An 'activator-repressor' loop controls the anthocyanin biosynthesis in red-skinned pear.

The color of red-skinned pear (Pyrus spp.) is primarily attributed to accumulation of anthocyanins, which provide nutritional benefits for human health and are closely associated with the commercial value of fruits. Here, we reported the functional characterization of a R2R3-MYB repressor PyMYB107, which forms an 'activator-repressor' loop to control anthocyanin accumulation in the red-skinned pear. PyMYB107 overexpression inhibited anthocyanin biosynthesis in both pear calli and fruits, while virus-induced gene silencing of PyMYB107 increased anthocyanin accumulation in pear fruits. Furthermore, ectopic expression of PyMYB107 decreased anthocyanin accumulation in tomato, strawberry and tobacco. PyMYB107 can competitively bind to PybHLH3 with PyMYB10/MYB114, thereby suppressing the transcriptional activation of key anthocyanin biosynthesis genes, PyANS and PyUFGT. Site-directed mutagenesis showed that mutations within the R3 domain and EAR motif of PyMYB107 eliminated its repressive activity. Additionally, PyMYB107 exhibited a comparable expression pattern to PyMYB10/MYB114 and was transcriptionally activated by them. Our finding advanced comprehension of the repression mechanism underlying anthocyanin accumulation, providing valuable molecular insights into improving quality of pear fruits.

Comparison of structural variants detected by PacBio-CLR and ONT sequencing in pear.

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.

Map-based cloning of the pear gene MYB114 identifies an interaction with other transcription factors to coordinately regulate fruit anthocyanin biosynthesis.

Red fruits are popular and widely accepted by consumers because of an enhanced appearance and enriched anthocyanins. The molecular mechanism of anthocyanin regulation in red-skinned pear (Pyrus) has been studied, and the genes encoding the biosynthetic steps and several transcription factors (TFs) have been characterized. In this study, a candidate R2R3 MYB TF, PyMYB114, was identified by linkage to the quantitative trait loci (QTL) for red skin color on linkage group 5 in a population of Chinese pear (Pyrus bretschneideri). The function of PyMYB114 was verified by transient transformation in tobacco (Nicotinana tabacum) leaves and strawberry (Fragaria) and pear fruits, resulting in the biosynthesis of anthocyanin. Suppression of PyMYB114 could inhibit anthocyanin biosynthesis in red-skinned pears. The ERF/AP2 TF PyERF3 was found to interact with PyMYB114 and its partner PybHLH3 to co-regulate anthocyanin biosynthesis, as shown by a dual luciferase reporter system and a yeast two-hybrid assay. In addition, the transcript abundance of PyMYB114 and PyMYB10 were correlated, and co-transformation of these two genes into tobacco and strawberry led to enhanced anthocyanin biosynthesis. This interaction network provides insight into the coloration of fruits and the interaction of different TFs to regulate anthocyanin biosynthesis.