DataSheet_2_Combined analysis of multi-omics reveals the potential mechanism of flower color and aroma formation in Macadamia integrifolia.xlsx

IntroductionMacadamia integrifolia Maiden & Betche is a domesticated high-value nut crop. The development of nut flower affects the fruit setting rate, yield and quality of nuts. Therefore, in this experiment, two varieties with different flower color, flowering time, flowering quantity and nut yield (single fruit weight) were selected as the research objects.MethodsTranscriptome (RNA-Seq) and metabolome (LC-MS/MS, GC-MS) analyses were performed to study the regulatory mechanisms of nut flower development, color and aroma.ResultsThe results indicated that plant hormone signal transduction, starch sucrose metabolism, phenylpropanoid metabolism, flavonoid biosynthesis, and anthocyanin biosynthesis pathways were related to nut flower development and flower color formation. In the early stage of flowering, most of the differentially expressed genes (DEGs) are involved in the IAA signal transduction pathway, while in the later stage, the brassinolide signal pathway is mainly involved. In starch and sugar metabolism, DEGs are mainly involved in regulating and hydrolyzing stored starch into small molecular sugars in flower tissues. In the phenylpropanoid biosynthesis pathway, DEGs are mainly related to the color and aroma (volatile organic compounds, VOCs) formation of nut flowers. Four color formation metabolites (anthocyanins) in nut flowers were found by LC-MS/MS detection. In addition, the VOCs showed no significant difference between red nut flowers (R) and white nut flowers (W), which was mainly reflected in the aroma formation stage (flowering time). And 12 common differentially accumulation metabolites (DAMs) were detected by GC-MS and LC-MS/MS. At the same time, the DEGs, AAT, LOX and PAL genes, were also identified to regulate key metabolite synthesis during nut flower development. These genes were further verified by qRT-PCR.ConclusionOur results provide insights to clarify the molecular mechanism of color and aroma formation during M. integrifolia flower development that pave the way for nut quality and yield breeding.</p

DataSheet_1_Combined analysis of multi-omics reveals the potential mechanism of flower color and aroma formation in Macadamia integrifolia.docx

IntroductionMacadamia integrifolia Maiden & Betche is a domesticated high-value nut crop. The development of nut flower affects the fruit setting rate, yield and quality of nuts. Therefore, in this experiment, two varieties with different flower color, flowering time, flowering quantity and nut yield (single fruit weight) were selected as the research objects.MethodsTranscriptome (RNA-Seq) and metabolome (LC-MS/MS, GC-MS) analyses were performed to study the regulatory mechanisms of nut flower development, color and aroma.ResultsThe results indicated that plant hormone signal transduction, starch sucrose metabolism, phenylpropanoid metabolism, flavonoid biosynthesis, and anthocyanin biosynthesis pathways were related to nut flower development and flower color formation. In the early stage of flowering, most of the differentially expressed genes (DEGs) are involved in the IAA signal transduction pathway, while in the later stage, the brassinolide signal pathway is mainly involved. In starch and sugar metabolism, DEGs are mainly involved in regulating and hydrolyzing stored starch into small molecular sugars in flower tissues. In the phenylpropanoid biosynthesis pathway, DEGs are mainly related to the color and aroma (volatile organic compounds, VOCs) formation of nut flowers. Four color formation metabolites (anthocyanins) in nut flowers were found by LC-MS/MS detection. In addition, the VOCs showed no significant difference between red nut flowers (R) and white nut flowers (W), which was mainly reflected in the aroma formation stage (flowering time). And 12 common differentially accumulation metabolites (DAMs) were detected by GC-MS and LC-MS/MS. At the same time, the DEGs, AAT, LOX and PAL genes, were also identified to regulate key metabolite synthesis during nut flower development. These genes were further verified by qRT-PCR.ConclusionOur results provide insights to clarify the molecular mechanism of color and aroma formation during M. integrifolia flower development that pave the way for nut quality and yield breeding.</p

Laser-scanning confocal microscopy of strain YIM 63111 tagged with <i>egfp</i> for colonization analysis.

<p>a) Wild-type strain YIM 63111; b) <i>egfp</i>-tagged strain YIM 63111; c) Outer colonization of the root 14 days after inoculation; d) Outer and inner colonization of the root 14 days after inoculation. EGFP was excited with a 488-nm laser, and fluorescence was detected at 505–530 nm.</p

Artemisinin content in <i>A</i>. <i>annua</i> plants grown for 74 days after YIM 63111 inoculation compared with untreated plants (No. 0).

<p>Nos. 1–10 indicate the seedlings that were inoculated with serially diluted bacterial suspensions: 1.84×10⁹, 3.68×10⁸, 7.38×10⁷, 1.48×10⁷, 2.96×10⁶, 5.90×10⁵, 1.18×10⁵, 2.36×10⁴, 4.72×10³ and 900∼1.0×10³ CFU ml⁻¹. * indicates a significant difference (<i>P</i><0.05, and the error bars represent the standard deviation.</p

Transcript abundance of the <i>ADS</i>, <i>ASQS</i>, <i>CPR</i> and <i>CYP71AV1</i> genes and artemisinin content in <i>A</i>. <i>annua</i> plants grown for 64 days after YIM 63111 inoculation compared with untreated plants (No. 0).

<p>Nos. 7 & 8 indicate the seedlings that were inoculated with serially diluted bacterial suspensions, 2.0×10⁴ CFU ml⁻¹, 4.0×10³ CFU ml⁻¹. * indicates a significant difference, and the error bars represent the standard deviation.</p

Special Enrichment Strategies Greatly Increase the Efficiency of Missing Proteins Identification from Regular Proteome Samples

As part of the Chromosome-Centric Human Proteome Project (C-HPP) mission, laboratories all over the world have tried to map the entire missing proteins (MPs) since 2012. On the basis of the first and second Chinese Chromosome Proteome Database (CCPD 1.0 and 2.0) studies, we developed systematic enrichment strategies to identify MPs that fell into four classes: (1) low molecular weight (LMW) proteins, (2) membrane proteins, (3) proteins that contained various post-translational modifications (PTMs), and (4) nucleic acid-associated proteins. Of 8845 proteins identified in 7 data sets, 79 proteins were classified as MPs. Among data sets derived from different enrichment strategies, data sets for LMW and PTM yielded the most novel MPs. In addition, we found that some MPs were identified in multiple-data sets, which implied that tandem enrichments methods might improve the ability to identify MPs. Moreover, low expression at the transcription level was the major cause of the “missing” of these MPs; however, MPs with higher expression level also evaded identification, most likely due to other characteristics such as LMW, high hydrophobicity and PTM. By combining a stringent manual check of the MS₂ spectra with peptides synthesis verification, we confirmed 30 MPs (neXtProt PE2 ∼ PE4) and 6 potential MPs (neXtProt PE5) with authentic MS evidence. By integrating our large-scale data sets of CCPD 2.0, the number of identified proteins has increased considerably beyond simulation saturation. Here, we show that special enrichment strategies can break through the data saturation bottleneck, which could increase the efficiency of MP identification in future C-HPP studies. All 7 data sets have been uploaded to ProteomeXchange with the identifier PXD002255

Tissue-Based Proteogenomics Reveals that Human Testis Endows Plentiful Missing Proteins

Investigations of missing proteins (MPs) are being endorsed by many bioanalytical strategies. We proposed that proteogenomics of testis tissue was a feasible approach to identify more MPs because testis tissues have higher gene expression levels. Here we combined proteomics and transcriptomics to survey gene expression in human testis tissues from three post-mortem individuals. Proteins were extracted and separated with glycine- and tricine-SDS-PAGE. A total of 9597 protein groups were identified; of these, 166 protein groups were listed as MPs, including 138 groups (83.1%) with transcriptional evidence. A total of 2948 proteins are designated as MPs, and 5.6% of these were identified in this study. The high incidence of MPs in testis tissue indicates that this is a rich resource for MPs. Functional category analysis revealed that the biological processes that testis MPs are mainly involved in are sexual reproduction and spermatogenesis. Some of the MPs are potentially involved in tumorgenesis in other tissues. Therefore, this proteogenomics analysis of individual testis tissues provides convincing evidence of the discovery of MPs. All mass spectrometry data from this study have been deposited in the ProteomeXchange (data set identifier PXD002179)