Meiotic Messenger RNA and Noncoding RNA Targets of the RNA-Binding Protein Translin (TSN) in Mouse Testis1

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Abstract
In postmeiotic male germ cells, TSN, formerly known as testis brain-RNA binding protein, is found in the cytoplasm and functions as a posttranscriptional regulator of a group of genes transcribed by the transcription factor CREM-tau. In contrast, in pachytene spermatocytes, TSN is found predominantly in nuclei. Tsn-null males show a reduced sperm count and high levels of apoptosis in meiotic cells, suggesting a critical function for TSN during meiosis. To identify meiotic target RNAs that associate in vivo with TSN, we reversibly cross-linked TSN to RNA in testis extracts from 17-day-old and adult mice and immunoprecipitated the complexes with an affinity-purified TSN antibody. Extracts from Tsn-null mice were used as controls. Cloning and sequencing the immunoprecipitated RNAs, we identified four new TSN target mRNAs, encoding diazepam-binding inhibitor-like 5, arylsulfatase A, a tetratricopeptide repeat structure-containing protein, and ring finger protein 139. In contrast to the population of postmeiotic translationally delayed mRNAs that bind TSN, these four mRNAs are initially expressed in pachytene spermatocytes. In addition, anti-TSN also precipitated a nonprotein-coding RNA (ncRNA), which is abundant in nuclei of pachytene spermatocytes and has a putative polyadenylation signal, but no open reading frame. A second similar ncRNA is adjacent to a GGA repeat, a motif frequently associated with recombination hot spots. RNA gel-shift assays confirm that the four new target mRNAs and the ncRNA specifically bind to TSN in testis extracts. These studies have, for the first time, identified both mRNAs and a ncRNA as TSN targets expressed during meiosis.

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