Evidence for the biogenesis of more than 1,000 novel human microRNAs.
BACKGROUND
MicroRNAs (miRNAs) are established regulators of development, cell identity and disease. Although nearly two thousand human miRNA genes are known and new ones are continuously discovered, no attempt has been made to gauge the total miRNA content of the human genome.


RESULTS
Employing an innovative computational method on massively pooled small RNA sequencing data, we report 2,469 novel human miRNA candidates of which 1,098 are validated by in-house and published experiments. Almost 300 candidates are robustly expressed in a neuronal cell system and are regulated during differentiation or when biogenesis factors Dicer, Drosha, DGCR8 or Ago2 are silenced. To improve expression profiling, we devised a quantitative miRNA capture system. In a kidney cell system, 400 candidates interact with DGCR8 at transcript positions that suggest miRNA hairpin recognition, and 1,000 of the new miRNA candidates interact with Ago1 or Ago2, indicating that they are directly bound by miRNA effector proteins. From kidney cell CLASH experiments, in which miRNA-target pairs are ligated and sequenced, we observe hundreds of interactions between novel miRNAs and mRNA targets. The novel miRNA candidates are specifically but lowly expressed, raising the possibility that not all may be functional. Interestingly, the majority are evolutionarily young and overrepresented in the human brain.


CONCLUSIONS
In summary, we present evidence that the complement of human miRNA genes is substantially larger than anticipated, and that more are likely to be discovered in the future as more tissues and experimental conditions are sequenced to greater depth.
Marc R
MR
Friedländer
Esther
E
Lizano
Anna J S
AJ
Houben
Daniela
D
Bezdan
Mónica
M
Báñez-Coronel
Grzegorz
G
Kudla
Elisabet
E
Mateu-Huertas
Birgit
B
Kagerbauer
Justo
J
González
Kevin C
KC
Chen
Emily M
EM
LeProust
Eulàlia
E
Martí
Xavier
X
Estivill
D060565:Argonaute Proteins / Q000378:metabolism; D039642:Eukaryotic Initiation Factors / Q000378:metabolism; D015894:Genome, Human; D006801:Humans; D061986:MCF-7 Cells; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D016601:RNA-Binding Proteins / Q000378:metabolism
C115848:AGO1 protein, human; C466422:AGO2 protein, human; D060565:Argonaute Proteins; C474043:DGCR8 protein, human; D039642:Eukaryotic Initiation Factors; D035683:MicroRNAs; D016601:RNA-Binding Proteins