Improved Fully Convolutional Siamese Networks for Visual Object Tracking Based on Response Behaviour Analysis.

Siamese networks have recently attracted significant attention in the visual tracking community due to their balanced accuracy and speed. However, as a result of the non-update of the appearance model and the changing appearance of the target, the problem of tracking drift is a regular occurrence, particularly in background clutter scenarios. As a means of addressing this problem, this paper proposes an improved fully convolutional Siamese tracker that is based on response behaviour analysis (SiamFC-RBA). Firstly, the response map of the SiamFC is normalised to an 8-bit grey image, and the isohypse contours that represent the candidate target region are generated through thresholding. Secondly, the dynamic behaviour of the contours is analysed in order to check if there are distractors approaching the tracked target. Finally, a peak switching strategy is used as a means of determining the real tracking position of all candidates. Extensive experiments conducted on visual tracking benchmarks, including OTB100, GOT-10k and LaSOT, demonstrated that the proposed tracker outperformed the compared trackers such as DaSiamRPN, SiamRPN, SiamFC, CSK, CFNet and Staple and achieved state-of-the-art performance. In addition, the response behaviour analysis module was embedded into DiMP, with the experimental results showing the performance of the tracker to be improved through the use of the proposed architecture.

Pairwise Registration Algorithm for Large-Scale Planar Point Cloud Used in Flatness Measurement.

In this paper, an optimized three-dimensional (3D) pairwise point cloud registration algorithm is proposed, which is used for flatness measurement based on a laser profilometer. The objective is to achieve a fast and accurate six-degrees-of-freedom (6-DoF) pose estimation of a large-scale planar point cloud to ensure that the flatness measurement is precise. To that end, the proposed algorithm extracts the boundary of the point cloud to obtain more effective feature descriptors of the keypoints. Then, it eliminates the invalid keypoints by neighborhood evaluation to obtain the initial matching point pairs. Thereafter, clustering combined with the geometric consistency constraints of correspondences is conducted to realize coarse registration. Finally, the iterative closest point (ICP) algorithm is used to complete fine registration based on the boundary point cloud. The experimental results demonstrate that the proposed algorithm is superior to the current algorithms in terms of boundary extraction and registration performance.

CmLOX10 positively regulates drought tolerance through jasmonic acid -mediated stomatal closure in oriental melon (Cucumis melo var. makuwa Makino).

Drought stress severely impairs plant growth and production. Lipoxygenase (LOX), a master regulator for lipid peroxidation, is critical for direct or indirect response to abiotic stresses. Here, we found that drought stress induced the transcription of CmLOX10 in leaves of oriental melon seedlings. Reverse genetic approaches and physiological analyses revealed that silencing CmLOX10 increased drought susceptibility and stomatal aperture in oriental melon seedlings, and that ectopic overexpression of CmLOX10 in Arabidopsis enhanced drought tolerance and decreased the stomatal aperture. Moreover, the transcription of jasmonic acid (JA)-related genes and JA accumulation were significantly induced in CmLOX10-overexpressed Arabidopsis, which were reversely suppressed in CmLOX10-silenced seedlings during the stage of drought stress. Foliar application of JA further verified that JA enhanced drought tolerance and induced stomatal closure in leaves of melon seedlings. In addition, the feedback regulation of CmLOX10 was induced by JA signaling, and the expression level of CmMYC2 was increased by JA and drought treatment. Yeast one-hybrid analysis showed that CmMYC2 directly bound to the promoter of CmLOX10. In summary, we identified the important roles of CmLOX10 in the regulation of drought tolerance in oriental melon seedlings through JA- mediated stomatal closure and JA signaling-mediated feedback through CmMYC2.

Identification of a Novel Melon Transcription Factor CmNAC60 as a Potential Regulator of Leaf Senescence.

NAC transcription factors (TFs) play important roles in plants' responses to abiotic stresses and developmental processes, including leaf senescence. Oriental melon (Cucumis melo var. makuwa Makino) is an important vegetable crop in China and eastern Asia countries. However, little is known about the functions of the melon NAC family members. In this study, a phylogenetic tree was constructed to show that CmNAC60 and the senescence regulator AtNAP were in the same cluster, which implied that CmNAC60 might be a NAC related to leaf senescence. The expression analysis of CmNAC60 in different melon organs showed that the expression of CmNAC60 was highest in the male flowers and lowest in the hypocotyl. In addition, the expression level of CmNAC60 in the senescing leaves was significantly higher than in the non-senescing leaves. Similarly, the expression level of CmNAC60 in the dark-treated leaves was significantly higher than in the untreated leaves. Furthermore, the subcellular localization and transcriptional activation assays indicated that CmNAC60 was a nucleus localized NAC transcription factor with a C-terminal transactivation domain. An analysis of the tissue specific expression showed that the promoter of CmNAC60 may contain cis-acting regulatory elements responsive to leaf senescence. CmNAC60 overexpressing lines of Arabidopsis showed a precocious senescence compared with the wild type (WT). Collectively, our results showed that CmNAC60 was associated with leaf senescence, and could be potentially utilized in molecular breeding to improve melon yield or to extend the postharvest shelf life by delaying leaf senescence.

Isolation and functional validation of the CmLOX08 promoter associated with signalling molecule and abiotic stress responses in oriental melon, Cucumis melo var. makuwa Makino.

BACKGROUND: Lipoxygenases (LOXs) play significant roles in abiotic stress responses, and identification of LOX gene promoter function can make an important contribution to elucidating resistance mechanisms. Here, we cloned the CmLOX08 promoter of melon (Cucumis melo) and identified the main promoter regions regulating transcription in response to signalling molecules and abiotic stresses. RESULTS: The 2054-bp promoter region of CmLOX08 from melon leaves was cloned, and bioinformatic analysis revealed that it harbours numerous cis-regulatory elements associated with signalling molecules and abiotic stress. Five 5'-deletion fragments obtained from the CmLOX08 promoter-2054 (LP1), 1639 (LP2), 1284 (LP3), 1047 (LP4), and 418 bp (LP5)-were fused with a GUS reporter gene and used for tobacco transient assays. Deletion analysis revealed that in response to abscisic acid, salicylic acid, and hydrogen peroxide, the GUS activity of LP1 was significantly higher than that of the mock-treated control and LP2, indicating that the - 2054- to - 1639-bp region positively regulates expression induced by these signalling molecules. However, no deletion fragment GUS activity was induced by methyl jasmonate. In response to salt, drought, and wounding treatments, LP1, LP2, and LP4 promoted significantly higher GUS expression compared with the control. Among all deletion fragments, LP4 showed the highest GUS expression, indicating that - 1047 to - 1 bp is the major region regulating promoter activity and that the - 1047 to - 418-bp region positively regulates expression induced by salt, drought, and wounding, whereas the - 1284 to - 1047-bp region is a negative regulatory segment. Interestingly, although the GUS activity of LP1 and LP2 was not affected by temperature changes, that of LP3 was significantly induced by heat, indicating that the - 1284- to - 1-bp region is a core sequence responding to heat and the - 2054- to - 1284-bp region negatively regulates expression induced by heat. Similarly, the - 1047- to - 1-bp region is the main sequence responding to cold, whereas the - 2054- to - 1047-bp region negatively regulates expression induced by cold. CONCLUSIONS: We cloned the CmLOX08 promoter and demonstrated that it is a signalling molecule/stress-inducible promoter. Furthermore, we identified core and positive/negative regulatory regions responding to three signalling molecules and five abiotic stresses.

Identification and Characterization of LRR-RLK Family Genes in Potato Reveal Their Involvement in Peptide Signaling of Cell Fate Decisions and Biotic/Abiotic Stress Responses.

Leucine-rich repeat receptor-like kinases (LRR-RLKs) represent the largest subfamily of receptor-like kinases (RLKs) and play important roles in regulating growth, development, and stress responses in plants. In this study, 246 LRR-RLK genes were identified in the potato (Solanum tuberosum) genome, which were further classified into 14 subfamilies. Gene structure analysis revealed that genes within the same subgroup shared similar exon/intron structures. A signature small peptide recognition motif (RxR) was found to be largely conserved within members of subfamily IX, suggesting that these members may recognize peptide signals as ligands. 26 of the 246 StLRR-RLK genes were found to have arisen from tandem or segmental duplication events. Expression profiling revealed that StLRR-RLK genes were differentially expressed in various organs/tissues, and several genes were found to be responsive to different stress treatments. Furthermore, StLRR-RLK117 was found to be able to form homodimers and heterodimers with StLRR-RLK042 and StLRR-RLK052. Notably, the overlapping expression region of StLRR-RLK117 with Solanum tuberosum WUSCHEL (StWUS) suggested that the CLV3⁻CLV1/BAM⁻WUS feedback loop may be conserved in potato to maintain stem cell homeostasis within the shoot apical meristem.

Melon13-lipoxygenase CmLOX18 may be involved in C6 volatiles biosynthesis in fruit.

To better understand the function role of the melon CmLOX18 gene in the biosynthesis of C6 volatiles during fruit ripening, we biochemically characterized CmLOX18 and identified its subcellular localization in transgenic tomato plants. Heterologous expression in yeast cells showed that the molecular weight of the CmLOX18 protein was identical to that predicted, and that this enzyme possesseed lipoxygenase activity. Linoleic acid was demonstrated to be the preferred substrate for the purified recombinant CmLOX18 protein, which exhibited optimal catalytic activity at pH 4.5 and 30 °C. Chromatogram analysis of the reaction product indicated that the CmLOX18 protein exhibited positional specificity, as evidenced by its release of only a C-13 oxidized product. Subcellular localization analysis by transient expression in Arabidopsis protoplasts showed that CmLOX18 was localized to non-chloroplast organelles. When the CmLOX18 gene was transgenically expressed in tomato via Agrobacterium tumefaciens-mediated transformation, it was shown to enhance expression levels of the tomato hydroperoxide lyase gene LeHPL, whereas the expression levels of six TomLox genes were little changed. Furthermore, transgenic tomato fruits exhibited increases in the content of the C6 volatiles, namely hexanal, (Z)-3-hexanal, and (Z)-3-hexen-1-ol, indicating that CmLOX18 probably plays an important role in the synthesis of C6 compounds in fruits.

The Relationship between CmADHs and the Diversity of Volatile Organic Compounds of Three Aroma Types of Melon (Cucumis melo).

Alcohol dehydrogenase (ADH) plays an important role in aroma volatile compounds synthesis of plants. In this paper, we tried to explore the relationship between CmADHs and the volatile organic compounds (VOCs) in oriental melon. Three different aroma types of melon were used as materials. The principle component analysis of three types of melon fruit was conducted. We also measured the CmADHs expression level and enzymatic activities of ADH and alcohol acyl-transferase (AAT) on different stages of fruit ripening. An incubation experiment was carried out to investigate the effect of substrates and inhibitor (4-MP, 4-methylpyrazole) on CmADHs expression, ADH activity, and the main compounds of oriental melon. The results illustrated that ethyl acetate, hexyl acetate (E,Z)-3,6-nonadien-1-ol and 2-ethyl-2hexen-1-ol were the four principal volatile compounds of these three types of melon. AAT activity was increasing with fruit ripening, and the AAT activity in CH were the highest, whereas ADH activity peaked on 32 DAP, 2 days before maturation, and the ADH activity in CB and CG were higher than that in CH. The expression pattern of 11 CmADH genes from 24 to 36 day after pollination (DAP) was found to vary in three melon varieties. CmADH4 was only expressed in CG and the expression levels of CmADH3 and CmADH12 in CH and CB were much higher than that in CG, and they both peaked 2 days before fruit ripening. Ethanol and 4-MP decreased the reductase activity of ADH, the expression of most CmADHs and ethyl acetate or hexyl acetate contents of CB, except for 0.1 mM 4-MP, while aldehyde improved the two acetate ester contents. In addition, we found a positive correlation between the expression of CmADH3 and CmADH12 and the key volatile compound of CB. The relationship between CmADHs and VOCs synthesis of oriental melon was discussed.

The Alcohol Dehydrogenase Gene Family in Melon (Cucumis melo L.): Bioinformatic Analysis and Expression Patterns.

Alcohol dehydrogenases (ADH), encoded by multigene family in plants, play a critical role in plant growth, development, adaptation, fruit ripening and aroma production. Thirteen ADH genes were identified in melon genome, including 12 ADHs and one formaldehyde dehydrogenease (FDH), designated CmADH1-12 and CmFDH1, in which CmADH1 and CmADH2 have been isolated in Cantaloupe. ADH genes shared a lower identity with each other at the protein level and had different intron-exon structure at nucleotide level. No typical signal peptides were found in all CmADHs, and CmADH proteins might locate in the cytoplasm. The phylogenetic tree revealed that 13 ADH genes were divided into three groups respectively, namely long-, medium-, and short-chain ADH subfamily, and CmADH1,3-11, which belongs to the medium-chain ADH subfamily, fell into six medium-chain ADH subgroups. CmADH12 may belong to the long-chain ADH subfamily, while CmFDH1 may be a Class III ADH and serve as an ancestral ADH in melon. Expression profiling revealed that CmADH1, CmADH2, CmADH10 and CmFDH1 were moderately or strongly expressed in different vegetative tissues and fruit at medium and late developmental stages, while CmADH8 and CmADH12 were highly expressed in fruit after 20 days. CmADH3 showed preferential expression in young tissues. CmADH4 only had slight expression in root. Promoter analysis revealed several motifs of CmADH genes involved in the gene expression modulated by various hormones, and the response pattern of CmADH genes to ABA, IAA and ethylene were different. These CmADHs were divided into ethylene-sensitive and -insensitive groups, and the functions of CmADHs were discussed.

Heterologous Expression and Biochemical Characterization of Two Lipoxygenases in Oriental Melon, Cucumis melo var. makuwa Makino.

Lipoxygenases (LOXs) are a class of non-heme iron-containing dioxygenases that catalyse oxidation of polyunsaturated fatty acids to produce hydroperoxidation that are in turn converted to oxylipins. Although multiple isoforms of LOXs have been detected in several plants, LOXs in oriental melon have not attracted much attention. Two full-length LOX cDNA clones, CmLOX10 and CmLOX13 which have been isolated from oriental melon (Cucumis melo var. makuwa Makino) cultivar "Yumeiren", encode 902 and 906 amino acids, respectively. Bioinformatics analysis showed that CmLOX10 and CmLOX13 included all of the typical LOX domains and shared 58.11% identity at the amino acid level with each other. The phylogenetic analysis revealed that CmLOX10 and CmLOX13 were members of the type 2 13-LOX subgroup which are known to be involved in biotic and abiotic stress. Heterologous expression of the full-length CmLOX10 and truncated CmLOX13 in Escherichia coli revealed that the encoded exogenous proteins were identical to the predicted molecular weights and possessed the lipoxygenase activities. The purified CmLOX10 and CmLOX13 recombinant enzymes exhibited maximum activity at different temperature and pH and both had higher affinity for linoleic acid than linolenic acid. Chromatogram analysis of reaction products from the CmLOX10 and CmLOX13 enzyme reaction revealed that both enzymes produced 13S-hydroperoxides when linoleic acid was used as substrate. Furthermore, the subcellular localization analysis by transient expression of the two LOX fusion proteins in tobacco leaves showed that CmLOX10 and CmLOX13 proteins were located in plasma membrane and chloroplasts respectively. We propose that the two lipoxygenases may play different functions in oriental melon during plant growth and development.

The Effect of CmLOXs on the Production of Volatile Organic Compounds in Four Aroma Types of Melon (Cucumis melo).

Lipoxygenases (LOXs) play important role in the synthesis of volatile organic compounds (VOCs), which influence the aroma of fruit. In this study, we elucidate that there is a positive relationship between LOXs activity and VOC production in melon (Cucumis melo), and CmLOX genes are involved in fruit aroma generation in melon. To this end, we tested four aroma types of melon that feature a thin pericarp: two aromatic cultivars of the oriental melons (C. melo var. makuwa Makino), 'Yu Meiren' (YMR) and 'Cui Bao' (CB); a non-aromatic oriental pickling melon (C. melo var. conomon), 'Shao Gua' (SHAO); and a non-aromatic snake melon (C. melo L. var. flexuosus Naud), 'Cai Gua' (CAI). A principal component analysis (PCA) revealed that the aromas of SHAO and CAI are similar in nature because their ester contents are lower than those of YMR and CB. Ethyl acetate, benzyl acetate, (E, Z)-2, 6-nonadienal and menthol are four principal volatile compounds that affect the aromatic characteristics of these four types of melons. The LOX activity and total ester content in YMR were the highest among the examined melon varieties. The expression patterns of 18 CmLOX genes were found to vary based on the aromatic nature of the melon. Four of them were highly expressed in YMR. Moreover, we treated the fruit disks of YMR with LOX substrates (linoleic acid and linolenic acid) and LOX inhibitors (n-propyl gallate and nordihydroguariaretic acid). Substrate application promoted LOX activity and induced accumulation of hexanal, (2E)-nonenal and straight-chain esters, such as ethyl acetate. In contrast, LOX inhibitors decreased the levels of these compounds. The effect of CmLOXs in the biosynthesis of esters in melons are discussed.

The cinnamyl alcohol dehydrogenase gene family in melon (Cucumis melo L.): bioinformatic analysis and expression patterns.

Cinnamyl alcohol dehydrogenase (CAD) is a key enzyme in lignin biosynthesis. However, little was known about CADs in melon. Five CAD-like genes were identified in the genome of melons, namely CmCAD1 to CmCAD5. The signal peptides analysis and CAD proteins prediction showed no typical signal peptides were found in all CmCADs and CmCAD proteins may locate in the cytoplasm. Multiple alignments implied that some motifs may be responsible for the high specificity of these CAD proteins, and may be one of the key residues in the catalytic mechanism. The phylogenetic tree revealed seven groups of CAD and melon CAD genes fell into four main groups. CmCAD1 and CmCAD2 belonged to the bona fide CAD group, in which these CAD genes, as representative from angiosperms, were involved in lignin synthesis. Other CmCADs were distributed in group II, V and VII, respectively. Semi-quantitative PCR and real time qPCR revealed differential expression of CmCADs, and CmCAD5 was expressed in different vegetative tissues except mature leaves, with the highest expression in flower, while CmCAD2 and CmCAD5 were strongly expressed in flesh during development. Promoter analysis revealed several motifs of CAD genes involved in the gene expression modulated by various hormones. Treatment of abscisic acid (ABA) elevated the expression of CmCADs in flesh, whereas the transcript levels of CmCAD1 and CmCAD5 were induced by auxin (IAA); Ethylene induced the expression of CmCADs, while 1-MCP repressed the effect, apart from CmCAD4. Taken together, these data suggested that CmCAD4 may be a pseudogene and that all other CmCADs may be involved in the lignin biosynthesis induced by both abiotic and biotic stresses and in tissue-specific developmental lignification through a CAD genes family network, and CmCAD2 may be the main CAD enzymes for lignification of melon flesh and CmCAD5 may also function in flower development.