The R2R3 MYB Ruby1 is activated by two cold responsive ethylene response factors, via the retrotransposon in its promoter, to positively regulate anthocyanin biosynthesis in citrus.

Anthocyanins are natural pigments and dietary antioxidants that play multiple biological roles in plants and are important in animal and human nutrition. Low temperature (LT) promotes anthocyanin biosynthesis in many species including blood orange. A retrotransposon in the promoter of Ruby1, which encodes an R2R3 MYB transcription factor, controls cold-induced anthocyanin accumulation in blood orange flesh. However, the specific mechanism remains unclear. In this study, we characterized two LT-induced ETHYLENE RESPONSE FACTORS (CsERF054 and CsERF061). Both CsERF054 and CsERF061 can activate the expression of CsRuby1 by directly binding to a DRE/CRT cis-element within the retrotransposon in the promoter of CsRuby1, thereby positively regulating anthocyanin biosynthesis. Further investigation indicated that CsERF061 also forms a protein complex with CsRuby1 to co-activate the expression of anthocyanin biosynthetic genes, providing a dual mechanism for the upregulation of the anthocyanin pathway. These results provide insights into how LT mediates anthocyanin biosynthesis and increase the understanding of the regulatory network of anthocyanin biosynthesis in blood orange.

DNA methylation controlling abscisic acid catabolism responds to light to mediate strawberry fruit ripening.

Phytohormones, epigenetic regulation and environmental factors regulate fruit ripening but their interplay during strawberry fruit ripening remains to be determined. In this study, bagged strawberry fruit exhibited delayed ripening compared with fruit grown in normal light, correlating with reduced abscisic acid (ABA) accumulation. Transcription of the key ABA catabolism gene, ABA 8'-hydroxylase FaCYP707A4, was induced in bagged fruit. With light exclusion whole genome DNA methylation levels were up-regulated, corresponding to a delayed ripening process, while DNA methylation levels in the promoter of FaCYP707A4 were suppressed, correlating with increases in transcript and decreased ABA content. Experiments indicated FaCRY1, a blue light receptor repressed in bagged fruit and FaAGO4, a key protein involved in RNA-directed DNA methylation, could bind to the promoter of FaCYP707A4. The interaction between FaCRY1 and FaAGO4, and an increased enrichment of FaAGO4 directed to the FaCYP707A4 promoter in fruit grown under light suggests FaCRY1 may influence FaAGO4 to modulate the DNA methylation status of the FaCYP707A4 promoter. Furthermore, transient overexpression of FaCRY1, or an increase in FaCRY1 transcription by blue light treatment, increases the methylation level of the FaCYP707A4 promoter, while transient RNA interference of FaCRY1 displayed opposite phenotypes. These findings reveal a mechanism by which DNA methylation influences ABA catabolism, and participates in light-mediated strawberry ripening.

PbRbohH/J mediates ROS generation to regulate the growth of pollen tube in pear.

Respiratory burst oxidase homolog (Rboh) family genes play crucial functions in development and growth. However, comprehensive and systematic investigation of Rboh family members in Rosaceae and their specific functions during pear pollen development are still limited. In the study, 63 Rboh genes were identified from eight Rosaceae genomes (Malus domestica, Pyrus bretschneideri, Pyrus communis, Prunus persica, Rubus occidentalis, Fragaria vesca, Prunus mume and Prunus avium) and divided into seven main subfamilies (I-VII) according to phylogenetic and structural features. Different modes of gene duplication led to the expansion of Rboh family, with purifying selection playing a vital role in the evolution of Rboh genes. In addition, RNA sequencing and qRT-PCR results indicated that PbRbohH and PbRbohJ were specifically high-expressed in pear pollen. Subsequently, subcellular localization revealed that PbRbohH/J distributed at the plasma membrane. Furthermore, by pharmacological analysis and antisense oligodeoxynucleotide assay, PbRbohH/J were demonstrated to mediate the formation of reactive oxygen species (ROS) to manage pollen tube growth. In conclusion, our results provide useful insights into the functions, expression patterns, evolutionary history of the Rboh genes in pear and other Rosaceae species.

Elucidation of the GAUT gene family in eight Rosaceae species and function analysis of PbrGAUT22 in pear pollen tube growth.

MAIN CONCLUSION: The phylogenetic relationship and evolutionary history of the GAUT gene family were identified in 8 Rosaseae species. PbrGAUT22 was involved in controlling pollen tube growth by regulating the content of pectins. In plants, galacturonosyltransferases (GAUTs) were involved in homogalacturonan biosynthesis and functioned in maintaining pollen tube cell wall integrity. However, the feature and evolutionary history of the GAUT gene family in Rosaceae species and candidates in pear pollen tube growth remain unclear. Here, we identified 190 GAUT genes in 8 Rosaceae species, including Chinese white pear (Pyrus bretschneideri), European pear (Pyrus communis), apple (Malus × domestica), peach (Prunus persica), Japanese apricot (Prunus mume), sweet cherry (Prunus avium), woodland strawberry (Fragaria vesca) and black raspberry (Rubus occidentalis). Members in GAUT gene family were divided into 4 subfamilies according to the phylogenetic and structural analysis. Whole-genome duplication events and dispersed duplicates drove the expansion of the GAUT gene family. Among 23 pollen-expressed PbrGAUT genes in pear, PbrGAUT22 showed increased expression level during 1-6 h post-cultured pollen tubes. PbrGAUT22 was localized to the cytoplasm and plasma membrane. Knockdown of PbrGAUT22 expression in pollen tubes caused the decrease of pectin content and inhibited pear pollen tubes growth. Taken together, we investigated the identification and evolution of the GAUT gene family in Rosaceae species, and found that PbrGAUT22 played an essential role in the synthesis of pectin and the growth of pear pollen tubes.

The Origin and Evolution of RNase T2 Family and Gametophytic Self-incompatibility System in Plants.

Ribonuclease (RNase) T2 genes are found widely in both eukaryotes and prokaryotes, and genes from this family have been revealed to have various functions in plants. In particular, S-RNase is known to be the female determinant in the S-RNase-based gametophytic self-incompatibility (GSI) system. However, the origin and evolution of the RNase T2 gene family and GSI system are not well understood. In this study, 785 RNase T2 genes were identified in 81 sequenced plant genomes representing broad-scale diversity and divided into three subgroups (Class I, II, and III) based on phylogenetic and synteny network analysis. Class I was found to be of ancient origin and to emerge in green algae, Class II was shown to originate with the appearance of angiosperms, while Class III was discovered to be eudicot-specific. Each of the three major classes could be further classified into several subclasses of which some subclasses were found to be lineage-specific. Furthermore, duplication, deletion, or inactivation of the S/S-like-locus was revealed to be linked to repeated loss and gain of self-incompatibility in different species from distantly related plant families with GSI. Finally, the origin and evolutionary history of S-locus in Rosaceae species was unraveled with independent loss and gain of S-RNase occurred in different subfamilies of Rosaceae. Our findings provide insights into the origin and evolution of the RNase T2 family and the GSI system in plants.

PbrCalS5, a callose synthase protein, is involved in pollen tube growth in Pyrus bretschneideri.

MAIN CONCLUSION: Identification of CalS genes in seven Rosaceae species and functional characterization of PbrCalS5 in pear pollen tube growth by regulating callose deposition. Callose exists widely in angiosperms and has significant functions in a range of developmental processes. Callose is synthesized by callose synthase (CalS). However, the members of the callose synthase gene family and their evolutionary profiles, along with their biological functions, in species of the Rosaceae remain unknown. In this study, a total of 69 members of the CalS gene family in seven Rosaceae species (Fragaria vesca, Malus × domestica, Prunus avium, Pyrus bretschneideri, Prunus mume, Prunus persica and Rubus occidentalis) were identified and divided into six clades. Different types of gene duplication events contributed to the expansions of the CalS gene family in the seven species, with purifying selection playing a key role in the evolution of the CalS genes. Tissue-specific expression patterns analysis revealed that PbrCalS5 was highly expressed in the pear pollen tube and was selected for further functional analysis. Subcellular localization indicated that PbrCalS5 was localized in the plasma membrane and cell wall. Antisense oligodeoxynucleotide (AS-ODN) assays resulted in the inhibition of PbrCalS5 expression, leading to the decreased callose deposition in the pollen tube wall and subsequent inhibition of pear pollen tube growth. These results provide the theoretical basis for exploring the functional roles of CalS genes in pear pollen tube growth.

Suppression of liver receptor homolog-1 by microRNA-451 represses the proliferation of osteosarcoma cells.

Liver receptor homolog-1 (LRH-1) plays an important role in the onset and progression of many cancer types. However, the role of LRH-1 in osteosarcoma has not been well investigated. In this study, the critical role of LRH-1 in osteosarcoma cells was described. Quantitative polymerase chain reaction and Western blot analysis results revealed that LRH-1 was highly overexpressed in osteosarcoma cells. LRH-1 was knocked down by small interfering RNA (siRNA), and this phenomenon significantly inhibited osteosarcoma cell proliferation. Bioinformatics analysis results showed that LRH-1 contained putative binding sites of microRNA-451 (miR-451); this result was further validated through a dual-luciferase activity reporter assay. miR-451 was overexpressed in osteosarcoma cells through transfection of miR-451 mimics; miR-451 overexpression then significantly inhibited LRH-1 expression and cell proliferation. The loss of LRH-1 by siRNA or miR-451 mimics significantly impaired Wnt/ß-catenin activity, leading to G0/G1 cell cycle arrest. Results showed that LRH-1 is implicated in osteosarcoma. Therefore, miR-451-induced suppression of LRH-1 can be a novel therapy to treat osteosarcoma.

Association between single nucleotide polymorphism of IL15RA gene with susceptibility to ossification of the posterior longitudinal ligament of the spine.

BACKGROUND: Previous studies reported the association between single nucleotide polymorphism (SNP) of IL15 receptor alpha (IL15RA) gene with susceptibility to ossification of the posterior longitudinal ligament of the spine (OPLL). However, the results were still in controversy. Therefore, the purpose of the present study was to investigate the association between SNPs of IL15RA gene with susceptibility to OPLL in a Chinese Han population. METHODS: A total of 235 OPLL patients and 250 age-matched healthy controls were recruited. All the subjects were genotyped using the PCR (polymerase chain reaction)-based invader assay. A case-control study was performed to define the contribution of rs2228059 and rs2296139 to predisposition of OPLL. We also performed subgroup analysis according to the different gender. RESULTS: A significant association of rs2228059 with OPLL was observed in the Chinese Han population (p <0.001, OR = 1.63, 95% CI = 1.26-2.11). The subgroup analysis showed that there was a significant association between the allele frequency of rs2228059 and the susceptibility of OPLL in males (p = 0.002, OR = 1.72, 95% CI = 1.23-2.42). However, there was no significant association between SNP of rs2296139 and susceptibility to OPLL. CONCLUSIONS: The present study demonstrates that the SNP of rs2228059 in IL15RA gene is associated with susceptibility to OPLL in a Chinese Han population, especially in males.