34 type II in group 3, and 36 type III in group 4. The CTA confirmed no arterial injuries in groups 1 and 2, 2 arterial injuries in group 3, and 35 in group 4. ELISA indicated that the levels of pro-inflammatory cytokines TNF-α and IL-1β were significantly higher in group 4 than in other groups, and the levels of anti-inflammatory cytokines TGF-β and IL-10 were significantly higher in surgery groups than in group 1 in later stage or throughout the entire process. HE, Masson, and caspase-3 stains confirmed the most severe inflammatory cell infiltration and apoptosis in group 4 which lasted longer than that in groups 2 and 3.ConclusionsThe novel apparatus was valuable in performing different fractures combined with soft tissue injuries in a rat tibial fracture model with high reproducibility and providing a new selection for trauma research in the future.

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The neat, uncontaminated wound margins and the fact that the amputated auricle was preserved in ice, even though it had been amputated 10 h before the operation, were prerequisites for successful auricle replantation. The microvascular anastomosis technique played a very important role in the survival of the amputated auricle. The postoperative treatment, observation and nursing were difficult but vital aspects of the procedure. The establishment of an effective venous return was a sign of the survival of the amputated auricle.