CT55 variants impair GABARAP-dependent autophagosome maturation to cause asthenoteratozoospermia in humans

Abstract

Asthenoteratozoospermia (AST) is a major cause of male infertility, yet the molecular connection between autophagy and sperm structural remodeling remains incompletely defined. Here, we identify two novel hemizygous cancer-testis antigen 55 (CT55) variants in unrelated men with severe AST: a multi-exonic truncation (exon1_4del; p.Met1_Glu180del) and a missense mutation (c.433C > T; p.Pro145Ser). Using integrated ultrastructural, immunofluorescence, and biochemical analyses, we show that these variants are associated with the altered CT55-mediated regulation of GABARAP-dependent autophagosome maturation through potentially distinct mechanisms. The truncating variant produces a shortened CT55 isoform that retains measurable GABARAP interaction but may impair downstream autophagosome maturation, whereas the missense variant reduces CT55 expression and weakens the CT55–GABARAP interaction in cell-based assays. Patient spermatozoa exhibit p62 accumulation, elevated LC3-II/LC3-I ratios, and broad cytoplasmic remodeling defects, including acrosomal malformation, midpiece disorganization, and axonemal instability. These findings support CT55 dysfunction as a cause of human AST and suggest that the disrupted CT55-associated autophagic regulation may contribute to multi-compartment sperm abnormalities. Our work expands the genetic and mechanistic framework of CT55-related male infertility and provides insights for molecular diagnosis and genetic counseling.