MicroRNA discovery and analysis of pinewood nematode Bursaphelenchus xylophilus by deep sequencing.
BACKGROUND
MicroRNAs (miRNAs) are considered to be very important in regulating the growth, development, behavior and stress response in animals and plants in post-transcriptional gene regulation. Pinewood nematode, Bursaphelenchus xylophilus, is an important invasive plant parasitic nematode in Asia. To have a comprehensive knowledge about miRNAs of the nematode is necessary for further in-depth study on roles of miRNAs in the ecological adaptation of the invasive species.


RESULTS
Five small RNA libraries were constructed and sequenced by Illumina/Solexa deep-sequencing technology. A total of 810 miRNA candidates (49 conserved and 761 novel) were predicted by a computational pipeline, of which 57 miRNAs (20 conserved and 37 novel) encoded by 53 miRNA precursors were identified by experimental methods. Ten novel miRNAs were considered to be species-specific miRNAs of B. xylophilus. Comparison of expression profiles of miRNAs in the five small RNA libraries showed that many miRNAs exhibited obviously different expression levels in the third-stage dispersal juvenile and at a cold-stressed status. Most of the miRNAs exhibited obviously down-regulated expression in the dispersal stage. But differences among the three geographic libraries were not prominent. A total of 979 genes were predicted to be targets of these authentic miRNAs. Among them, seven heat shock protein genes were targeted by 14 miRNAs, and six FMRFamide-like neuropeptides genes were targeted by 17 miRNAs. A real-time quantitative polymerase chain reaction was used to quantify the mRNA expression levels of target genes.


CONCLUSIONS
Basing on the fact that a negative correlation existed between the expression profiles of miRNAs and the mRNA expression profiles of their target genes (hsp, flp) by comparing those of the nematodes at a cold stressed status and a normal status, we suggested that miRNAs might participate in ecological adaptation and behavior regulation of the nematode. This is the first description of miRNAs in plant parasitic nematodes. The results provide a useful resource for further in-depth study on molecular regulation and evolution of miRNAs in plant parasitic nematodes.
College of Life Sciences, Beijing Normal University, Beijing, China.
Qi-Xing
QX
Huang
Xin-Yue
XY
Cheng
Zhen-Chuan
ZC
Mao
Yun-Sheng
YS
Wang
Li-Lin
LL
Zhao
Xia
X
Yan
Virginia R
VR
Ferris
Ru-Mei
RM
Xu
Bing-Yan
BY
Xie
D000465:Algorithms; D000818:Animals; D001483:Base Sequence; D020869:Gene Expression Profiling; D035683:MicroRNAs / Q000235:genetics*; D008969:Molecular Sequence Data; D009348:Nematoda / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid
D035683:MicroRNAs