AUTHOR=Anbazhagan Rajakumar , Kavarthapu Raghuveer , Coon Steven L. , Dufau Maria L. 

TITLE=Role of Phosphorylated Gonadotropin-Regulated Testicular RNA Helicase (GRTH/DDX25) in the Regulation of Germ Cell Specific mRNAs in Chromatoid Bodies During Spermatogenesis

JOURNAL=Frontiers in Cell and Developmental Biology

VOLUME=8

YEAR=2020

URL=https://www.frontiersin.org/journals/cell-and-developmental-biology/articles/10.3389/fcell.2020.580019

DOI=10.3389/fcell.2020.580019

ISSN=2296-634X

ABSTRACT=<p>GRTH/DDX25 is a member of the DEAD-box family of RNA helicases that play an essential role in spermatogenesis. GRTH knock-in (KI) mice with the human mutant GRTH gene (R242H) show loss of the phospho-species from cytoplasm with preservation of the non-phospho form in the cytoplasm and nucleus. GRTH KI mice are sterile and lack elongated spermatids and spermatozoa, with spermatogenic arrest at step 8 of round spermatids which contain chromatoid body (CB) markedly reduced in size. We observed an absence of phospho-GRTH in CB of GRTH KI mice. RNA-Seq analysis of mRNA isolated from CB revealed that 1,421 genes show differential abundance, of which 947 genes showed a decrease in abundance and 474 genes showed an increase in abundance in GRTH KI mice. The transcripts related to spermatid development, differentiation, and chromatin remodeling (<italic>Tnp1/2</italic>, <italic>Prm1/2/3</italic>, <italic>Spem1/2</italic>, <italic>Tssk 2/3/6</italic>, <italic>Grth</italic>, <italic>tAce</italic>, and <italic>Upf2</italic>) were reduced, and the transcripts encoding for factors involved in RNA transport, regulation, and surveillance and transcriptional and translational regulation (<italic>Eef1a1, Ppp1cc</italic>, <italic>Pabpc1</italic>, <italic>Ybx3</italic>, <italic>Tent5b</italic>, <italic>H2al1m</italic>, <italic>Dctn2</italic>, and <italic>Dync1h1</italic>) were increased in the CB of KI mice and were further validated by qPCR. In the round spermatids of wild-type mice, mRNAs of <italic>Tnp2</italic>, <italic>Prm2</italic>, and <italic>Grth</italic> were abundantly co-localized with MVH protein in the CB, while in GRTH KI mice these were minimally present. In addition, GRTH binding to <italic>Tnp1/2</italic>, <italic>Prm1/2</italic>, <italic>Grth</italic>, and <italic>Tssk6</italic> mRNAs was found to be markedly decreased in KI. These results demonstrate the importance of phospho-GRTH in the maintenance of the structure of CB and its role in the storage and stability of germ cell-specific mRNAs during spermiogenesis.</p>