Allelic variation of BBX24 is a dominant determinant controlling red coloration and dwarfism in pear.

Variation in anthocyanin biosynthesis in pear fruit provides genetic germplasm resources for breeding, while dwarfing is an important agronomic trait, which is beneficial to reduce the management costs and allow for the implementation of high-density cultivation. Here, we combined bulked segregant analysis (BSA), quantitative trait loci (QTL), and structural variation (SV) analysis to identify a 14-bp deletion which caused a frame shift mutation and resulted in the premature translation termination of a B-box (BBX) family of zinc transcription factor, PyBBX24, and its allelic variation termed PyBBX24ΔN14. PyBBX24ΔN14 overexpression promotes anthocyanin biosynthesis in pear, strawberry, Arabidopsis, tobacco, and tomato, while that of PyBBX24 did not. PyBBX24ΔN14 directly activates the transcription of PyUFGT and PyMYB10 through interaction with PyHY5. Moreover, stable overexpression of PyBBX24ΔN14 exhibits a dwarfing phenotype in Arabidopsis, tobacco, and tomato plants. PyBBX24ΔN14 can activate the expression of PyGA2ox8 via directly binding to its promoter, thereby deactivating bioactive GAs and reducing the plant height. However, the nuclear localization signal (NLS) and Valine-Proline (VP) motifs in the C-terminus of PyBBX24 reverse these effects. Interestingly, mutations leading to premature termination of PyBBX24 were also identified in red sports of un-related European pear varieties. We conclude that mutations in PyBBX24 gene link both an increase in pigmentation and a decrease in plant height.

Lactobacillus casei combined with Lactobacillus reuteri alleviate pancreatic cancer by inhibiting TLR4 to promote macrophage M1 polarization and regulate gut microbial homeostasis.

BACKGROUND: Pancreatic cancer is a highly lethal disease with no effective treatments. Lactobacillus casei (L. casei) and Lactobacillus reuteri (L. reuteri) exhibited therapeutic effects on several cancers, but their roles in pancreatic cancer are unknown. This study aims to explore how L. casei & L. reuteri influence pancreatic cancer and the underlying mechanisms. METHODS: Pancreatic cancer cells were treated with L. casei & L. reuteri and co-cultured with macrophages in a transwell system in vitro. Pancreatic cancer xenograft model was established and L. casei & L. reuteri was used to treat mice in vivo. MTT, CCK-8 assay or immunohistochemical staining were used to determine the proliferation of pancreatic cancer cells or tumor tissues. Transwell assay was applied to test the migration and invasion of pancreatic cells. RT-qPCR was utilized to assess TLR4 and MyD88 expressions in pancreatic cells or tumor tissues. WB, immunofluorescence staining, or flow cytometry was used to evaluate the M1/M2 polarization of macrophages. Besides, the composition of gut microbiota of tumor-bearing mice was determined by 16 S rRNA sequencing, and ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) untargeted metabolomics was used to evaluate the metabolic profiles of feces. RESULTS: L. casei & L. reuteri inhibited the proliferation, migration, invasion of pancreatic cancer cells and pancreatic cancer cell-induced M2 polarization of macrophages by suppressing TLR4. Meanwhile, L. casei & L. reuteri repressed pancreatic cancer growth and promoted M1 macrophage polarization. Besides, L. casei & L. reuteri reduced fecal Alloprevotella and increased fecal azelate and glutamate in nude mice, while TLR4 inhibitor TAK-242 increased Clostridia UCG-014, azelate, uridine, methionine sulfoxide, oxypurinol, and decreased glyceryl monoester in the feces of pancreatic tumor-bearing mice. Fecal oxypurinol and glyceryl monoester levels were positively or negatively associated with gut Clostridia UCG-014 abundance, respectively. CONCLUSION: L. casei & L. reuteri alleviate pancreatic cancer by inhibiting TLR4 to promote macrophage M1 polarization and regulate gut microbial homeostasis.

Contrasting genetic variation and positive selection followed the divergence of NBS-encoding genes in Asian and European pears.

BACKGROUND: The NBS disease-related gene family coordinates the inherent immune system in plants in response to pathogen infections. Previous studies have identified NBS-encoding genes in Pyrus bretschneideri ('Dangshansuli', an Asian pear) and Pyrus communis ('Bartlett', a European pear) genomes, but the patterns of genetic variation and selection pressure on these genes during pear domestication have remained unsolved. RESULTS: In this study, 338 and 412 NBS-encoding genes were identified from Asian and European pear genomes. This difference between the two pear species was the result of proximal duplications. About 15.79% orthologous gene pairs had Ka/Ks ratio more than one, indicating two pear species undergo strong positive selection after the divergence of Asian and European pear. We identified 21 and 15 NBS-encoding genes under fire blight and black spot disease-related QTL, respectively, suggesting their importance in disease resistance. Domestication caused decreased nucleotide diversity across NBS genes in Asian cultivars (cultivated 6.23E-03; wild 6.47E-03), but opposite trend (cultivated 6.48E-03; wild 5.91E-03) appeared in European pears. Many NBS-encoding coding regions showed Ka/Ks ratio of greater than 1, indicating the role of positive selection in shaping diversity of NBS-encoding genes in pear. Furthermore, we detected 295 and 122 significantly different SNPs between wild and domesticated accessions in Asian and European pear populations. Two NBS genes (Pbr025269.1 and Pbr019876.1) with significantly different SNPs showed >5x upregulation between wild and cultivated pear accessions, and > 2x upregulation in Pyrus calleryana after inoculation with Alternaria alternata. We propose that positively selected and significantly different SNPs of an NBS-encoding gene (Pbr025269.1) regulate gene expression differences in the wild and cultivated groups, which may affect resistance in pear against A. alternata. CONCLUSION: Proximal duplication mainly led to the different number of NBS-encoding genes in P. bretschneideri and P. communis genomes. The patterns of genetic diversity and positive selection pressure differed between Asian and European pear populations, most likely due to their independent domestication events. This analysis helps us understand the evolution, diversity, and selection pressure in the NBS-encoding gene family in Asian and European populations, and provides opportunities to study mechanisms of disease resistance in pear.

Mining and evolution analysis of lateral organ boundaries domain (LBD) genes in Chinese white pear (Pyrus bretschneideri).

BACKGROUND: The lateral organ boundaries domain (LBD) gene is a plant-specific transcription factor that plays a critical role in diverse biological processes. However, the evolution and functional divergence of the LBD gene family has not yet been characterized for the Chinese White Pear. RESULTS: In our study, a total of 60 PbrLBDs were identified in the pear genome. The PbrLBD gene family was divided into two classes based on gene structure and phylogenetic analysis: class I (53) and class II (7). Cis-acting element analysis results suggested that PbrLBDs may participate in various biological processes, such as flavonoid biosynthetic and stress response. Synteny analysis results indicated that segmental duplication played a key role in the expansion of the PbrLBD gene family. The mean Ks and 4DTv values showed that the PbrLBD gene family had undergone only one recent whole-genome duplication event occurring at 30-45 MYA. Purifying selection was a primary force during the PbrLBD gene family evolution process. Transcriptome data analysis revealed that 10 PbrLBDs were expressed in all six examined tissues, and 73.33% of members in the PbrLBD gene family were expressed in pear sepal. qRT-PCR was conducted to verify the expression levels of 11 PbrLBDs in these six tissues. Specifically, PbrLBD20, PbrLBD35 and PbrLBD53 genes were down-regulated when anthocyanin concentrations were high, whereas PbrLBD33 was significantly up-regulated in pear when anthocyanin concentrations were high. Furthermore, PbrLBD20, one of the candidate genes related to anthocyanins was localized in the nucleus. CONCLUSIONS: Our analysis provides valuable information for understanding the evolution of the PbrLBD gene family, and provides new insights into the regulation of pear pigment metabolism and lays a foundation for the future disclosure of the molecular mechanism of LBD gene regulating flavonoid metabolism.