Assessment of piRNA biogenesis and function in testicular germ cell tumors and their precursor germ cell neoplasia in situ

Ildar V Gainetdinov,Yulia V Skvortsova,Tatyana L Azhikina,Alexey Klimov,Alexey A Tryakin,Sofia A Kondratieva

doi:10.1186/s12885-017-3945-6

Ildar V Gainetdinov, Yulia V Skvortsova + Show 4 more

Open Access

https://doi.org/10.1186/s12885-017-3945-6

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Abstract

BackgroundAberrant overexpression of PIWI/piRNA pathway proteins is shown for many types of tumors. Interestingly, these proteins are downregulated in testicular germ cell tumors (TGCTs) compared to normal testis tissues. Here, we used germline and TGCT markers to assess the piRNA biogenesis and function in TGCTs and their precursor germ cell neoplasia in situ (GCNIS).MethodsWe used small RNA deep sequencing, qRT-PCR, and mining public RNAseq/small RNA-seq datasets to examine PIWI/piRNA gene expression and piRNA biogenesis at four stages of TGCT development: (i) germ cells in healthy testis tissues, (ii) germ cells in testis tissues adjacent to TGCTs, (iii) GCNIS cells and (iv) TGCT cells. To this end, we studied three types of samples: (a) healthy testis, (b) testis tissues adjacent to two types of TGCTs (seminomas and nonseminomas) and containing both germ cells and GCNIS cells, as well as (c) matching TGCT samples.ResultsBased on our analyses of small RNA-seq data as well as the presence/absence of expression correlation between PIWI/piRNA pathway genes and germline or TGCT markers, we can suggest that piRNA biogenesis is intact in germ cells present in healthy adult testes, and adjacent to TGCTs. Conversely, GCNIS and TGCT cells were found to lack PIWI/piRNA pathway gene expression and germline-like piRNA biogenesis. However, using an in vitro cell line model, we revealed a possible role for a short PIWIL2/HILI isoform expressed in TGCTs in posttranscriptional regulation of the youngest members of LINE and SINE classes of transposable elements. Importantly, this regulation is also implemented without involvement of germline-like biogenesis of piRNAs.ConclusionsThough further studies are warranted, these findings suggest that the conventional germline-like PIWI/piRNA pathway is lost in transition from germ cells to GCNIS cells.

Highlights

Aberrant overexpression of PIWI/PIWI-interacting RNAs (piRNAs) pathway proteins is shown for many types of tumors
Expression of piRNA biogenesis genes in germ cells in testis samples adjacent to Testicular germ cell tumor (TGCT) To study the role of PIWI/piRNA pathway in development of TGCTs, we wanted to assess its function at four stages of neoplastic transformation using three types of samples: (i) normal germ cells, (ii) germ cells and (iii) germ cell neoplasia in situ (GCNIS) cells adjacent to TGCTs and (iv) TGCT cells
Because testis samples adjacent to TGCTs contain both germ cells and GCNIS cells, we distinguished between those using correlation of piRNA pathway gene expression with either germline (DDX4 and DAZL [30,31,32]) or TGCT markers (NANOG and POU5F1 [33,34,35,36,37])

Summary

Introduction

Aberrant overexpression of PIWI/piRNA pathway proteins is shown for many types of tumors. Testicular germ cell tumors (TGCTs) are the most frequent malignancies among male adolescents and young adults, with rising incidence [1, 2] They are very heterogeneous cancers that are histologically classified into seminomas and nonseminomas [3]. Ferreira et al found a correlation between hypermethylation of PIWI gene promoters and loss of their expression in TGCTs compared to normal testis of healthy individuals [12]. This group demonstrated a concomitant hypomethylation of L1 retrotransposons in TGCTs [12]. Another study by Rounge et al presented deep sequencing data of small RNAs for a set of normal testis, GCNIS samples and TGCTs, and stated that the loss of piRNAs is a hallmark of TGCT samples [13]

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