ATAC-Seq

Partial results are shown below:

1. What type of controls are necessary for ATAC-Seq experiments?

In the realm of ATAC-Seq experiments, the incorporation of diverse controls is imperative to uphold precision and dependability. The utilization of an input DNA control serves the purpose of normalizing potential biases in DNA preparation and sequencing processes. Technical replicates play a crucial role in confirming the reproducibility and reliability of the outcomes. The inclusion of positive controls, representing established open chromatin regions, acts as a mechanism to authenticate the protocol. On the other hand, negative controls, indicative of anticipated closed chromatin regions, aid in the detection and mitigation of background noise.

2. How does ATAC-Seq compare to other chromatin accessibility assays like DNase-Seq and FAIRE-Seq?

DNase-Seq, ATAC-Seq, and FAIRE-Seq are all techniques utilized for studying open chromatin regions. DNase-Seq employs the use of DNase I endonuclease to identify accessible chromatin regions, FAIRE-Seq involves sonication followed by phenol-chloroform enrichment, and ATAC-Seq utilizes Tn5 transposase for enrichment and amplification. The high activity of Tn5 transposase makes ATAC-Seq a straightforward, efficient method requiring only 500-50,000 cells. The sensitivity and specificity of ATAC-Seq are comparable to DNase-Seq and superior to FAIRE-Seq.

3. What techniques are commonly combined with ATAC-seq for research?

ATAC-seq + RNA-seq: Generally, RNA-seq is performed prior to ATAC-seq to identify differentially expressed genes and enriched pathways, which suggest correlations. To determine which factors regulate these target genes, motif analysis via ATAC-seq can identify potential regulatory elements. Subsequent validation can be carried out through additional experiments or ChIP-seq.

ATAC-seq + Hi-C: For studies investigating the effects of higher-order chromatin structures on biological processes, ATAC-seq is often used alongside Hi-C. Hi-C provides information on higher-order structures like compartment A/B, TADs, and loops, which indicate correlations. ATAC-seq can further identify promoters, enhancers, and other elements, elucidating how these structures influence gene expression.

ATAC-seq + Histone Modifications: While ATAC-seq can predict the accessibility of specific sites and potential transcription factor binding, it does not reveal whether these factors promote or inhibit gene expression. Combining ATAC-seq with histone modification data (e.g., H3K27ac for activation or H3K27me3 for repression) can provide a more comprehensive understanding, making the data more robust and reliable.

Multiomics analyses of the effects of LED white light on the ripening of apricot fruits

Journal: Journal of Advanced Research
Impact factor: 12.822
Published: Available online 9 January 2024

Background

Apricot (Prunus armeniaca L.), native to Central Asia and widely cultivated in regions like the Mediterranean and China, is popular for its color, taste, and nutrients but has a short storage life of 3-5 days. The ripening and senescence process, regulated by ethylene, involves complex physiological changes. Despite extensive research, the metabolite changes and regulatory mechanisms during postharvest storage are not fully understood. Light-emitting diodes (LEDs) offer an eco-friendly method to extend shelf life by delaying senescence and maintaining quality. This study investigates the effects of LED white light on apricot fruit quality, transcriptome, metabolome, and chromatin accessibility, aiming to improve storage methods and prolong shelf life.

Materials & Methods

Results

In this study, fruit transcriptomics utilized PCC as an index to assess biological replicate correlation, showing high values (>0.9), indicating strong similarity (Fig. 1F). Analysis revealed differential gene expression patterns across comparisons: CK-0 d vs. CK-8 d (6,256 DEGs), CK-0 d vs. LED-8 d (8,110 DEGs), and CK-8 d vs. LED-8 d (1,792 DEGs), with varying overlaps (Fig. 1I). KEGG and GO enrichment highlighted pathways like secondary metabolite biosynthesis and regulatory roles of DEGs in flavor, texture, color, ethylene biosynthesis, transcription factors, and oxidative processes. LED light regulated 56 DEGs in fruit ripening, impacting metabolic pathways and transcriptional activities.

Fig. 1. Effects of LED treatment on the physiological quality and transcription of apricot fruits.

Combined transcriptomic and metabolomic analyses revealed that under LED irradiation, UDP-sugar pyrophosphorylase (USP) was upregulated, enhancing D-glucuronate synthesis, while inositol oxygenase (MIOX) and aldehyde dehydrogenase (ALDH) were downregulated, reducing myo-inositol accumulation. LED treatment also decreased L-dehydroascorbate accumulation by affecting L-ascorbate peroxidase (APX) and L-ascorbate oxidase (AO) activities. Additionally, LED-induced increases in cinnamoyl-CoA levels upregulated chalcone synthase (CHS) and phlorizin synthase (PTG1), altering flavonoid metabolism and promoting anthocyanin biosynthesis through dihydroflavonol 4-reductase (DFR) upregulation.

Fig. 2. The metabolic pathway of flavonoid biosynthesis was analyzed by combining the transcriptome and metabolome data.

ATAC-seq analysis of apricot fruits post LED treatment revealed 616.85 million clean reads with high Q30 base percentages, indicating robust data quality. Correlation analysis showed strong reproducibility among biological replicates within each group. Differential analysis identified 4492, 1769, and 123 differentially accessible regions (DARs) in CK-0 d vs. CK-8 d, CK-0 d vs. LED-8 d, and CK-8 d vs. LED-8 d, respectively. Motif analysis highlighted 76 transcription factors (TFs), including ethylene-responsive factors (ERF2, ERF9, ERF069, ERF104, and MYB124), implicated in regulating DARs and target genes. These findings underscore LED-induced changes in chromatin accessibility and TF regulation, influencing gene expression and fruit development in apricots.

Fig. 3. Overview of the ATAC-seq data of apricot fruit ripening.

Fig. 4. (A) Differential gene expression and heatmap of chromatin open regions. (B) The metabolic pathway of phenylpropanoid biosynthesis was analyzed by combining the transcriptome, metabolome, and ATAC-seq data.

In LED-treated apricot fruits, integrating transcriptomic and ATAC-seq analyses revealed genes with altered chromatin accessibility and expression levels. HCT exhibited reduced chromatin accessibility and decreased gene expression, while POD and 4CL showed variable changes in accessibility paired with mixed upregulation and downregulation of gene expression.

Metabolomic analysis highlighted phenylpropanoid biosynthesis as significantly impacted by LED treatment. Changes in HCT expression influenced specific metabolite accumulation like p-coumaroyl shikimic acid and caffeoyl quinic acid. POD and 4CL, with altered chromatin accessibility and varied gene expression, played roles in lignin synthesis pathways crucial for maintaining apricot fruit quality during storage.

Fig. 5 Model of the regulatory network of LED light during ripening of harvested apricot fruits.

Conclusion

LED treatment altered pathways for ascorbic acid and uronate metabolism, flavonoid biosynthesis, and phenylpropanoid biosynthesis in apricot fruits, influencing gene expression related to plant hormone signaling, fruit quality, and antioxidant activity. This research highlights LED's potential to extend storage and shelf life in fruits and vegetables.

Reference:

1. Bai C, Zheng Y, Watkins CB, et al. Multiomics analyses of the effects of LED white light on the ripening of apricot fruits. Journal of Advanced Research. 2024.