Abstract
Piwi- interacting RNAs (piRNAs), a new class of small RNAs discovered from mammalian testes, are involved in transcriptional silencing of retrotransposons and other genetic elements in germ line cells. In order to identify a full transcriptome set of piRNAs expressed in the sexually mature porcine testes, small RNA fractions were extracted and were subjected to a Solexa deep sequencing. We cloned 6,913,561 clean reads of Sus Scrofa small RNAs (18–30 nt) and performed functional characterization. Sus Scrofa small RNAs showed a bimodal length distribution with two peaks at 21 nt and 29 nt. Then from 938,328 deep-sequenced small RNAs (26–30 nt), 375,195 piRNAs were identified by a k-mer scheme and 326 piRNAs were identified by homology searches. All piRNAs predicted by the k-mer scheme were then mapped to swine genome by Short Oligonucleotide Analysis Package (SOAP), and 81.61% of all uniquely mapping piRNAs (197,673) were located to 1124 defined genomic regions (5.85 Mb). Within these regions, 536 and 501 piRNA clusters generally distributed across only minus or plus genomic strand, 48 piRNA clusters distributed on two strands but in a divergent manner, and 39 piRNA clusters distributed on two strands in an overlapping manner. Furthermore, expression pattern of 7 piRNAs identified by homology searches showed 5 piRNAs displayed a ubiquitous expression pattern, although 2 piRNAs were specifically expressed in the testes. Overall, our results provide new information of porcine piRNAs and their specific expression pattern in porcine testes suggests that piRNAs have a role in regulating spermatogenesis.
Highlights
Spermatogenesis is a complex process of cellular divisions and developmental changes that occur within the seminiferous tubules of the testes [1]
Of high-quality reads in this library, 6,913,561 (91.30%) clean reads of Sus Scrofa small RNA fraction (18–30 nt) were used to map the swine genome assembly using the Short Oligonucleotide Analysis Package (SOAP), leading to 4,527,258 genome-matched reads (Table S1)
PiRNA candidates by a k-mer scheme and 326 PIWI-interacting RNAs (piRNAs) candidates by homology searches from 938,328 small RNAs sequenced by generation method (26–30 nt)
Summary
Spermatogenesis is a complex process of cellular divisions and developmental changes that occur within the seminiferous tubules of the testes [1]. Spermatogenesis can be divided into three major phases: mitotic process of stem cells (spermatogonia) to form spermatocytes, meiosis to reduce the number of chromosomes to form spermatids, and spermiogenesis in which haploid spermatids develop into spermatozoa [1,2]. Meiosis and haploid stages are unique to germ cells, and often require specific genes to execute unique regulatory roles [3]. The patterns of gene expression in meiotic and haploid germ cells could be repressed by post-transcriptional control [3]. This repression is partly achieved by some regulatory elements through binding mRNA un-translated regions [3,4]. Some regulatory small RNAs including small interfering RNAs (siRNAs), PIWI-interacting RNAs (piRNAs) and microRNAs (miRNAs) have emerged as important regulators of eukaryotic gene expression and have been used in elucidating the molecular mechanisms regulating spermatogenesis [5,6]
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