Transcriptome Gene Expression Profiling Research Articles

Molecular Basis of Cerebral Vasospasm: What Can We Learn from Transcriptome and Temporal Gene Expression Profiling in Intracranial Aneurysm?

Cerebral vasospasm (CV) is a significant complication following aneurysmal subarachnoid hemorrhage (aSAH), and lacks a comprehensive molecular understanding. Given the temporal trajectory of intracranial aneurysm (IA) formation, its rupture, and development of CV, altered gene expression might be a molecular substrate that runs through these clinical events, influencing both disease inception and progression. Utilizing RNA-Seq, we analyzed tissue samples from ruptured IAs with and without vasospasm to identify the dysregulated genes. In addition, temporal gene expression analysis was conducted. We identified seven dysregulated genes in patients with ruptured IA with vasospasm when compared with those without vasospasm. We found 192 common genes when the samples of each clinical subset of patients with IA, that is, unruptured aneurysm, ruptured aneurysm without vasospasm, and ruptured aneurysm with vasospasm, were compared with control samples. Among these common genes, TNFSF13B, PLAUR, OSM, and LAMB3 displayed temporal expression (progressive increase) with the pathological progression of disease that is formation of aneurysm, its rupture, and consequently the development of vasospasm. We validated the temporal gene expression pattern of OSM at both the transcript and protein levels and OSM emerges as a crucial gene implicated in the pathological progression of disease. In addition, RSAD2 and ATP1A2 appear to be pivotal genes for CV development. To the best of our knowledge, this is the first study to compare the transcriptome of aneurysmal tissue samples of aSAH patients with and without CV. The findings collectively provide new insights on the molecular basis of IA and CV and new leads for translational research.

Transcriptomic analysis reveals the crosstalk between type 2 diabetes and chronic pancreatitis.

Mounting evidence highlights a strong association between chronic pancreatitis (CP) and type 2 diabetes (T2D), although the exact mechanism of interaction remains unclear. This study aimed to investigate the crosstalk genes and pathogenesis between CP and T2D. Transcriptomic gene expression profiles of CP and T2D were extracted from Gene Expression Omnibus, respectively, and the common differentially expressed genes (DEGs) were subsequently identified. Further analysis, such as Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), protein-protein interaction, transcription factors (TFs), microRNA (miRNAs), and candidate chemicals identification, was performed to explore the possible common signatures between the two diseases. In total, we acquired 281 common DEGs by interacting CP and T2D datasets, and identified 10 hub genes using CytoHubba. GO and KEGG analyses revealed that endoplasmic reticulum stress and mitochondrial dysfunction were closely related to these common DEGs. Among the shared genes, EEF2, DLD, RAB5A, and SLC30A9 showed promising diagnostic value for both diseases based on receiver operating characteristic curve and precision-recall curves. Additionally, we identified 16 key TFs and 16 miRNAs that were strongly correlated with the hub genes, which may serve as new molecular targets for CP and T2D. Finally, candidate chemicals that might become potential drugs for treating CP and T2D were screened out. This study provides evidence that there are shared genes and pathological signatures between CP and T2D. The genes EEF2, DLD, RAB5A, and SLC30A9 have been identified as having the highest diagnostic efficiency and could be served as biomarkers for these diseases, providing new insights into precise diagnosis and treatment for CP and T2D.

De novo full length transcriptome analysis and gene expression profiling to identify genes involved in phenylethanol glycosides biosynthesis in Cistanche tubulosa

BackgroundThe dried stem of Cistanche, is a famous Chinese traditional medicine. The main active pharmacodynamic components are phenylethanol glycosides (PhGs). Cistanche tubulosa produces higher level of PhGs in its stems than that of Cistanche deserticola. However, the key genes in the PhGs biosynthesis pathway is not clear in C. tubulosa.ResultsIn this study, we performed the full-length transcriptome sequencing and gene expression profiling of C. tubulosa using PacBio combined with BGISEQ-500 RNA-seq technology. Totally, 237,772 unique transcripts were obtained, ranging from 199 bp to 31,857 bp. Among the unique transcripts, 188,135 (79.12%) transcripts were annotated. Interestingly, 1080 transcripts were annotated as 22 enzymes related to PhGs biosynthesis. We measured the content of echinacoside, acteoside and total PhGs at two development stages, and found that the content of PhGs was 46.74% of dry matter in young fleshy stem (YS1) and then decreased to 31.22% at the harvest stage (HS2). To compare with YS1, 13,631 genes were up-regulated, and 15,521 genes were down regulated in HS2. Many differentially expressed genes (DEGs) were identified to be involved in phenylpropanoid biosynthesis pathway, phenylalanine metabolism pathway, and tyrosine metabolism pathway.ConclusionsThis is the first report of transcriptome study of C. tubulosa which provided the foundation for understanding of PhGs biosynthesis. Based on these results, we proposed a potential model for PhGs biosynthesis in C. tubulosa.

ALL-061 Delineating the Genomic and Proteomic Landscape of BCR-ABL1-like Acute Lymphoblastic Leukemia Subtype: Bench-to-Bedside Translational Research

Baseline detection of Ph/BCR-ABL1-like-ALL cases is extremely beneficial for hematologists/physicians to design personalized medicine and to treat these patients efficiently. Worldwide, researchers use patented TaqMan low-density array (TLDA) gene expression profiling (GEP) to identify these cases. In developing countries, its detection is very challenging and expensive due to the unavailability of patented TLDA and there is a paucity of literature on the identification and characterization of Ph-like ALL cases. Stepwise identification followed by genomic and proteomic characterization of Ph-like ALL cases. This study was conducted at PGIMER, India (2016-2021). Flowcytometric-immunophenotyping and multiplex RT-PCR was performed to detect four recurrent fusion transcripts (n=1021). Transcriptome GEP was performed on 12 Ph-positive ALL cases. Ph-like-ALL cases identified using nCounter Platform (n=96) and RQ-PCR (n=536) were characterized using FISH, DD-PCR, RQ-PCR and MRD-positivity. Nine HKGs' expression was evaluated using 6 algorithms. Furthermore, we built and validated PHi-RACE classifier (n=629 cases) for its rapid detection. Lastly, we delineated the proteomic landscape of Ph-like ALL cases using LC-MS/MS. We studied 1021 treatment-naive and newly diagnosed B-ALL cases in this study cohort. Multiplex assay revealed 18.66% BCR-ABL1, 3.85% TCF3-PBX1, 5.27% ETV6-RUNX1, and 1.21% KMT2A-AFF1. Using transcriptome GEP, we identified 32.05% Ph-like-ALL cases using NanoString and 26.67% using RQ-PCR. These cases were substantially older and represented higher MRD positivity. In CRLF2 subgroup of Ph-like ALLs, we identified chimeric gene fusions including 10% JAK2 rearrangement, 8.33% CRLF2-IGH and CRLF2-P2RY8 translocation with concomitant R683S mutation. We found 8.33% IKZF1 (Δ4-7) deletion using DD-PCR. Furthermore, we developed an online logistic classifier entitled PHi-RACE (sensitivity=95.20%, specificity=83.70%, cut-off >0.28) for its detection using logistic regression. We have validated PHi-RACE classifier (n=93) and identified 56 Ph-like ALL cases. Six algorithms' findings revealed that EEF2, GAPDH and PGK1 represent the best HKGs for its detection. Lastly, from LC-MS/MS findings, we identified 3079 proteins, spliceosomal mediated mRNA splicing pathway and four miRNAs were significantly expressed in Ph-like ALL cases. The authors created and published the PHi-RACE classifier to detect Ph-like ALL cases, which has direct clinical implications. Further, we have delineated the genomic and proteomic landscape of Ph-like-ALL for the first time from developing countries.

Transcriptomic signatures responding to PKM2 activator TEPP-46 in the hyperglycemic human renal proximal epithelial tubular cells.

Pyruvate kinase M2 (PKM2), as the terminal and last rate-limiting enzyme of the glycolytic pathway, is an ideal enzyme for regulating metabolic phenotype. PKM2 tetramer activation has shown a protective role against diabetic kidney disease (DKD). However, the molecular mechanisms involved in diabetic tubular have not been investigated so far. In this study, we performed transcriptome gene expression profiling in human renal proximal tubular epithelial cell line (HK-2 cells) treated with 25 mM high D-glucose (HG) for 7 days before the addition of 10 μM TEPP-46, an activator of PKM2 tetramerization, for a further 1 day in the presence of HG. Afterwards, we analyzed the differentially expressed (DE) genes and investigated gene relationships based on weighted gene co-expression network analysis. The results showed that 2,902 DE genes were identified (adjusted P-value ≤ 0.05), where 2,509 DE genes (86.46%) were co-expressed in the key module. Four extremely downregulated DE genes (HSPA8, HSPA2, HSPA1B, and ARRB1) and three extremely upregulated DE genes (GADD45A, IGFBP3, and SIAH1) enriched in the downregulated endocytosis (hsa04144) and upregulated p53 signaling pathway (hsa04115), respectively, were validated by qRT-PCR experiments. The qRT-PCR results showed that the relative expression levels of HSPA8 [adjusted P-value = 4.45 × 10-34 and log2(FC) = -1.12], HSPA2 [adjusted P-value = 6.09 × 10-14 and log2(FC) = -1.27], HSPA1B [adjusted P-value = 1.14 × 10-11 and log2(FC) = -1.02], and ARRB1 [adjusted P-value = 2.60 × 10-5 and log2(FC) = -1.13] were significantly different (P-value < 0.05) from the case group to the control group. Furthermore, the interactions and predicted microRNAs of the key genes (HSPA8, HSPA2, HSPA1B, and ARRB1) were visualized in networks. This study identified the key candidate transcriptomic biomarkers and biological pathways in hyperglycemic HK-2 cells responding to the PKM2 activator TEPP-46 that can highlight a possibility of PKM2 tetramerization reshaping the interplay among endocytic trafficking through the versatile networks of Hsp70s and rewiring the crosstalk between EGFR signal transduction circuits and metabolic stress to promote resilience, which will be valuable for further research on PKM2 in DKD.

Transcriptome Analysis and Gene Expression Profiling of the Peanut Small Seed Mutant Identified Genes Involved in Seed Size Control.

Seed size is a key factor affecting crop yield and a major agronomic trait concerned in peanut (Arachis hypogaea L.) breeding. However, little is known about the regulation mechanism of peanut seed size. In the present study, a peanut small seed mutant1 (ssm1) was identified through irradiating peanut cultivar Luhua11 (LH11) using 60Coγ ray. Since the globular embryo stage, the embryo size of ssm1 was significantly smaller than that of LH11. The dry seed weight of ssm1 was only 39.69% of the wild type LH14. The seeds were wrinkled with darker seed coat. The oil content of ssm1 seeds were also decreased significantly. Seeds of ssm1 and LH11 were sampled 10, 20, and 40 days after pegging (DAP) and were used for RNA-seq. The results revealed that genes involved in plant hormones and several transcription factors related to seed development were differentially expressed at all three stages, especially at DAP10 and DAP20. Genes of fatty acid biosynthesis and late embryogenesis abundant protein were significantly decreased to compare with LH11. Interestingly, the gene profiling data suggested that PKp2 and/or LEC1 could be the key candidate genes leading to the small seed phenotype of the mutant. Our results provide valuable clues for further understanding the mechanisms underlying seed size control in peanut.

Ferroptosis is partially responsible for dexamethasone-induced T cell ablation, but not osteoporosis in larval zebrafish

Glucocorticoids (GCs) have been widely detected in the aquatic system. However, the hazardous effects of GCs on aquatic organisms were underestimated, and the mechanisms of GCs-induced toxic effects in fish were largely unknown. The zebrafish larvae at 3 dpf were exposed to dexamethasone (DEX) for 48 h, and the toxic effects and the underlying mechanisms were investigated in the current study. The T cells were ablated in zebrafish larvae after being treated with 1, 3, 10, 30 and 100 μM of DEX for 48 h. In addition, osteoporosis was induced and the regeneration of the caudal fin was inhibited, by 48 h-exposure to 10, 30 and 100 μM of DEX. The transcriptomic analysis, biochemical parameters and gene expression profiles revealed that ferroptosis possibly contributed to the DEX-induced toxic effects in zebrafish larvae. Finally, Fer-1 treatment partially attenuated the DEX-induced T cell ablation, but not osteoporosis in zebrafish larvae. Taken together, the current study proved the toxic effects of DEX on zebrafish larvae, and elucidated that ferroptosis was involved in DEX-induced toxicity, providing strong evidence for the toxic effects of GCs on aquatic organisms.

3-keto-DON, but Not 3-epi-DON, Retains the in Planta Toxicological Potential after the Enzymatic Biotransformation of Deoxynivalenol.

Deoxynivalenol (DON) is a secondary fungal metabolite that is associated with many adverse toxicological effects in agriculture as well as human/animal nutrition. Bioremediation efforts in recent years have led to the discovery of numerous bacterial isolates that can transform DON to less toxic derivatives. Both 3-keto-DON and 3-epi-DON were recently shown to exhibit reduced toxicity, compared to DON, when tested using different cell lines and mammalian models. In the current study, the toxicological assessment of 3-keto-DON and 3-epi-DON using in planta models surprisingly revealed that 3-keto-DON, but not 3-epi-DON, retained its toxicity to a large extent in both duckweeds (Lemna minor L.) and common wheat (Triticum aestivum L.) model systems. RNA-Seq analysis revealed that the exposure of L. minor to 3-keto-DON and DON resulted in substantial transcriptomic changes and similar gene expression profiles, whereas 3-epi-DON did not. These novel findings are pivotal for understanding the environmental burden of the above metabolites as well as informing the development of future transgenic plant applications. Collectively, they emphasize the fundamental need to assess both plant and animal models when evaluating metabolites/host interactions.

Transcriptome Analysis Reveal Candidate Genes and Pathways Responses to Lactate Dehydrogenase Inhibition (Oxamate) in Hyperglycemic Human Renal Proximal Epithelial Tubular Cells.

Diabetic kidney disease (DKD) is the leading cause of both chronic kidney disease (CKD) and end-stage renal disease (ESRD). Previous studies showed that oxamate could regulate glycemic homeostasis and impacted mitochondria respiration in a hyperglycemia-dependent manner in the rat proximal tubular cells. To explore the transcriptome gene expression profiling of kidney tissues in human renal proximal epithelial tubular cell line (HK-2), we treated HK-2 cells with high D-glucose (HG) for 7 days before the addition of 40 mM oxamate for a further 24 hours in the presence of HG in this study. Afterwards, we identified 3,884 differentially expressed (DE) genes based on adjusted P-value ≤ 0.05 and investigated gene relationships based on weighted gene co-expression network analysis (WGCNA). After qRT-PCR validations, MAP1LC3A, MAP1LC3B (P-value < 0.01) and BECN1 were found to show relatively higher expression levels in the treated groups than the control groups, while PGC1α (P-value < 0.05) showed the lower expressions. Accordingly, enrichment analyses of GO terms and KEGG pathways showed that several pathways [e.g., lysosome pathway (hsa04142) and p53 signaling pathway (hsa04115)] may be involved in the response of HK-2 cells to oxamate. Moreover, via WGCNA, we identified two modules: both the turquoise and blue modules were enriched in pathways associated with lysosome. However, the p53 signaling pathway was only found using all 3,884 DE genes. Furthermore, the key hub genes IGFBP3 (adjusted P-value = 1.34×10-75 and log2(FC) = 2.64) interacted with 6 up-regulated and 12 down-regulated DE genes in the network that were enriched in the p53 signaling pathway. This is the first study reporting co-expression patterns of a gene network after lactate dehydrogenase inhibition in HK-2 cells. Our results may contribute to our understanding of the underlying molecular mechanism of in vitro reprogramming under hyperglycemic stress that orchestrates the survival and functions of HK-2 cells.

Better Agreement of Human Transcriptomic and Proteomic Cancer Expression Data at the Molecular Pathway Activation Level.

Previously, we have shown that the aggregation of RNA-level gene expression profiles into quantitative molecular pathway activation metrics results in lesser batch effects and better agreement between different experimental platforms. Here, we investigate whether pathway level of data analysis provides any advantage when comparing transcriptomic and proteomic data. We compare the paired proteomic and transcriptomic gene expression and pathway activation profiles obtained for the same human cancer biosamples in The Cancer Genome Atlas (TCGA) and the NCI Clinical Proteomic Tumor Analysis Consortium (CPTAC) projects, for a total of 755 samples of glioblastoma, breast, liver, lung, ovarian, pancreatic, and uterine cancers. In a CPTAC assay, expression levels of 15,112 protein-coding genes were profiled using the Thermo QE series of mass spectrometers. In TCGA, RNA expression levels of the same genes were obtained using the Illumina HiSeq 4000 engine for the same biosamples. At the gene level, absolute gene expression values are compared, whereas pathway-grade comparisons are made between the pathway activation levels (PALs) calculated using average sample-normalized transcriptomic and proteomic profiles. We observed remarkably different average correlations between the primary RNA- and protein expression data for different cancer types: Spearman Rho between 0.017 (p = 1.7 × 10−13) and 0.27 (p < 2.2 × 10−16). However, at the pathway level we detected overall statistically significantly higher correlations: averaged Rho between 0.022 (p < 2.2 × 10−16) and 0.56 (p < 2.2 × 10−16). Thus, we conclude that data analysis at the PAL-level yields results of a greater similarity when comparing high-throughput RNA and protein expression profiles.

Weighted gene co-expression network analysis of embryos and first instar larvae of the horseshoe crab Tachypleus tridentatus uncovers development gene networks

Horseshoe crabs are marine chelicerates that have existed on Earth for about 450 million years, and they are often used as an experimental model for studying marine invertebrate embryology. In this study, we performed transcriptome gene expression profiling of four continuous embryonic stages (Stages 18–21) and first instar larvae of Tachypleus tridentatus. A mean of 50,742,995 high-quality clean reads was obtained from each library. We then conducted weighted gene co-expression network analysis (WGCNA) for 13,698 genes with fragments per kilobase of exon per million mapped fragments values >5. We identified 17 modules, six of which likely play critical roles in development. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of differentially expressed genes was performed on the biologically significant modules. We found that several pathways, such as hedgehog signaling pathway, VEGF signaling pathway, dorso-ventral axis formation, may be involved in the embryonic development process of T. tridentatus. We also identified hub genes that were highly connected in the six critical modules. This is the first study to apply WGCNA to horseshoe crabs to identify hub genes that may play critical roles in development, and our results provide new insight into the mechanisms underlying early development in horseshoe crabs.

Differential gene expression analysis related to sperm storage in spermathecas of Amphioctopus fangsiao

Sperm storage in the female body is an important strategy in animal reproductive behavior. Amphioctopus fangsiao is an economically important cephalopod that has a sperm storage period of up to seven months. There are few studies concerning the mechanism of sperm storage in A. fangsiao. In this study, we performed transcriptome gene expression profiling of the oviductal glands at different phases (presence and absence of sperm storage). In total, 7943 differentially expressed genes (DEGs) comprising 4737 upregulated and 3206 downregulated genes were identified. GO and KEGG enrichment analyses were used to search for sperm storage-related genes. A protein interaction network was constructed to examine the interactions between genes. Nineteen genes associated with immunity, apoptosis, and autophagy were obtained and verified by qRT-PCR. This is the first comprehensive analysis of sperm storage-related genes in A. fangsiao. The results provide basic insights into the complex sperm storage mechanism of A. fangsiao.

Inflammatory molecular endotypes of nasal polyps derived from White and Japanese populations

Emerging evidence suggests that chronic rhinosinusitis with nasal polyps (CRSwNP) is a highly heterogeneous disease with disparate inflammatory characteristics between different racial groups and geographies. Currently, little is known about possible underlying distinguishing factors between these inflammatory differences. Our aim was to interrogate differences in CRSwNP disease between White/non-Asian patients and Japanese patients by using whole transcriptome and single-cell RNA gene expression profiling of nasal polyps (NPs). We performed whole transcriptome RNA sequencing with endotype stratification of NPs from 8 White patients (residing in the United States) and 9 Japanese patients (residing in Japan). Reproducibility was confirmed by quantitative PCR in an independent validation set of 46 White and 31 Japanese patients. Single-cell RNA sequencing (scRNAseq) was used to stratify key cell types for contributory transcriptional signatures. Unsupervised clustering analysis identified 2 major endotypes that were present within both cohorts of patients with NPs and had previously been reported at the cytokine level: (1) type 2 endotype and (2) non-type 2 endotype. Importantly, there was a statistically significant difference in the proportion of these endotypes between these geographically distinct subgroups with NPs (P= .03). Droplet-based single-cell RNA sequencing further identified prominent type 2 inflammatory transcript expression: C-C motif chemokine ligand 13 (CCL13) and CCL18 in M2 macrophages, as well as cystatin SN (CST1) and CCL26 in basal, suprabasal, and secretory epithelial cells. NPs from both racial groups harbor the same 2 major endotypes, which we have determined to be present in differing ratios between each cohort with CRSwNP disease. Distinct inflammatory and epithelial cells contribute to the type 2 inflammatory profiles observed.

Genetics Responses to Hypoxia and Reoxygenation Stress in Larimichthys crocea Revealed via Transcriptome Analysis and Weighted Gene Co-Expression Network.

Simple SummaryHypoxia, which occurs frequently in aquaculture, can cause serious harm to all aspects of the growth, reproduction and metabolism of cultured fish. Due to the intolerance of Larimichthys crocea to hypoxia, Larimichthys crocea often floats head or even dies under hypoxic environment. However, the molecular mechanism of hypoxia tolerance in Larimichthys crocea has not been fully described. Therefore, the aim of this study was to explore the hub regulatory genes under hypoxic stress environment by transcriptome analysis of three key tissues (liver, blood and gill) in Larimichthys crocea. We identified a number of important genes that exercise different regulatory functions. Overall, this study will provide important clues to the molecular mechanisms of hypoxia tolerance in Larimichthys crocea.The large yellow croaker (Larimichthys crocea) is an important marine economic fish in China; however, its intolerance to hypoxia causes widespread mortality. To understand the molecular mechanisms underlying hypoxia tolerance in L. crocea, the transcriptome gene expression profiling of three different tissues (blood, gills, and liver) of L. crocea exposed to hypoxia and reoxygenation stress were performed. In parallel, the gene relationships were investigated based on weighted gene co-expression network analysis (WGCNA). Accordingly, the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed that several pathways (e.g., energy metabolism, signal transduction, oxygen transport, and osmotic regulation) may be involved in the response of L. crocea to hypoxia and reoxygenation stress. In addition, also, four key modules (darkorange, magenta, saddlebrown, and darkolivegreen) that were highly relevant to the samples were identified by WGCNA. Furthermore, some hub genes within the association module, including RPS16, EDRF1, KCNK5, SNAT2, PFKL, GSK-3β, and PIK3CD, were found. This is the first study to report the co-expression patterns of a gene network after hypoxia stress in marine fish. The results provide new clues for further research on the molecular mechanisms underlying hypoxia tolerance in L. crocea.

New insights of low-temperature plasma effects on seeds germination of Platycodon grandiflorum

Abstract Low temperature plasma (LTP) exerted beneficial effects on seed germination and crop growth. With application of LTP on seed germination of Platycodon grandiflorum, germination indexes and physiological parameters and transcriptome profile were investigated in this experiment. The results showed the seed germination rate and germinating potential of Platycodon grandiflorum in LTP increased by 24.7% and 30.7% comparing to CK respectively (p &lt; 0.05). The SOD and CAT activity of the seed sprouts increased by 26.78% and 12.41% comparing to CK. Transcriptome results showed that 14342 up-regulated and 4366 down-regulated different expressing genes (DEGs) at LTP and CK. A considerable number of DEGs related to the plant hormone signal transduction, peroxisome, oxidative phosphorylation, starch and surose in response to low temperature plasma were identified in this study. The transcriptomic gene expression profiles present a valuable genomic tool to the molecular mechanisms of Platycodon grandiflorum seed germination underlying low temperature plasma. which can provide theoretical basis and practical guidance for cultivation and planting of Platycodon grandiflorum.

English

Sicyos angulatus has become an important invasive plant exhibiting good ecological adaptability and strong competitive ability. However, studies on this plant at the molecular level are limited by a lack of sequencing data. The present study obtained transcriptome sequences and gene expression profiles using RNA-Seq during S. angulatus seed germination. In total, RNA-Seq generated 491,967,468 reads, which were de novo assembled and 127,874 unigenes with N50 length of 807 bp. About 34.9% of the unigenes (44,660) were annotated against the protein databases, and 35,176 coding sequences were determined. During S. angulatus seed germination, over 127,860 unigenes were expressed and 66,664 unigenes differentially expressed genes (DEGs), among which 8919 DEGs were similar in pairwise comparison. Gene Ontology (GO) analysis of DEGs revealed that genes related to post-embryonic development, meristem development, and photosynthesis were enriched. In addition, the GO term “plant hormone signal transduction pathway” was also enriched in the DEGs. Important changes in genes expression related to auxin and gibberellin signal transduction might possibly be associated with S. angulatus seed germination. The findings of this study provide a foundation for research on S. angulatus that may contribute to prevent further invasion of this plant, consequently protecting the environment. © 2021 Friends Science Publishers

Abstract PD5-08: The human tumor atlas network (HTAN) breast pre cancer atlas: A multi-omic integrative analysis of ductal carcinoma in situ (DCIS) and correlation with clinical outcomes

Abstract Introduction. As nonobligate precursors of invasive disease, pre-cancers provide a unique vantage point from which to study the molecular pathways and evolutionary dynamics that lead to the development of life-threatening cancers. Ductal carcinoma in situ (DCIS) is the most commonly diagnosed precursor of breast cancer, with variable propensity for invasive progression. In order to address the problems of over- and under-treatment, we performed a multimodal, integrated profile of DCIS with clinical outcomes with which to develop and validate predictors of invasive progression. Methods. We present observations on DNA, RNA, and protein expression on two independent patient cohorts of DCIS, diagnosed from 1981 to 2014, from the Translational Breast Cancer Research Consortium (TBCRC 038) and the Washington University Repository of Archival Human Breast Tissue (RAHBT). Patients initially diagnosed with DCIS, with either DCIS or invasive recurrence (cases; mean follow up 5.8 years) were matched to those without recurrence (controls; mean follow up 10.3 years), based upon age at diagnosis and year of diagnosis. Results. We present genomic and cellular changes that correlate with both disease states and patient outcomes in DCIS. DCIS can be clustered by classification systems developed for IBC. Specific immune cell types and pathways correlate with longitudinal outcome. Luminal cell adhesion and metabolism pathways are upregulated in controls and cases, respectively. Highly multiplexed ion beam imaging (MIBI) was used to validate RNA seq findings, and to provide single cell-level spatial context for molecular alterations.Conclusion. We have performed an integrated multi-omic analysis of DCIS and associated tumor micorenvironment. Our multi-scale approach employs in situ methods to generate a spatially resolved atlas of breast precancers where different modalities can be directly compared to each other, and correlated with conventional pathology findings and clinical outcome. The PreCancer Atlas represents a complex multi-modal database for DCIS study, whose design allows for future discovery and hypothesis generation. Table 1. Breast Pre-cancer Atlas Multi-scale Characterization AssaysAssayScaleType of DataIntegration and validation with other assaysRNA-seq (Single duct, single cell, TME)Cell, duct, organ, normal tissue1. Whole transcriptome gene expression profiling per single duct (also enabling CNV and cell type prediction)2. Whole transcriptome gene expression profiling per single duct1. Prediction of CNV confirmed by DNA-seq (single duct) and FISH (single cell)2. Prediction of cell type composition (Cibersort) confirmed by multiplex IHC and multicolor flow cytometryLow-pass whole genome DNA-seqDuct and adjacent normalCNV profiling per single ductAnalysis of CNV supported by RNA-seq (single duct) and MIBI (single cell)Whole genome sequencingDuct and adjacent normalMutation status per single ductMutational analysis confirmed by RNA-seqMultiplex IHC (MIBI &amp; Cyclic multicolor)Cell1. Cell type2. Proteomic analysisAnalysis of cell type supported by RNA-seq of ducts (Cibersort) and single cellsH&amp;E MorphometricsCell, duct, organSpatial location of cell types, organization of ductsAnalysis of H&amp;E images correlated with FISH data Citation Format: Shelley Hwang, Siri H Strand, Belen Rivero, Lorraine King, Tyler Risom, Bryan Harmon, Fergus Couch, Kristalyn Gallagher, Mark Kilgore, Shi Wei, Angela DeMichele, Tari King, Priscilla McAuliffe, Julie Nangia, Ana Maria Storniolo, Alastair Thompson, Gaorav Gupta, Joanna Lee, Jennifer Tseng, Robyn Burns, ChunFang Zhu, Magda Matusiak, Shirley X Zhu, Jason Wang, Jose Seoane, Jen Tappenden, Daisy Ding, Dadong Zhang, Jingqin Luo, Sujay Vennam, Sushama Varma, Lunden Simpson, Luis Cisneros, Timmothy Hardman, Lauren Anderson, Cody Straub, Sucheta Srivastava, Deb J Veis, Christina Curtis, Rob Tibshirani, Robert Michael Angelo, Allison Hall, Kouros Owzar, Kornelia Polyak, Carlo Maley, Jeff Marks, Graham Colditz, Robert B West. The human tumor atlas network (HTAN) breast pre cancer atlas: A multi-omic integrative analysis of ductal carcinoma in situ (DCIS) and correlation with clinical outcomes [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PD5-08.

Transcriptome sequencing and gene expression profiling of Pinus sibirica under different cold stresses.

Cold stress is a major abiotic factor that affects plant growth and geographical distribution. Pinus sibirica is extremely frigostable tree species. To understand the molecular mechanisms of cold tolerance by P. sibirica, physiological responses were analyzed and transcriptome profiling was conducted to the plants treated by cold stress. The physiological data showed that membrane permeability relative conductivity (REC), reactive oxygen species (ROS), malonaldehyde (MDA) content, peroxidase (POD) and catalase (CAT) activity, soluble sugar, soluble protein and proline contents were increased significantly (p < 0.05) in response to cold stress. Transcriptome analysis identified a total of 871, 1397 and 872 differentially expressed genes (DEGs) after cold treatment for 6 h, 24 h and 48 h at –20°C, respectively. The signaling pathway mediated by Ca2+ as a signaling molecule and abscisic acid pathways were the main cold signal transduction pathways in P. sibirica. The APETALA2/Ethylene-Responsive Factor (AP2/ERF) and MYB transcription factor families also play an important role in the transcriptional regulation of P. sibirica. In addition, many genes related to photosynthesis were differentially expressed under cold stress. We also validated the reliability of transcriptome data with quantitative real-time PCR. This study lays the foundation for understanding the molecular mechanisms related to cold responses in P. sibirica.

Transcriptome Analysis of Female and Male Henosepilachna vigintioctopunctata (Coleoptera: Coccinellidae) Identifies Sex-related Genes

Henosepilachna vigintioctopunctata is an emerging horticultural pest of Solanaceae plants in Asia. Here, we employed transcriptome sequencing of three female and three male H. vigintioctopunctata libraries to identify sex-related genes. We recorded 281 599 294 bp of clean reads, and de novo assembly generated 85 206 unigenes, with N50 of 1 311 bp and average length of 763 bp. We found 5 002 genes highly expressed in males and 2 179 genes highly expressed in females. Expression profiles of six unigenes specific to females, six unigenes specific to males, and ten unigenes common to both sexes were verified using semi-quantitive RT-PCR; results showed expression levels of these 22 unigenes were relatively consistent with the transcriptome analysis. Using all unigenes as references, we identified 6 657 putative simple sequence repeats. The transcriptome sequences and gene expression profiles of female and male H. vigintioctopunctata help explore genetic mechanisms of sexual dimorphism.

Transcriptome sequencing and gene expression profiling of Populus wutunensis, a natural hybrid, during salinity stress

When Populus wutunensis seedlings are salt stressed, many genes involved in cellular metabolism, signal transduction, transport, osmotic regulation, and reactive oxygen species scavenging show elevated expression levels. Populus wutunensis, a natural hybrid individual resulting from a cross between Populus × xiaozhuanica W. Y. Hsu et Liang, shows salt and drought tolerance and is fast-growing. It is widely cultivated in the Three-North Shelterbelt Forest area in China. To determine the transcriptome changes in P. wutunensis under salt stress, RNA-seq was conducted in salt-treated leaves and non-salt-treated control leaves. A total of 81,188 unigenes, with an average length of 982 bp, were obtained. In salt-treated leaves, 2466 differentially expressed genes were identified that differed from those in control leaves at any time point. Among them, 391 were upregulated and 44 were downregulated during all salt stress time points. They were mainly involved in cellular and metabolic processes, signal transduction, transport, osmotic regulation, and reactive oxygen species (ROS) scavenging. In addition, 2543 transcription factor genes were identified and divided into 60 families. qRT-PCR analysis revealed that the levels of differentially expressed genes were consistent with those from sequencing of the transcriptome under salt stress. These findings will increase our understanding of gene regulation in response to salt stress in P. wutunensis and are beneficial for future research on molecular-assisted breeding.