Revealing RNA poly(A) tail dynamics during spermiogenesis and its function to support male fertility [dRNA-seq_mutants]

TENT5C is a terminal nucleotidyltransferase with mRNA polyadenylation activity, required for the fastening of the flagella and the head of the sperm. However, TENT5C's role in shaping the poly(A) tail of the elongating spermatid transcriptome remains limited. Here we show that the poly(A) polymerase activity of TENT5C is required for fertility and spermiogenesis: male mice expressing catalytically inactive TENT5C (Tent5cdCat/dCat) are sterile and produce headless spermatozoa phenocopying TENT5C-deficient animals. We found that the transcript encoding for the structural proteins SMCP and ODF1 are directly targeted by TENT5C. We propose a model in which TENT5C secures the fastening of the flagella to the head of the sperm by targeting ODF1 in elongating spermatids. Overall design: Round (RS) and elongated (ES) spermatids were isolated from adult mice testis using Hoescht/PI staining and FACS sorting. Cells were isolated from wt/wt male mice or mice expressing a catalytically inactive TENT5C (Tent5cdcat/dcat). Each replicate is representative of a pool of 3 individual mice - 3 replicates per condition were considered for analysis. Nanopore technologies were used to sequence the transcriptome of round and elongated spermatids. Data were processed to measure the poly(A) tail length and the presence of 3'-end addition to mRNAs.