SUPPLEMENTARY DATA to: Next Generation Sequencing Analysis of Human Platelet PolyA+ mRNAs and rRNA-Depleted Total RNA (original) (raw)
Transcriptomic landscape of blood platelets in healthy donors
Scientific Reports
Blood platelet RNA-sequencing is increasingly used among the scientific community. Aberrant platelet transcriptome is common in cancer or cardiovascular disease, but reference data on platelet RNA content in healthy individuals are scarce and merit complex investigation. We sought to explore the dynamics of platelet transcriptome. Datasets from 204 healthy donors were used for the analysis of splice variants, particularly with regard to age, sex, blood storage time, unit of collection or library size. Genes B2M, PPBP, TMSB4X, ACTB, FTL, CLU, PF4, F13A1, GNAS, SPARC, PTMA, TAGLN2, OAZ1 and OST4 demonstrated the highest expression in the analysed cohort, remaining substantial transcription consistency. CSF3R gene was found upregulated in males (fold change 2.10, FDR q
Blood, 2020
Genome-wide association studies have identified common variants associated with platelet-related phenotypes, but because these variants are largely intronic or intergenic, their link to platelet biology is unclear. In 290 normal subjects from the GeneSTAR Research Study (110 African Americans [AAs] and 180 European Americans [EAs]), we generated whole-genome sequence data from whole blood and RNA sequence data from extracted nonribosomal RNA from 185 induced pluripotent stem cell-derived megakaryocyte (MK) cell lines (platelet precursor cells) and 290 blood platelet samples from these subjects. Using eigenMT software to select the peak single-nucleotide polymorphism (SNP) for each expressed gene, and meta-analyzing the results of AAs and EAs, we identify (q-value < 0.05) 946 cis-expression quantitative trait loci (eQTLs) in derived MKs and 1830 cis-eQTLs in blood platelets. Among the 57 eQTLs shared between the 2 tissues, the estimated directions of effect are very consistent (98...
Circular RNA enrichment in platelets is a signature of transcriptome degradation
Blood, 2015
In platelets, splicing and translation occur in the absence of a nucleus. However, the integrity and stability of mRNAs derived from megakaryocyte progenitor cells remain poorly quantified on a transcriptome-wide level. As circular RNAs (circRNAs) are resistant to degradation by exonucleases, their abundance relative to linear RNAs can be used as a surrogate marker for mRNA stability in the absence of transcription. Here we show that circRNAs are enriched in human platelets 17-188 fold relative to nucleated tissues, and 14-26 fold relative to samples digested with RNAse R to selectively remove linear RNA. We compare RNAseq read depths inside and outside circRNAs to provide in silico evidence of transcript circularity, show that exons within circRNAs are enriched on average 12.7X in platelets relative to nucleated tissues, and identify 3162 genes significantly enriched for circRNAs, including some where all RNAseq reads appear to be derived from circular molecules. We also confirm th...
Molecular & Cellular Proteomics, 2003
Platelets, while anucleate, contain RNA, some of which is translated into protein upon activation. Hypothesising that the platelet proteome is reflected in the transcriptome, we identified 82 proteins secreted from activated platelets and compared these, as well as published proteomic data, to the transcriptional profile. We also compared the transcriptome of platelets to other tissues to identify platelet-specific genes and used ontology to determine gene categories over-represented in platelets. RNA was isolated from highly pure platelet preparations for hybridization to Affymetrix oligonucleotide arrays. We identified 2,928 distinct messages as being present in platelets. The platelet transcriptome was compared with the proteome by relating both to UniGene clusters. Platelet proteomic data correlated well with the transcriptome, with 69% of secreted proteins detectable at the mRNA level, and similar concordance was obtained using two published datasets. While many of the most abundant mRNAs are for known platelet proteins, messages were detected for proteins not previously reported in platelets. Some of these may represent residual megakaryocyte messages; however, proteomic analysis confirmed the expression of many previously unreported genes in platelets. Transcripts for well-described platelet proteins are among the most platelet-specific messages. Ontological categories related to signal transduction, receptors, ion channels, and membranes are over-represented in platelets, while categories involved in protein synthesis are depleted. Despite the absence of gene transcription, the platelet proteome is mirrored in the transcriptome. Conversely, transcriptional analysis predicts the presence of novel proteins in the platelet. Transcriptional analysis is relevant to platelet biology, providing insights into platelet function and the mechanisms of platelet disorders.
Transcriptomic profiling of platelet senescence and platelet extracellular vesicles
Transfusion, 2016
Platelets (PLTs) are derived from megakaryocytes during PLT shedding. Senescent or activated PLTs are expanded in vascular and neurological diseases and release PLT extracellular vesicles (PL-EVs). A systematic analysis of regular messenger RNA (mRNA) and small RNA composition in PLTs and PL-EVs during in vitro PLT senescence has not yet been published. We isolated PLTs, total PL-EVs, and PL-EV subsets on Days 0 and 5 from human stored donor platelet concentrates. Isolated mRNA species and microRNA (miRNA) species were analyzed by microarrays and deep sequencing. Correlation of mRNA and miRNA species (miR) and miRNA target analyses were performed using bioinformatics. During in vitro PLT senescence, residual PLT mRNA species were decreased and partially converted to miRNA species. Residual mRNAs included encoded genes relevant for atherosclerosis, inflammation (matrix metallopeptidase 14 [MMP-14], granulin [GRN], angiopoietin like 2 [ANGPTL2]), and neurotransmission (dopamine recept...
Blood, 2012
Antiplatelet treatment is of fundamental importance in combatting functions/dysfunction of platelets in the pathogenesis of cardiovascular and inflammatory diseases. Dysfunction of anucleate platelets is likely to be completely attributable to alterations in posttranslational modifications and protein expression. We therefore examined the proteome of platelets highly purified from fresh blood donations, using elaborate protocols to ensure negligible contamination by leukocytes, erythrocytes, and plasma. Using quantitative mass spectrometry, we created the first comprehensive and quantitative human platelet proteome, comprising almost 4000 unique proteins, estimated copy numbers for ∼ 3700 of those, and assessed intersubject (4 donors) as well as intrasubject (3 different blood samples from 1 donor) variations of the proteome. For the first time, our data allow for a systematic and weighted appraisal of protein networks and pathways in human platelets, and indicate the feasibility of...
Platelet MicroRNAs: An Overview
Transfusion Medicine Reviews, 2015
MicroRNAs are short ~ 22 nucleotide non-coding RNA that have been found to influence the expression of many genes and cellular processes by either repressing translation or degrading mRNA transcripts. Platelet miRNA expression has been shown to be perturbed during ex vivo storage of platelets and in platelet associated disorders. While bioinformatics-based miRNA target predictions have been established, direct experimental validation of the role of miRNAs in platelet biology has been rather slow. Target prediction studies are nonetheless valuable in directing the design of appropriate experiments to test specific miRNA:mRNA interactions relevant to the underlying mechanisms of platelet function in general, and in disease as well as in ex vivo storage-associated 'storage lesions' (SL), a collective term used to include physiological, biochemical and morphological changes that occur in stored platelets. This brief review will focus on emerging human platelet miRNA studies to emphasize their potential role relevant to transfusion medicine field in terms of regulating platelet signaling pathways, markers of platelet associated disorders, and remote impactors of gene expression (intercellular bio-modulators) as well as potential platelet quality markers of storage and pathogen reduction treatments.