TY - JOUR
T1 - The complex transcriptional landscape of the anucleate human platelet.
AU - Bray, Paul F.
AU - McKenzie, Steven E.
AU - Edelstein, Leonard C.
AU - Nagalla, Srikanth
AU - Delgrosso, Kathleen
AU - Ertel, Adam
AU - Kupper, Joan
AU - Jing, Yi
AU - Londin, Eric
AU - Loher, Phillipe
AU - Chen, Huang Wen
AU - Fortina, Paolo
AU - Rigoutsos, Isidore
N1 - Funding Information:
We wish to thank Ms. Lin Ma and Dr. Xianguo Kong for excellent technical assistance. This work was supported in part by the Cardeza Foundation for Hematological Research (Philadelphia, PA; PFB. LCE, SEM), the Kimmel Cancer Center (Philadelphia, PA; PF, KD, JK and AE), and by grants from the NIH-NCI Cancer Center (P30-CA-56036 to PF, KD and AE), NIH-NHLBI (R01-HL-102482 to PB), NIH-NIAID (2U19AI056363-06/2030984 to IR), the William M. Keck Foundation (IR) and TJU funds (IR, YJ, EL, PL, H-WC). The authors declare no conflicts of interest.
PY - 2013
Y1 - 2013
N2 - Human blood platelets are essential to maintaining normal hemostasis, and platelet dysfunction often causes bleeding or thrombosis. Estimates of genome-wide platelet RNA expression using microarrays have provided insights to the platelet transcriptome but were limited by the number of known transcripts. The goal of this effort was to deep-sequence RNA from leukocyte-depleted platelets to capture the complex profile of all expressed transcripts. From each of four healthy individuals we generated long RNA (≥40 nucleotides) profiles from total and ribosomal-RNA depleted RNA preparations, as well as short RNA (<40 nucleotides) profiles. Analysis of ~1 billion reads revealed that coding and non-coding platelet transcripts span a very wide dynamic range (≥16 PCR cycles beyond β-actin), a result we validated through qRT-PCR on many dozens of platelet messenger RNAs. Surprisingly, ribosomal-RNA depletion significantly and adversely affected estimates of the relative abundance of transcripts. Of the known protein-coding loci, ~9,500 are present in human platelets. We observed a strong correlation between mRNAs identified by RNA-seq and microarray for well-expressed mRNAs, but RNASeq identified many more transcripts of lower abundance and permitted discovery of novel transcripts. Our analyses revealed diverse classes of non-coding RNAs, including: pervasive antisense transcripts to protein-coding loci; numerous, previously unreported and abundant microRNAs; retrotransposons; and thousands of novel un-annotated long and short intronic transcripts, an intriguing finding considering the anucleate nature of platelets. The data are available through a local mirror of the UCSC genome browser and can be accessed at: http://cm.jefferson.edu/platelets_2012/.
AB - Human blood platelets are essential to maintaining normal hemostasis, and platelet dysfunction often causes bleeding or thrombosis. Estimates of genome-wide platelet RNA expression using microarrays have provided insights to the platelet transcriptome but were limited by the number of known transcripts. The goal of this effort was to deep-sequence RNA from leukocyte-depleted platelets to capture the complex profile of all expressed transcripts. From each of four healthy individuals we generated long RNA (≥40 nucleotides) profiles from total and ribosomal-RNA depleted RNA preparations, as well as short RNA (<40 nucleotides) profiles. Analysis of ~1 billion reads revealed that coding and non-coding platelet transcripts span a very wide dynamic range (≥16 PCR cycles beyond β-actin), a result we validated through qRT-PCR on many dozens of platelet messenger RNAs. Surprisingly, ribosomal-RNA depletion significantly and adversely affected estimates of the relative abundance of transcripts. Of the known protein-coding loci, ~9,500 are present in human platelets. We observed a strong correlation between mRNAs identified by RNA-seq and microarray for well-expressed mRNAs, but RNASeq identified many more transcripts of lower abundance and permitted discovery of novel transcripts. Our analyses revealed diverse classes of non-coding RNAs, including: pervasive antisense transcripts to protein-coding loci; numerous, previously unreported and abundant microRNAs; retrotransposons; and thousands of novel un-annotated long and short intronic transcripts, an intriguing finding considering the anucleate nature of platelets. The data are available through a local mirror of the UCSC genome browser and can be accessed at: http://cm.jefferson.edu/platelets_2012/.
UR - http://www.scopus.com/inward/record.url?scp=84872183938&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84872183938&partnerID=8YFLogxK
U2 - 10.1186/1471-2164-14-1
DO - 10.1186/1471-2164-14-1
M3 - Article
C2 - 23323973
AN - SCOPUS:84872183938
VL - 14
JO - Seminars in Fetal and Neonatal Medicine
JF - Seminars in Fetal and Neonatal Medicine
SN - 1744-165X
ER -