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Background: Structural variations (SVs) have recently become a topic of great interest in the area of genetic diversity and trait regulation. As genomic sequencing technologies have rapidly advanced, longer reads have been used to identify SVs at high resolution and with increased accuracy. It is important to choose a suitable sequencing platform and appropriate sequencing depth for SV detection in the pear genome. Results: In this study, two types of long reads from sequencing platforms, continuous long reads from Pacific Biosciences (PB-CLR) and long reads from Oxford Nanopore Technologies (ONT), were used to comprehensively analyze and compare SVs in the pear genome. The mapping rate of long reads was higher when the program Minimap2 rather than the other three mapping tools (NGMLR, LRA and Winnowmap2) was used. Three SV detection programs (Sniffles_v2, CuteSV, and Nanovar) were compared, and Nanovar had the highest sensitivity in detecting SVs at low sequencing depth (10–15×). A sequencing depth of 15× was suitable for SV detection in the pear genome using Nanovar. SVs detected by Sniffles_v2 and CuteSV with ONT reads had the high overlap with presence/absence variations (PAVs) in the pear cultivars 'Bartlett' and 'Dangshansuli', both of them with 38% of insertions and 55% of deletions overlapping with PAVs at sequencing depth of 30×. For the ONT sequencing data, over 37,526 SVs spanning ~ 28 Mb were identified by all three software packages for the 'Bartlett' and 'Dangshansuli' genomes. Those SVs were annotated and combined with transcriptome profiles derived from 'Bartlett' and 'Dangshansuli' fruit flesh at 60 days after cross-pollination. Several genes related to levels of sugars, acid, stone cells, and aromatic compounds were identified among the SVs. Transcription factors were then predicted among those genes, and results included bHLH, ERF, and MYB genes. Conclusion: SV detection is of great significance in exploring phenotypic differences between pear varieties. Our study provides a framework for assessment of different SV software packages and sequencing platforms that can be applied in other plant genome studies. Based on these analyses, ONT sequencing data was determined to be more suitable than PB-CLR for SV detection in the pear genome. This analysis model will facilitate screening of genes related to agronomic traits in other crops. [ABSTRACT FROM AUTHOR]

Background: Osteosarcoma (OS) is the most common primary bone cancer in adolescents and lung metastasis is the leading cause of death in patients with OS. However, the molecular mechanisms that promote OS growth and metastasis remain unknown. Methods: We investigated the expression of myosin light chain kinase family members between metastasis and non-metastasis patients in the TARGET database and ensured that only myosin light chain kinase family member 4 (MYLK4) had higher expression in metastatic osteosarcoma patients. Then we confirmed the results by immunohistochemistry (IHC) and Western blotting (WB) of OS tissues. The effect of MYLK4 on the metastasis and proliferation of OS cells was investigated by wound healing, Transwell and colony-formation assays. Mass spectrum analysis was used to ensure the new binding protein of MYLK4. Tissue microarrays analysis was used to show the correlation between MYLK4 and pEGFR (Y1068). A series of in vivo experiments were conducted to reveal the mechanisms by which MYLK4 modulated the metastasis and proliferation of OS. Results: Myosin Light Chain Kinase Family Member 4 (MYLK4) was significantly upregulated in metastatic human OS tissues. Growth and metastasis of OS could be accelerated by MYLK4 overexpression, whereas silencing MYLK4 expression resulted in decreased cell growth and metastasis. Mechanistically, mass spectrum analysis showed that MYLK4 interacted with the epidermal growth factor receptor (EGFR) in osteosarcoma cells and promoted growth and metastasis via the EGFR signaling pathway. Tissue microarrays analysis also showed that MYLK4 expression had a positive correlation with the expression of pEGFR (Y1068). Moreover, the EGFR inhibitor gefitinib could partially reverse the effect of cell proliferation and metastasis caused by MYLK4 overexpression. Importantly, the combination of MYLK4 and EGFR inhibitors had synergistic effects on growth and metastasis of OS in vitro and in vivo. Conclusion: Our results indicate that MYLK4 promotes OS growth and metastasis by activating the EGFR signaling pathway and can be a novel therapeutic target for the treatment of OS patients. [ABSTRACT FROM AUTHOR]

Background: Osteosarcoma (OS) is a common malignant cancer in children and adolescents and has a cure rate that has not improved in the last two decades. CYT997 (lexibulin) is a novel potent microtubule-targeting agent with various anticancer activities, such as proliferation inhibition, vascular disruption, and cell cycle arrest and apoptosis induction, in multiple cancers. However, the direct cytotoxic mechanisms of CYT997 have not yet been fully characterized. Methods: We evaluated apoptosis and autophagy in human osteosarcomas after treatment with CYT997 and investigated the underlying mechanisms. To explore relationships, we used the reactive oxygen species (ROS) scavenger N-acetyl cysteine (NAC), PERK inhibitor GSK2606414, ERO1 inhibitor EN460 and mitochondrial targeted protection peptide elamipretide. BALB/c-nu mice were inoculated with 143B tumor cells to investigate the in vivo effect of CYT997. Results: We explored the efficacy and mechanism of CYT997 in osteosarcoma (OS) in vitro and in vivo and demonstrated that CYT997 potently suppresses cell viability and induces apoptosis and autophagy. CYT997 triggered production of ROS and exerted lethal effects via endoplasmic reticulum (ER) stress in OS cells. NAC attenuated these effects. The PERK inhibitor GSK2606414, which can block the ER stress pathway, reduced ROS production and enhanced cell viability. Moreover, activation of ERO1 in the ER stress pathway was responsible for inducing ROS production. ROS produced by the mitochondrial pathway also aggravate ER stress. Protection of mitochondria can reduce apoptosis and autophagy. Finally, CYT997 prominently reduced tumor growth in vivo. Conclusions: This study suggests that CYT997 induces apoptosis and autophagy in OS cells by triggering mutually enhanced ER stress and ROS and may thus be a promising agent against OS. [ABSTRACT FROM AUTHOR]